Cell therapy, and phenotype marker for related method

Abstract

To provide a cell therapy associated with administration of one more doses of T-cell composition for treatment, and a method, a composition and a product to be used in the therapy.SOLUTION: Cells of a T-cell composition expresses a recombinant receptor which is a chimeric receptor such as a chimeric antigen receptor (CAR) for example, or another transgenic receptor such as a T-cell receptor (TCR) for example. Features of aspects of the disclosure including a dose of cells to be administered or a unit of cells, and / or a phenotype of cells to be administered give various advantages which are consistent dosage, reduction of toxicity risk, and / or increase of response in a subject to which the T-cell composition is administered for example.SELECTED DRAWING: Figure 3C

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority from U.S. Provisional Application No. 62 / 596,775, entitled "PHENOTYPIC MARKERS FOR CELL THERAPY AND RELATED METHODS," filed December 8, 2017; U.S. Provisional Application No. 62 / 643,165, entitled "PHENOTYPIC MARKERS FOR CELL THERAPY AND RELATED METHODS," filed March 14, 2018; U.S. Provisional Application No. 62 / 657,716, entitled "PHENOTYPIC MARKERS FOR CELL THERAPY AND RELATED METHODS," filed April 13, 2018; and U.S. Provisional Application No. 62 / 716,967, entitled "PHENOTYPIC MARKERS FOR CELL THERAPY AND RELATED METHODS," filed August 9, 2018, the contents of which are incorporated by reference in their entireties.

[0002] INCORPORATION-BY-REFERENCE TO SEQUENCE LISTING This application is filed with an electronic Sequence Listing, which is provided in a file named 735042014540SeqList.txt, created on December 7, 2018, and is 35,461 bytes in size. The information in the electronic Sequence Listing is incorporated by reference in its entirety.

[0003] Field The present disclosure relates to cell therapy methods involving the administration of one or more doses of a therapeutic T cell composition, as well as methods, compositions, and articles of manufacture for use therein. The cells of the T cell composition express a recombinant receptor, such as a chimeric receptor, e.g., a chimeric antigen receptor (CAR), or other transgenic receptor, such as a T cell receptor (TCR). Features of embodiments of the present disclosure, including the dose or unit of cells administered and / or the phenotype of the administered cells, provide various advantages, such as consistent dosing, lower risk of toxicity, and / or increased response in subjects administered the T cell composition. [Background technology]

[0004] background A variety of immunotherapy and / or cell therapy methods are available for treating disease and pathology.For example, adoptive cell therapy (including the administration of cells that express chimeric receptors specific to the disease or disorder of interest, such as chimeric antigen receptors (CAR) and / or other recombinant antigen receptors, as well as other adoptive immune cell and adoptive T cell therapies) can be beneficial in the treatment of cancer or other diseases or disorders.There is a need for improved approaches.Methods and products that meet this need are provided. Summary of the Invention

[0005] overview A therapeutic composition comprising T cells expressing a recombinant receptor, wherein at least 40% or at least about 40%, or 40% or about 40%, or at least 50% or at least about 50%, or 50% or about 50%, or at least 60% or at least about 60%, or 60% or about 60%, or at least 70% or at least about 70%, or 70% or about 70%, or at least 80% or at least about 80%, or 80% or about 80%, or at least 85% or at least about 85%, or 85% or about 85%, or at least 90% or at least about 90%, or 90% or about 90%, or at least 91% or at least about 91%, or 91% or about 91%, or at least 92% of the total number of T cells in the composition or of the total number of T cells in the composition expressing the recombinant receptor. or at least about 92%, or 92% or about 92%, or at least 93% or at least about 93%, or 93% or about 93%, or at least 94% or at least about 94%, or 94% or about 94%, or at least 95% or at least about 95%, or 95% or about 95%, or at least 96% or at least about 96%, or 96% or about 96%, or at least 97% or at least about 97%, or 97% or about 97%, or at least 98% or at least about 98%, or 98% or about 98%, or at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100% are surface positive for CCR7 and / or CD27. In some embodiments, at least 50% or at least about 50%, or 50% or about 50% of the total number of T cells in the composition or of the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27.In some embodiments, at least 60%, or at least about 60%, or 60% or about 60% of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27. In some embodiments, at least 70%, or at least about 70%, or 70% or about 70% of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27. In some embodiments, at least 80%, or at least about 80%, or 80% or about 80% of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27.

[0006] In some embodiments, the T cells and / or T cells expressing the recombinant receptor in the composition comprise or consist of CD8+ T cells; the T cells and / or T cells expressing the recombinant receptor in the composition comprise or consist of CD4+ T cells; the T cells and / or T cells expressing the recombinant receptor in the composition comprise or consist of CD8+ and CD4+ T cells; the T cells and / or T cells expressing the recombinant receptor in the composition are at least 30% or at least about 30%, at least 50% or at least about 50%, at least 95% or at least about 95%, at least 96% or at least about 96%, at least 97% or at least about 97%, at least 98% or at least about 98%, or at least 99% or at least about 99%, or at least 100% or at least about 100% CD8+ the T cells and / or T cells expressing the recombinant receptor in the composition comprise or consist of at least 30%, at least 50%, at least 95%, at least 96%, at least 97%, at least 97%, at least 98%, or at least 98%, or at least 99%, or at least 100% or at least 100% CD4+ T cells; and / or the T cells and / or T cells expressing the recombinant receptor in the composition comprise or consist of about 1:1, or about 1:3-3:1, or about 1:2-2:1 CD4+:CD8+ T cells.

[0007] CD8 + T cell subtypes or CD4 + Defined numbers of T cell subtypes or those cells expressing recombinant receptors, such as CD8 + and CD4 +Provided herein are methods, compositions, and articles of manufacture involving or comprising a unit dose of cells based on the ratio of T cell subtypes. In some embodiments, the total number of T cells in the composition, or the total number of T cells in the composition expressing a recombinant receptor, are surface-positive for CCR7. In some embodiments, the total number of T cells in the composition, or the total number of T cells in the composition expressing a recombinant receptor, are surface-positive for CD27. In some embodiments, the total number of T cells in the composition, or the total number of T cells in the composition expressing a recombinant receptor, are surface-positive for CCR7 and CD27. In some embodiments, the composition comprises one or more unit doses of cells.

[0008] In some embodiments, the cell subtype is positive for a cell surface marker, such as a marker for a central memory phenotype, and / or the cell surface marker is CCR7 and / or CD27. In some embodiments, the provided methods, compositions and articles of manufacture are for use in connection with cell therapy, such as engineered T cell therapy, for the treatment of various diseases and conditions, including tumors.

[0009] Articles of manufacture are provided herein. In some aspects, the provided articles of manufacture comprise a plurality of CD8 receptor-expressing (optionally engineered to express) a recombinant CD8 receptor. + and / or CD4 + One or more unit doses of cells present in one or more therapeutic compositions, including T cells, expressing (optionally surface-expressing) a defined number of recombinant receptor-expressing CD8 T cells that express (optionally surface-expressing) CC chemokine receptor type 7 (CCR7). + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor+ / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + The container includes a unit dose of cells, including a subset of cells in a cell-pair composition, and instructions for administering the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, to a subject having a disease or condition.

[0010] Articles of manufacture are provided herein. In some aspects, the provided articles of manufacture comprise a plurality of CD8 receptor-expressing (optionally engineered to express) a recombinant CD8 receptor. + and / or CD4 + One or more unit doses of cells present in one or more therapeutic compositions, including T cells, expressing (optionally surface-expressing) a defined number of recombinant receptor-expressing CD8 T cells that express (optionally surface-expressing) Cluster of Differentiation 27 (CD27). + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 +The container includes a unit dose of cells, including a subset of cells in a cell-pair composition, and instructions for administering the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, to a subject having a disease or condition.

[0011] Also provided herein are therapeutic compositions. In some embodiments, the therapeutic composition comprises T cells expressing a recombinant receptor, wherein at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least 94% or less of the total number of T cells in the composition (or of the total number of T cells in the composition expressing the recombinant receptor). at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or 95% or about 95%, at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100% are surface positive for CCR7 and / or CD27, and optionally the T cells and / or T cells expressing the recombinant receptor in the composition are surface positive for CCR7 and / or CD27, + comprising or consisting of T cells; the T cells and / or T cells expressing the recombinant receptor in the composition are CD4 + comprising or consisting of T cells; the T cells and / or T cells expressing the recombinant receptor in the composition are CD8 + and CD4 +comprising or consisting of T cells; the T cells and / or T cells expressing the recombinant receptor in the composition are at least 30% or at least about 30%, at least 50% or at least about 50%, at least 95% or at least about 95%, at least 96% or at least about 96%, at least 97% or at least about 97%, at least 98% or at least about 98%, or at least 99% or at least about 99%, or at least 100% or at least about 100% CD8 + or at least about 100% of the CD4 T cells in the composition; ... + the T cells and / or T cells expressing the recombinant receptor in the composition are in a ratio of about 1:1, or about 1:3 to 3:1, or about 1:2 to 2:1 CD4 + :CD8 + Comprising or consisting of T cells.

[0012] In some aspects, the unit dose of cells comprises any of the therapeutic compositions described herein.

[0013] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + and / or the unit dose of cells contains a defined number of CD4 + / CCR7 + In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CD27 + and / or the unit dose of cells contains a defined number of CD4 + / CD27 +In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + / CD27 + and / or the unit dose of cells contains a defined number of CD4 + / CCR7 + / CD27 + Contains cells.

[0014] In some embodiments, the unit dose or composition is 1×10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + The unit dose or composition may contain about 1 x 10 cells and / or 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 105 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells.

[0015] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the unit dose is 3×10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the total receptor in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptors in the unit dose +At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + is.

[0016] In some embodiments, the unit dose or composition of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0017] In some embodiments, the unit dose or composition of cells comprises a defined ratio of receptors. + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + The cells include 1:1 or about 1:1 or about 1:3 to about 3:1 CCR7 and / or CD45RA cells, optionally at a ratio of 1:1 or about 1:1 or about 1:3 to about 3:1. In some embodiments, the specified number or ratio is further based on the expression or absence of CCR7 and / or CD45RA on the cells. In some embodiments, the specified number of cells further express or do not express CCR7 and / or CD45RA, and optionally, the specified number of cells further express or do not express CCR7 and / or CD45RA. + or CD45RA - It is a cell.

[0018] In some embodiments, the defined number or ratio is further based on the expression or absence of CD27 and / or CD45RA on the cells. In some embodiments, the defined number of cells further express or do not express CD27 and / or CD45RA, and optionally, the defined number of cells further express CD27 + In some embodiments, the defined number further comprises cells that do or do not express CD27 and / or CD45RA, and optionally, the defined number further comprises cells that do or do not express CD27 and / or CD45RA. + or CD45RA - In some embodiments, the unit dose or composition of cells contains a defined number of receptors. + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD8 + / CCR7 + / CD45RA - In some embodiments, the unit dose or composition of cells contains a defined number of receptors. + / CD4 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - Contains cells.

[0019] In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA- , receptor + / CD4 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD45RA - is.

[0020] Also, multiple CD8 receptors, optionally engineered to express recombinant receptors, + and / or CD4 + One or more unit doses of cells present in one or more therapeutic compositions, including T cells, and a defined number of recombinant receptor-expressing CD8 T cells expressing CCR7 and CD27. + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 +Provided herein are articles of manufacture that include a container containing a unit dose of cells, including a subset of cells in a cell-pair composition, and instructions for administering the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, to a subject having a disease or condition. In some embodiments, the unit dose of cells is a defined number of CD8 + / CCR7 + / CD27 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 1 x 10 cells, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 105 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0021] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + is.

[0022] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 +cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0023] In some embodiments, the defined number or ratio is further based on the expression or absence of CD45RA on the cells. In some embodiments, the defined number of cells further express or do not express CD45RA, and optionally, the defined number of cells further express CD45RA. - In some embodiments, the defined number further includes cells that do or do not express CD45RA, and optionally, the defined number further includes cells that do or do not express CD45RA. - Contains cells.

[0024] In some embodiments, among the plurality of articles or unit doses or compositions, optionally produced according to the same method, there are provided cells expressing CCR7 (CCR7 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0025] In some embodiments, the unit dose or number of cells expressing the recombinant receptor is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD3 expressing the recombinant receptor between + Cells (receptors + / CD3 + cells) or total CD3 +In some embodiments, the unit dose or number of cells expressing the recombinant receptor is about 5 x 10 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + Contains cells.

[0026] In some embodiments, CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA -The total number of cells is the total number of such cells that are alive or viable. + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that do not express the apoptotic marker and / or are apoptotic marker negative ( - ) is the total number of such cells, where the apoptotic marker is annexin V or activated caspase 3.

[0027] In some embodiments, the instructions specify administering a plurality of unit doses contained in a plurality of separate compositions. In some embodiments, the plurality of separate compositions comprises a CD8 + T cells and CD4 +a first composition comprising one of T cells and CD8 + T cells and CD4 + In some embodiments, the first composition comprises a second composition comprising another of the CD8 T cells. + In some embodiments, the first composition comprises CD4 T cells. + Contains T cells.

[0028] In some embodiments, the instructions include instructions for use in a CD8 + a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, and + It is specified that the T cell-containing composition or one or more unit doses thereof and / or an amount corresponding to one or more such unit doses are administered within 2 hours, 1 hour, 30 minutes, 15 minutes, 10 minutes or 5 minutes apart or simultaneously.

[0029] In some embodiments, the instructions include instructions for use to + Prior to administering a composition containing CD8 cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, + In some embodiments, the instructions specify administering a composition containing CD8 T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses. + CD4 cells are administered prior to administration of one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses. + It is specified that a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more such unit doses is administered.

[0030] In some embodiments, the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor. In some embodiments, the recombinant receptor is capable of binding to a target antigen associated with, specific to, and / or expressed on a cell or tissue of a disease, disorder, or condition. In some embodiments, the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer. In some embodiments, the target antigen is a tumor antigen.

[0031] In some embodiments, the target antigen is αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (also known as CA9, CAIX, or G250), cancer testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, CD138, CD171, epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutant (EGFR), or EGFR mutated type III (EGFR mutated type III). vIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5;Fc receptor homolog 5 or FCRH5), fetal acetylcholine receptor (fetal AchR), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (gp100), G protein-coupled receptor 5D (GPCR5D), Her2 / neu (receptor tyrosine kinase erbB2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high-molecular-weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-AI), human leukocyte antigen A2 (HLA-A2), IL-22 receptor Interleukin-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), kinase insert domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, mesothelin, c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen (oncofetal antigen), melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific antigens, or antigens associated with universal tags, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV or other pathogens;

[0032] In some embodiments, the recombinant receptor is or comprises a functional non-TCR antigen receptor or TCR or an antigen-binding fragment thereof, hi some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR).

[0033] In some embodiments, the recombinant receptor comprises an extracellular domain comprising an antigen-binding domain. In some embodiments, the antigen-binding domain is or comprises an antibody or antibody fragment (optionally a single-chain fragment) thereof. In some embodiments, the fragment comprises antibody variable regions linked by a flexible linker. In some embodiments, the fragment comprises an scFv.

[0034] In some embodiments, the recombinant receptor comprises an intracellular signaling region. In some embodiments, the intracellular signaling region comprises an intracellular signaling domain. In some embodiments, the intracellular signaling domain is or comprises a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain comprising an immunoreceptor tyrosine-based activation motif (ITAM). In some embodiments, the intracellular signaling domain is or comprises the intracellular signaling domain of a CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof.

[0035] In some embodiments, the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region. In some embodiments, the intracellular signaling region further comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

[0036] In some embodiments, the T cells are primary T cells obtained from the subject. In some embodiments, the T cells are autologous to the subject. In some embodiments, the T cells are allogeneic to the subject.

[0037] Also provided herein are methods of treatment. In some embodiments, the methods of treatment involve administering to a subject having a disease or condition any of the therapeutic compositions described herein or a plurality of CD8 + and / or CD4 + The method involves administering one or more unit doses of a therapeutic composition comprising T cells and / or an amount corresponding to such a unit dose. In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 T cells expressing CC chemokine receptor type 7 (CCR7). + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + and / or a group of subjects treated according to the method, comprising a subset of cells expressing CCR7 (CCR7 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0038] Also provided herein are methods of treatment. In some embodiments, the methods of treatment involve administering to a subject having a disease or condition a therapeutic composition described herein or a plurality of CD8 + and / or CD4 + The method involves administering one or more unit doses of a therapeutic composition comprising T cells and / or an amount corresponding to such a unit dose, and optionally, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 T cells expressing cluster of differentiation 27 (CD27). + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 + and / or a group of subjects treated according to the method, including a subset of cells expressing CD27 (CD27 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0039] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CD27 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CD27 + Contains cells.

[0040] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + In some embodiments, the unit dose comprises 1 x 10 cells, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 +Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells.

[0041] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the unit dose is 3×10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the total receptor in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose +Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + is.

[0042] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0043] In some embodiments, the defined number or ratio is further based on the expression or absence of expression on the cells of CD27 and / or CD45RA. + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD8 + / CCR7 + / CD45RA - In some embodiments, a unit dose of cells contains a defined number of receptors. + / CD4 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 +The ratio of CCR7 and / or CD45RA to the total receptors in the unit dose is optionally 1:1 or about 1:1, or about 1:3 to about 3:1. In some embodiments, the specified number or ratio is further based on the expression or absence of CCR7 and / or CD45RA on the cells. In some embodiments, the total receptors in the unit dose are + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD8 + / CCR7 + / CD45RA - is.

[0044] Also provided in some embodiments are methods of treatment involving administering to a subject having a disease or condition one or more unit doses of any of the compositions described herein.

[0045] In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 +Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + and a group of subjects treated according to the method, comprising a subset of cells expressing CCR7 (CCR7 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0046] Also, multiple CD8 receptors engineered to express recombinant receptors in subjects with a disease or condition are provided. + and / or CD4 + Provided herein are methods of treatment that involve administering a therapeutic composition comprising T cells and / or an amount corresponding to such a unit dose. In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 T cells that express CCR7 and CD27. + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors+ / CD4 + / CCR7 + / CD27 + and a group of subjects treated according to the method, comprising a subset of cells expressing CCR7 (CCR7 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0047] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + / CD27 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 1 x 10 cells, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0048] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells; and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 106 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells; and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + is.

[0049] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor+ / CD4 + / CCR7 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0050] In some embodiments, the defined number or ratio is further based on the expression or absence of expression on the cells of CD45RA. - In some embodiments, the defined number of cells further express or do not express CD45RA, and optionally, the defined number of cells further express or do not express CD45RA. - It is a cell.

[0051] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD3 expressing the recombinant receptor between + Cells (receptors + / CD3 + cells) or total CD3 + In some embodiments, the unit dose comprises about 5 x 10 cells. 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5Total receptors below + / CD3 + Cellular or total CD3 + Contains cells.

[0052] In some embodiments, CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that are alive or viable. + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that do not express the apoptotic marker and / or are apoptotic marker negative ( - ) is the total number of such cells, where the apoptotic marker is annexin V or activated caspase 3.

[0053] In some embodiments, the methods involve administering multiple unit doses contained in multiple separate compositions.

[0054] In some embodiments, the plurality of separate compositions comprises CD8 + T cells and CD4 + a first composition comprising one of T cells and CD8 + T cells and CD4 + In some embodiments, the first composition comprises a second composition comprising another of the CD8 T cells. + In some embodiments, the first composition comprises CD4 T cells. + Contains T cells.

[0055] In some embodiments, the method comprises: + a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, and + It involves administering a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more such unit doses within 2 hours, 1 hour, 30 minutes, 15 minutes, 10 minutes or 5 minutes apart or simultaneously.

[0056] In some embodiments, the method comprises: + Prior to administering a composition containing CD8 cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, + It involves administering a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses.

[0057] In some embodiments, the method comprises: + CD4 cells are administered prior to administration of one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses. + It involves administering a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses.

[0058] In some embodiments, the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor. In some embodiments, the recombinant receptor is capable of binding to a target antigen associated with, specific to, and / or expressed on a cell or tissue of a disease, disorder, or condition. In some embodiments, the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer. In some embodiments, the target antigen is a tumor antigen.

[0059] In some embodiments, the target antigen is αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (also known as CA9, CAIX, or G250), cancer testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, CD123, CD138, CD171, epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutant (EGFR), or EGFR mutated type III (EGFR mutated type III). vIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5;Fc receptor homolog 5 or also known as FCRH5), fetal acetylcholine receptor (fetal AchR), folate binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (gp100), G protein-coupled receptor 5D (GPRC5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high High molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), kinase insert domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAA) MAGE)-A1, MAGE-A3, MAGE-A6, mesothelin, c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, preferentially expressed antigen in melanoma (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like antigen (RTK). The antigen is selected from among Fan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific antigens or antigens associated with universal tags, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV or other pathogens;

[0060] In some embodiments, the recombinant receptor is or comprises a functional non-TCR antigen receptor or TCR or an antigen-binding fragment thereof, hi some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR).

[0061] In some embodiments, the recombinant receptor comprises an extracellular domain comprising an antigen-binding domain. In some embodiments, the antigen-binding domain is or comprises an antibody or antibody fragment (optionally a single-chain fragment) thereof. In some embodiments, the fragment comprises antibody variable regions linked by a flexible linker. In some embodiments, the fragment comprises an scFv.

[0062] In some embodiments, the recombinant receptor comprises an intracellular signaling region. In some embodiments, the intracellular signaling region comprises an intracellular signaling domain. In some embodiments, the intracellular signaling domain is or comprises a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain comprising an immunoreceptor tyrosine-based activation motif (ITAM). In some embodiments, the intracellular signaling domain is or comprises the intracellular signaling domain of a CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof.

[0063] In some embodiments, the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region. In some embodiments, the intracellular signaling region further comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof.

[0064] In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain or a signaling portion thereof of CD28, 4-1BB, or ICOS, hi some embodiments, the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

[0065] In some embodiments, the T cells are primary T cells obtained from the subject. In some embodiments, the T cells are autologous to the subject. In some embodiments, the T cells are allogeneic to the subject.

[0066] Also provided herein are methods for determining a dosage unit of engineered T cells for treating a subject. In some embodiments, the methods involve determining a dosage unit of engineered T cells that express a recombinant receptor. + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and CCR7 are expressed (receptor + / CCR7 + In some embodiments, the method includes assessing the number, percentage, or ratio of receptor T cells. + / CCR7 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 C chemokine receptor type 7 (CCR7)-expressing cells; + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios+ / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + The method includes the step of including a subset of cells in a cell pair composition.

[0067] Also provided herein are methods for determining a dosage unit of engineered T cells for treating a subject. In some embodiments, the methods include: (a) determining a plurality of CD8 T cells engineered to express a recombinant receptor; + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and CD27 are expressed (receptor + / CD27 + (b) assessing the number, percentage, or ratio of T cells; and + / CD27 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 + The method involves including a subset of cells in a cell-pair composition.

[0068] Also provided herein are methods for producing a composition comprising a unit dose of a T cell composition. In some embodiments, the methods include producing a plurality of CD8 T cells engineered to express a recombinant receptor. + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and those expressing CCR7 and CD27 (receptors + / CCR7 + / CD27 + In some embodiments, the method includes filling all or a portion of the composition, optionally in a separate solution, into a container to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express CC chemokine receptor type 7 (CCR7). + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + In some embodiments, the method includes filling all or a portion of the composition, optionally a separate solution, into a container to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express Cluster of Differentiation 27 (CD27). + T cells (receptors + / CD8+ / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 + The method includes the step of including a subset of cells in a cell pair composition.

[0069] Also provided herein are methods for producing a therapeutic composition comprising a unit dose of a T cell composition. In some embodiments, the method includes filling a container with all or a portion of a T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express CC chemokine receptor type 7 (CCR7). + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7+ Cells and / or receptors + / CD4 + / CCR7 + The method includes the step of including a subset of cells in a cell pair composition.

[0070] Also provided herein are methods for producing a therapeutic composition comprising a unit dose of a T cell composition. In some embodiments, the method includes filling a container with all or a portion of a T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express Cluster of Differentiation 27 (CD27). + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 + The method includes the step of including a subset of cells in a cell pair composition.

[0071] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + Contains cells.

[0072] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8+ / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells.

[0073] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the unit dose is 3×10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8+ / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + In some embodiments, the total receptor in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 +is.

[0074] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0075] In some embodiments, the unit dose further comprises a defined number of cells, including cells that may or may not express CD27 and / or CD45, and optionally, the unit dose further comprises a defined number of cells that may or may not express CD27 and / or CD45. + or a defined number of cells including CD45RA- cells.

[0076] In some embodiments, the method further comprises measuring the number, percentage, or ratio of T cells that express or do not express CD27 and / or CD45RA in the therapeutic composition, optionally including CD27 + Or, assessing the number, percentage, or ratio of T cells that are CD45RA-.

[0077] In some embodiments, a unit dose of cells contains a defined number of receptors. + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD8 + / CCR7 + / CD45RA - In some embodiments, a unit dose of cells contains a defined number of receptors. + / CD4 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - Contains cells.

[0078] In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - , receptor + / CD4 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD45RA - is.

[0079] Also provided herein are methods for determining a unit dose of engineered T cells for treating a subject. In some embodiments, the methods involve determining a unit dose of engineered T cells engineered to express a recombinant receptor, such as CD8 + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and those expressing CCR7 and CD27 (receptors + / CCR7 + / CD27 + In some embodiments, the method includes assessing the number, percentage, or ratio of receptor T cells. + / CCR7 + / CD27 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CCR7 + / CD27+ cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + The method includes the step of including a subset of cells in a cell pair composition.

[0080] Also provided herein are methods for producing a composition comprising a unit dose of a T cell composition. In some embodiments, the methods include producing a plurality of CD8 T cells engineered to express a recombinant receptor. + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and those expressing CCR7 and CD27 (receptors + / CCR7 + / CD27 + In some embodiments, the method includes filling all or a portion of the composition, optionally in a separate solution, into a container to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express CCR7 and CD27. + T cells (receptors + / CD8 + / CCR7 + / CD27 +cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + The method includes the step of including a subset of cells in a cell pair composition.

[0081] Also provided herein are methods for producing a therapeutic composition comprising a unit dose of a T cell composition. In some embodiments, the method includes filling a container with all or a portion of a T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition to achieve a unit dose of the T cell composition, wherein the unit dose contains a defined number of recombinant receptor-expressing CD8 T cells that express CCR7 and CD27. + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27+ Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + The method includes the step of including a subset of cells in a cell pair composition.

[0082] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + / CD27 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + / CD27 + Contains cells.

[0083] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27+ Cells, or total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0084] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + / CD27 +In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27+ is.

[0085] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0086] In some embodiments, the unit dose further comprises a defined number of cells, including cells that express or do not express CD45, and optionally, the unit dose further comprises a defined number of cells, including CD45RA- cells. In some embodiments, the method further comprises assessing the number, percentage, or ratio of T cells in the therapeutic composition that express or do not express CD45RA, and optionally the number, percentage, or ratio of T cells that are CD45RA-.

[0087] Also provided are methods for producing cell compositions comprising genetically engineered cells. In some embodiments, the methods involve selecting a target percentage of CD8+ cells that express or do not express CCR7, CD27, and / or CD45RA from a biological sample from a subject. + Target percentages of T cells and / or CD4 +In some embodiments, the method involves providing an input composition comprising T cells. In some embodiments, the method involves introducing a polynucleotide encoding a recombinant receptor into the input composition. In some embodiments, the method involves stimulating the cells in the input composition before, during, and / or after the introducing step, the stimulation step comprising incubating the cells in the presence of one or more stimulatory agents, which results in activation and / or proliferation of the cells. In some embodiments, the method involves introducing recombinant receptor-expressing T cells (receptors) that do or do not express a defined ratio of CCR7, CD27, and / or CD45RA. + )CD8 + T cells versus receptors expressing or not expressing CCR7, CD27 and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing defined ratios of CCR7, CD27 and / or CD45RA + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27 and / or CD45RA + / CD4 + An output composition is produced that contains the distinct cell subsets in the T cell pair composition.

[0088] Also provided are methods for producing cell compositions comprising genetically engineered cells. In some embodiments, the methods include introducing a polynucleotide encoding a recombinant receptor into an input composition, wherein the input composition is provided from a subject-derived biological sample containing a target percentage of CD8+ cells that express or do not express CCR7, CD27, and / or CD45RA. + Target percentages of T cells and / or CD4 +and the input composition comprises T cells, and the input composition comprises a step in which, prior to, during and / or after said introducing step, the cells are stimulated by incubating the cells in the presence of one or more stimulatory substances, said stimulation resulting in activation and / or proliferation of the cells; and the method comprises introducing recombinant receptor-expressing T cells (receptors) that express or do not express a defined ratio of CCR7, CD27 and / or CD45RA. + )CD8 + T cells versus receptors expressing or not expressing CCR7, CD27 and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing defined ratios of CCR7, CD27 and / or CD45RA + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27 and / or CD45RA + / CD4 + An output composition is produced that contains the distinct cell subsets in the T cell pair composition.

[0089] Also provided are methods for producing cell compositions comprising genetically engineered cells. In some embodiments, the methods involve isolating CD8+ cells that express or do not express target numbers of CCR7, CD27, and / or CD45RA from a biological sample obtained from a subject. + T cells and / or CD4 cells with or without CCR7, CD27, and / or CD45RA expression in target numbers +The method involves isolating T cells, thereby creating an input composition. In some embodiments, the method involves introducing a polynucleotide encoding a recombinant receptor into the input composition. In some embodiments, the method involves stimulating the cells in the input composition before, during, and / or after the introducing step, the stimulation step comprising incubating the cells in the presence of one or more stimulatory agents, which results in cell activation and / or proliferation. In some embodiments, the method involves isolating T cells, thereby creating an input composition ... which express or do not express a defined ratio of CCR7, CD27, and / or CD45RA, + )CD8 + T cells versus receptors expressing or not expressing CCR7, CD27 and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing defined ratios of CCR7, CD27 and / or CD45RA + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27 and / or CD45RA + / CD4 + An output composition is produced that contains the distinct cell subsets in the T cell pair composition.

[0090] Also provided are methods for producing cell compositions comprising genetically engineered cells. In some embodiments, the methods include introducing a polynucleotide encoding a recombinant receptor into an input composition, wherein the input composition is selected from a biological sample obtained from a subject to identify CD8+ cells that express or do not express a target number of CCR7, CD27, and / or CD45RA. + T cells and / or CD4 cells with or without CCR7, CD27, and / or CD45RA expression in target numbers +The method is produced by isolating T cells, and the input composition involves a step in which, before, during and / or after the introducing step, the cells are stimulated by incubating the cells in the presence of one or more stimulatory substances, the stimulation resulting in activation and / or proliferation of the cells; and the method involves the introduction of recombinant receptor-expressing (receptor-expressing) T cells that express or do not express a defined ratio of CCR7, CD27 and / or CD45RA. + )CD8 + T cells versus receptors expressing or not expressing CCR7, CD27 and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing defined ratios of CCR7, CD27 and / or CD45RA + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27 and / or CD45RA + / CD4 + An output composition is produced that contains the distinct cell subsets in the T cell pair composition.

[0091] In some embodiments, the input composition comprises a target percentage of CD8+ cells that either express or do not express CCR7, CD27, and / or CD45RA. + Target percentages of T cells and / or CD4 + Contains T cells.

[0092] In some embodiments, the input composition is a target percentage of CD4 + / CCR7 + , CD4 + / CD27 + , CD4 + / CCR7 + / CD27 + , CD4 + / CCR7 + / CD45RA-, CD4 + / CCR7 + / CD45RA + , CD8 + / CCR7 + , CD8 + / CD27 + , CD8 + / CCR7 + / CD27 + , CD8 + / CCR7 + / CD45RA- and / or CD8 + / CCR7 + / CD45RA + Contains cells.

[0093] In some embodiments, the output composition comprises a defined ratio of receptors. + / CD8 + / CCR7 + , receptor + / CD8 + / CD27 + , receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA- and / or receptor + / CD8 + / CCR7 + / CD45RA + Cell vs. Receptor + / CD4 + / CCR7 + , receptor + / CD4 + / CD27 + , receptor + / CD4 + / CCR7 + / CD27 + , receptor + / CD4 + / CCR7 + / CD45RA- and / or receptor + / CD4 + / CCR7 + / CD45RA + cells; or receptors in a defined ratio + / CD8 + / CCR7 + , receptor + / CD8+ / CD27 + , receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA- and / or receptor + / CD8 + / CCR7 + / CD45RA + Cells and / or receptors + / CD4 + / CCR7 + , receptor + / CD4 + / CD27 + , receptor + / CD4 + / CCR7 + / CD27 + , receptor + / CD4 + / CCR7 + / CD45RA- and / or receptor + / CD4 + / CCR7 + / CD45RA + Cell pairs include subsets of different cells in a composition.

[0094] In some embodiments, the method also includes determining one or more unit doses of the T cell composition for administration to a subject having a disease or condition, the unit dose comprising a defined number of recombinant receptor-expressing CD8 T cells that express CC chemokine receptor type 7 (CCR7). + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + Another subset of cells in the cell-pair composition: a defined number of recombinant receptor-expressing CD8 that expresses cluster of differentiation 27 (CD27). + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 + Cells and / or receptors + / CD4 + / CD27 + a subset of cells in the cell pair composition; or a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27+ Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + The method involves including all or a portion of the output composition comprising a subset of distinct cells in the cell pair composition.

[0095] In some embodiments, the unit dose of cells comprises a defined number of CD8 + / CCR7 + cells, CD4 + / CCR7 + cells, CD8 + / CD27 + cells, CD4 + / CD27 + cells, CD8 + / CCR7 + / CD27 + cells and / or CD4 + / CCR7 + / CD27 + Contains cells.

[0096] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + Cell or total receptor+ / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, total receptors + / CD4 + / CD27 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0097] In some embodiments, the unit dose is at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells; and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 +and / or at least about 3 × 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells and / or at least about 3 x 10 6 , 4×10 6 , 5×10 6 , 6×10 6 , 7×10 6 , 8×10 6 , 9×10 6 Or 1 x 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0098] In some embodiments, the unit dose is 3×10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells; and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + , and / or 3×10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + Contains cells.

[0099] In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + and / or the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + and / or the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + is.

[0100] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + and / or a defined ratio of receptors, optionally in a ratio of 1:1 or about 1:1, or about 1:3 to about 3:1. + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + and / or a defined ratio of receptors, optionally in a ratio of 1:1 or about 1:1, or about 1:3 to about 3:1. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0101] In some embodiments, the unit dose further comprises a defined number of cells, including cells that express or do not express CD45, and optionally, the unit dose further comprises a defined number of cells, including CD45RA- cells. + / CD4 + / CCR7 + / CD45RA - Cells and / or receptors + / CD8 + / CCR7 + / CD45RA - Contains cells.

[0102] In some embodiments, the total receptors in the unit dose + At least 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7+ / CD45RA - or receptor + / CD8 + / CCR7 + / CD45RA - is.

[0103] In some embodiments, the method further comprises providing or isolating CD8 + T cells and / or CD4 with or without CCR7, CD27 and / or CD45RA expression + Determining the number, volumetric number, weight number, and / or percentage of T cells.

[0104] In some embodiments, the one or more stimulatory agents are selected from the group consisting of T cells, CD4 + T cells and / or CD8 + capable of activating T cells; capable of inducing signals through the TCR complex; and / or capable of activating T cells, CD4 + T cells and / or CD8 + It can induce T cell proliferation.

[0105] In some embodiments, the one or more stimulatory agents comprise a primary agent that binds to a member of the TCR complex, optionally specifically binds to CD3. In some embodiments, the one or more stimulatory agents further comprise a secondary agent that specifically binds to a T cell costimulatory molecule. In some embodiments, the costimulatory molecule is selected from the group consisting of CD28, CD137 (4-1-BB), OX40, or ICOS. In some embodiments, the primary and secondary agents comprise antibodies, and optionally, the one or more stimulatory agents include incubation with an anti-CD3 antibody and an anti-CD28 antibody. In some embodiments, the one or more stimulatory agents are present on a solid support, optionally a bead. In some embodiments, the one or more stimulatory agents are selected from the group consisting of a CD3-binding molecule; a CD28-binding molecule; recombinant IL-2; recombinant IL-15; and recombinant IL-7; a vaccine comprising an antigen specifically recognized by an antigen receptor; and an anti-idiotypic antibody that specifically binds to an antigen receptor, or a combination thereof.

[0106] In some aspects, the therapeutic composition is any of the compositions described herein.

[0107] In some embodiments, the method further comprises administering to a subject having the disease or condition a therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses.

[0108] In some embodiments, among a group of subjects treated according to the method, the number of cells expressing CCR7 (CCR7 + The number or ratio of cells varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

[0109] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD3 expressing the recombinant receptor between + Cells (receptors + / CD3 + cells) or total CD3 + In some embodiments, the unit dose comprises about 5 x 10 cells. 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + Contains cells.

[0110] In some embodiments, CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 +Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that are alive or viable. + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA -The total number of cells is the total number of such cells that do not express the apoptotic marker and / or are apoptotic marker negative ( - ) is the total number of such cells, where the apoptotic marker is annexin V or activated caspase 3.

[0111] In some embodiments, the methods involve administering multiple unit doses contained in multiple separate compositions.

[0112] In some embodiments, the plurality of separate compositions comprises CD8 + T cells and CD4 + a first composition comprising one of T cells and CD8 + T cells and CD4 + In some embodiments, the first composition comprises a second composition comprising another of the CD8 T cells. + In some embodiments, the first composition comprises CD4 T cells. + Contains T cells.

[0113] In some embodiments, the method comprises: + a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, and + It involves administering a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more such unit doses within 2 hours, 1 hour, 30 minutes, 15 minutes, 10 minutes or 5 minutes apart or simultaneously.

[0114] In some embodiments, the method comprises: + Prior to administering a composition containing CD8 cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, + In some embodiments, the method involves administering a composition containing CD8 T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses. +CD4 cells are administered prior to administration of one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses. + It involves administering a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses.

[0115] In some embodiments, the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor. In some embodiments, the recombinant receptor is capable of binding to a target antigen associated with, specific to, and / or expressed on a cell or tissue of a disease, disorder, or condition. In some embodiments, the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer. In some embodiments, the target antigen is a tumor antigen.

[0116] In some embodiments, the target antigen is αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (also known as CA9, CAIX, or G250), cancer testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, CD123, CD138, CD171, epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutant (EGFR), or EGFR mutated type III (EGFR mutated type III). vIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5;Fc receptor homolog 5 or also known as FCRH5), fetal acetylcholine receptor (fetal AchR), folate binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (gp100), G protein-coupled receptor 5D (GPRC5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high High molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), kinase insert domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAA) MAGE)-A1, MAGE-A3, MAGE-A6, mesothelin, c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, preferentially expressed antigen in melanoma (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like antigen (RTK). The antigen is selected from among Fan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific antigens or antigens associated with universal tags, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV or other pathogens;

[0117] In some embodiments, the recombinant receptor is or comprises a functional non-TCR antigen receptor or TCR, or an antigen-binding fragment thereof. In some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR). In some embodiments, the recombinant receptor comprises an extracellular domain comprising an antigen-binding domain. In some embodiments, the antigen-binding domain is or comprises an antibody or antibody fragment thereof (optionally a single-chain fragment). In some embodiments, the fragment comprises antibody variable regions linked by a flexible linker. In some embodiments, the fragment comprises an scFv.

[0118] In some embodiments, the recombinant receptor comprises an intracellular signaling region. In some embodiments, the intracellular signaling region comprises an intracellular signaling domain. In some embodiments, the intracellular signaling domain is or comprises a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain comprising an immunoreceptor tyrosine-based activation motif (ITAM). In some embodiments, the intracellular signaling domain is or comprises the intracellular signaling domain of a CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof.

[0119] In some embodiments, the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region.

[0120] In some embodiments, the intracellular signaling region further comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

[0121] In some embodiments, the T cells are primary T cells obtained from the subject. In some embodiments, the T cells are autologous to the subject. In some embodiments, the T cells are allogeneic to the subject.

[0122] In some aspects of any of the embodiments provided herein, the cells in the provided therapeutic compositions or unit doses are associated with certain functional attributes, including a particular recombinant receptor (e.g., CAR)-dependent activity that is contingent on the activity or presence of the recombinant receptor, e.g., an activity induced or mediated in the presence of an antigen recognized by the recombinant receptor. In some embodiments, the functional attribute includes higher levels of cytokine production, e.g., IL-2. In some aspects of any of the embodiments provided herein, the functional attribute includes lower levels of cytokine production, e.g., IFNγ or IL-13. In some embodiments, higher or lower levels of such cytokines are relative to a reference composition containing an undefined number of cells of a particular phenotype and / or a specific, undefined ratio of cells of a particular phenotype according to the embodiments provided herein. In some embodiments, the reference composition contains a greater frequency or number of T cells having an effector or differentiated phenotype.

[0123] In some aspects, when a therapeutic composition or unit dose is administered to one of a group of subjects according to any embodiment provided herein, multiple subjects in the group of subjects exhibit one or more characteristics associated with increased efficacy, durability, and / or reduced toxicity following administration. In some embodiments, multiple subjects in the group of subjects are likely to exhibit and / or are associated with pharmacokinetic parameters, reduced likelihood of toxicity or adverse events, and / or a particular response, efficacy, or survival outcome. In some embodiments, the subjects are subjects suspected of having a disease or condition. In some embodiments, the one or more characteristics are improved compared to a similar group of subjects treated with a reference composition containing an undefined number of cells of a particular phenotype and / or a particular, undefined ratio of cells of a particular phenotype according to embodiments provided herein. In some embodiments, the reference composition contains a greater frequency or number of T cells having an effector or differentiated phenotype.

[0124] In some aspects, upon administration of a therapeutic composition or unit dose to one of a group of subjects according to any embodiment provided herein, a plurality of subjects in the group of subjects experience a maximum (peak) plasma concentration (C max ), peak time (i.e., maximum plasma concentration (C max ) occurs;T max ), minimum plasma concentration (i.e., the therapeutic agent, e.g., CAR + Minimum plasma concentration between doses of T cells; C min ), elimination half-life (T 1 / 2 ) and area under the curve (i.e., therapeutic agent CAR + In some aspects, upon administration of a therapeutic composition or unit dose, multiple subjects in a group experience an increased maximum (peak) plasma concentration (C) of the administered cells. maxIn some embodiments, long-term persistence of the therapeutic composition or unit dose of cells is observed in multiple subjects in the group.

[0125] In some aspects, when a therapeutic composition or unit dose is administered to one of a group of subjects according to any embodiment provided herein, a plurality of subjects in the group of subjects are likely to exhibit and / or are associated with a reduced likelihood of occurrence of a toxicity or adverse event, such as cytokine release syndrome (CRS) or severe CRS (sCRS) or neurotoxicity (NT) or severe NT (sNT).

[0126] In some aspects, administration of a therapeutic composition or unit dose to one of a group of subjects according to any embodiment provided herein is likely to result in and / or is associated with a particular response, efficacy, or survival outcome, e.g., partial response (PR) or partial remission, complete response (CR) or complete remission, progression-free survival (PFS), objective response (OR), overall survival (OS), event-free survival (EFS), increased duration of response (DOR), or increased survival. In some aspects, administration of a therapeutic composition or unit dose is likely to result in or is associated with increased progression-free survival (PFS).

[0127] In some embodiments of any of the provided embodiments, the method and / or use and / or administration of a unit dose of cells according to the article of manufacture achieves a certain outcome and / or is associated with a certain reduced risk of toxicity, e.g., in a population of subjects treated according to the method or according to information provided in the article of manufacture. In some aspects, at least 35%, at least 40%, at least 50%, or at least 60% of subjects treated according to the method achieve a complete response (CR) or complete remission, progression-free survival (PFS), objective response (OR), overall survival (OS), event-free survival (EFS), or a response that is durable for more than 3 months, more than 6 months, or more than 12 months. In certain embodiments of any of the provided methods, more than 50% or more than about 50% of subjects treated according to the method do not exhibit Grade 3 or higher cytokine release syndrome (CRS) or Grade 3 or higher neurotoxicity. In some embodiments, such subjects do not exhibit early-onset CRS and / or neurotoxicity, e.g., do not exhibit onset of CRS before 3 days after initiation of administration and / or do not exhibit onset of neurotoxicity before 5 days after initiation of administration. In some embodiments of any of the provided methods, greater than or about 30%, greater than or about 35%, greater than or about 40%, or greater than or about 50% of the subjects treated according to the method do not exhibit any grade of cytokine release syndrome (CRS) or neurotoxicity.

[0128] Also provided are output compositions produced by any of the methods described herein.

[0129] Also provided are unit doses determined or produced by any of the methods described herein.

[0130] Also provided are pharmaceutical compositions comprising any of the unit doses described herein. In some embodiments, the pharmaceutical composition also comprises a pharmaceutical carrier.

[0131] Also provided are methods of treatment comprising administering to a mammalian subject all or a portion of any of the output compositions, unit doses or pharmaceutical compositions described herein.

[0132] Also provided is the use of all or a portion of any of the output compositions, unit doses or pharmaceutical compositions described herein to treat cancer.

[0133] Also provided is the use of all or a portion of any of the output compositions, unit doses or pharmaceutical compositions described herein in the manufacture of a medicament for treating cancer.

[0134] Also provided is all or a portion of any of the output compositions, unit doses or pharmaceutical compositions described herein for use in treating cancer.

[0135] A therapeutic cell composition containing T cells expressing a recombinant receptor that binds to an antigen, wherein after stimulation with a stimulatory agent, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least 94% or at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or 95% or about 95%, at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100% of the CD4+ cells in the composition expressing a recombinant receptor are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha; and / orProvided herein are therapeutic cell compositions in which at least 10% or at least about 10%, or 10% or about 10%, at least 15% or at least about 15%, or 15% or about 15%, at least 20% or at least about 20%, or 20% or about 20%, at least 25% or at least about 25%, or 25% or about 25%, at least 30% or at least about 30%, or 30% or about 30%, at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60% or more of the total number of T cells are polyfunctional for the production of two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha. In some embodiments, the CD4+ T cells in the composition expressing a recombinant receptorAt least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least 94% or at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or 95% or about 95%, at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100% of the CD4+ cells in the composition expressing the recombinant receptor are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha.At least 10%, or at least about 10%, or 10% or about 10%, at least 15%, or at least about 15%, or 15% or about 15%, at least 20%, or at least about 20%, or 20% or about 20%, at least 25%, or at least about 25%, or 25% or about 25%, at least 30%, or at least about 30%, or 30% or about 30%, at least 40%, or at least about 40%, or 40% or about 40%, at least 50%, or at least about 50%, or 50% or about 50%, at least 60%, or at least about 60%, or 60% or about 60% or more of the total number of T cells are polyfunctional for the production of two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha. In some embodiments, at least 10% or at least about 10%, or 10% or about 10%, at least 15% or at least about 15%, or 15% or about 15%, at least 20% or at least about 20%, or 20% or about 20%, at least 25% or at least about 25%, or 25% or about 25%, at least 30% or at least about 30%, or 30% or about 30%, at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60% or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are polyfunctional for the production of the cytokines interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

[0136] In some embodiments, the stimulator is a non-specific or non-antigen-dependent T cell stimulator. In some embodiments, the non-specific or non-antigen-dependent T cell stimulator is a polyclonal stimulator. In some embodiments, the non-specific or non-antigen-dependent T cell stimulator comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA). In some embodiments, the non-specific or non-antigen-dependent T cell stimulator comprises PMA / ionomycin. In some embodiments, the stimulator specifically binds to the recombinant receptor, and optionally, the stimulator is an antigen-specific stimulating reagent and / or contains an antigen or a portion thereof that is specifically recognized by the recombinant receptor. In some embodiments, cytokine production is measured in an intracellular cytokine assay.

[0137] In some embodiments, the composition comprises at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60%, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100%, which are surface positive for CCR7 and / or CD27, and optionally further negative for surface expression of CD45RA.

[0138] In some embodiments, the T cells are primary cells obtained from a subject. In some embodiments, the subject has cancer, and optionally, the cancer is leukemia or lymphoma. In some embodiments, the subject is considered to have or is known to have high tumor burden at the time the cells are obtained. In some embodiments, the tumor in the subject has a two-way product of densitometry (SPD) of 30 cm or less. 2 Or about 30cm 2, 40cm 2 Or about 40cm 2 , 50cm 2 Or about 50cm 2 , 60cm 2 Or about 60cm 2 or 70cm 2 Or about 70cm 2 and / or a subject has a high tumor burden if C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 or about 5 milligrams / liter, 10 or about 10 milligrams / liter, 15 or about 15 milligrams / liter, 20 or about 20 milligrams / liter, 25 or about 25 milligrams / liter, 30 or about 30 milligrams / liter, 40 or about 40 milligrams / liter, or 50 or about 50 milligrams / liter. In some embodiments, the SPD of a tumor in a subject is greater than 50 cm 2 or about 50cm 2 In some embodiments, a subject has high tumor burden if CRP is greater than or about 20 milligrams / liter in a biological sample.

[0139] In some embodiments, the therapeutic cell composition is enriched for CD4+ T cells and / or contains a total recombinant receptor agonist in the engineered composition. + At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least about 97%, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD4 + In some embodiments, the therapeutic cell composition is enriched for CD3+ T cells, enriched for CD4+ and CD8+ T cells, and / or contains a total recombinant receptor agonist in the engineered composition. +At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least about 97% of the cells, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD3+ or CD4+. + and CD8+. In some embodiments, the therapeutic cell composition contains CD4+ T cells and CD8+ T cells at a ratio of CD4+ T cells to CD8+ T cells of about 1:3 to about 3:1, optionally about 1:2 to 2:1.

[0140] In some embodiments, the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor. In some embodiments, the recombinant receptor is capable of binding to a target antigen associated with, specific to, and / or expressed on a cell or tissue of a disease, disorder, or condition. In some embodiments, the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer. In some embodiments, the target antigen is a tumor antigen. In some embodiments, the target antigen is αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (also known as CA9, CAIX, or G250), cancer testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, CD123, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutant (EGFR), or EGFR fusion protein (EGFR fusion protein). vIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (g p100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPRC5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Rα),IL-13 receptor alpha 2 (IL-13Rα2), kinase insert domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor The antigen is selected from among receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta-isomerase or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigens, or antigens associated with universal tags, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV, or other pathogens.

[0141] In some embodiments, the recombinant receptor is or contains a functional non-TCR antigen receptor or TCR, or an antigen-binding fragment thereof. In some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR). In some embodiments, the recombinant receptor contains an extracellular domain containing an antigen-binding domain and an intracellular signaling region containing an intracellular signaling domain. In some embodiments, the antigen-binding domain is or contains an antibody or antibody fragment thereof (optionally a single-chain fragment). In some embodiments, the fragment contains antibody variable regions linked by a flexible linker. In some embodiments, the fragment comprises an scFv. In some embodiments, the intracellular signaling domain is or contains a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain containing an immunoreceptor tyrosine-based activation motif (ITAM). In some embodiments, the intracellular signaling domain is or contains the intracellular signaling domain of a CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof. In some embodiments, the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region. In some embodiments, the intracellular signaling region further comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

[0142] In some embodiments, the T cells are primary T cells obtained from the subject. In some embodiments, the T cells are autologous to the subject.

[0143] Provided herein are articles of manufacture including a container containing a therapeutic cell composition described herein and instructions for administering the therapeutic cell composition, optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses, to a subject having a disease or condition. In some embodiments, the instructions specify administering multiple unit doses contained in multiple separate compositions. In some embodiments, the multiple separate compositions are CD8 + T cells and CD4 + a first composition containing a therapeutic cell composition containing one of CD8 T cells and + T cells and CD4 + In some embodiments, the instructions include a second composition containing another of the CD8 T cells, wherein the second composition contains a recombinant receptor or a cell expressing a recombinant receptor specific to an antigen or to a cell expressed by the disease or condition, and optionally the second composition is any composition described herein. + a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, and + The instructions specify that the T cell-containing composition or one or more unit doses thereof and / or amounts corresponding to one or more of such unit doses are administered no more than 48 hours apart, no more than 36 hours apart, no more than 24 hours apart, no more than 12 hours apart, no more than 6 hours apart, no more than 2 hours apart, no more than 1 hour apart, or simultaneously. + Prior to administering a composition containing CD8 cells or one or more unit doses thereof and / or an amount corresponding to one or more of such unit doses, + It is specified that a composition containing T cells or one or more unit doses thereof and / or an amount corresponding to one or more such unit doses is administered.

[0144] Provided herein are methods of treatment comprising administering to a subject having a disease or condition one or more unit doses of a therapeutic cell composition described herein. In some embodiments, the therapeutic composition is a first therapeutic cell composition containing one of CD8+ T cells and CD4+ T cells, and the one or more unit doses further contain a second therapeutic cell composition containing the other of CD8+ T cells and CD4+ cells, the second composition containing cells expressing a recombinant receptor or a recombinant receptor specific to an antigen or to cells expressed by the disease or condition, and optionally the second composition is any composition described herein.

[0145] In some embodiments, at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60%, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92%, at least 93%, or at least about 93%, or 93% or about 93%, at least 94%, or at least about 94%, or 94% or about 94%, at least 95%, or at least about 95%, or 95% or about 95%, at least 96%, or at least about 96%, or 96% or about 96%, at least 97%, or at least about 97%, or 97% or about 97%, at least 98%, or at least about 98%, or 98% or about 98%, at least 99%, or at least about 99%, or 99% or about 99%, or at least 100%, or at least about 100%, or 100% or about 100% are surface positive for CCR7 and / or CD27, and optionally further negative for surface expression of CD45RA. In some embodiments, the second therapeutic cell composition is enriched for CD8+ T cells and / or is negative for the total recombinant receptor agonist (CRG) expression in the engineered composition. +At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least about 97% of the cells, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD8 + is.

[0146] In some embodiments, the first and second therapeutic compositions are administered within 48 hours or about 48 hours, 36 hours or about 36 hours, 24 hours or about 24 hours, 12 hours or about 12 hours, 6 hours or about 6 hours, 2 hours or about 2 hours, or 1 hour or about 1 hour of each other. In some embodiments, the second therapeutic cell composition containing CD8+ T cells is administered before the first composition containing CD4+ T cells. In some embodiments, one or more unit doses of cells contain a defined ratio of CD4+ T cells to CD8+ T cells, the ratio being 1:3 and 3:1, or about 1:3 and about 3:1, optionally 1:2 and 2:1, or about 1:2 and 2:1, optionally 1:1 or about 1:1.

[0147] (A) The engineered cell composition containing recombinant receptor-expressing T cells is analyzed by measuring (i) the percentage of T cells expressing the recombinant receptor that are surface positive for CCR7 and / or CD27, and optionally surface negative for CD45RA; and / or (ii) the percentage of CD4+ T cells expressing the recombinant receptor that are capable of producing IL-2, TNF-alpha, and / or IFN-gamma after stimulation with a stimulator. and (B) administering a therapy to a subject having a disease or condition, comprising: (1) administering to the subject one or more unit doses of cells of the engineered cell composition containing T cells expressing the recombinant receptor if the factor indicative of the function or phenotype of cells of the engineered T cell composition is at or above a threshold value; or (2) administering to the subject one or more unit doses of cells of the engineered cell composition containing T cells expressing the recombinant receptor if the factor indicative of the function or phenotype of cells of the engineered T cell composition is below the threshold value for the factor, (a) one or more unit doses of cells of the engineered cell composition and an agent capable of increasing the expansion, proliferation, or efficacy of the T cells of the engineered cell composition in the subject, (b) comparing the increased number of cells of the engineered cell composition, optionally with one or more unit doses administered to a similarly situated subject having the same disease or condition but exhibiting at or above the threshold value for the factor. or (c) administering a therapy selected from an alternative therapeutic treatment for treating a disease or condition other than one or more unit doses of the engineered cell composition, wherein the threshold factor is (i) at or about 40%, 50%, or about 50%, 60%, or about 60%, 70%, or more of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor. are about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% surface positive for CCR7 and / or CD27;and / or (ii) at or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing interleukin 2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or (iii) at or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing interleukin 2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; Provided herein are methods wherein at or about 10%, 15% or about 15%, 20% or about 20%, 25% or about 25%, 30% or about 30%, 40% or about 40%, 50% or about 50%, 60% or about 60% or more of the total number of T cells are selected as being polyfunctional for the production of two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha after stimulation with a stimulator;

[0148] administering a therapy to a subject having a disease or condition, comprising: (1) administering to the subject one or more unit doses of cells of the engineered T cell composition when a factor indicative of the function or phenotype of cells of the engineered cell composition containing T cells expressing the recombinant receptor is at or above a threshold value; or (2) when the factor indicative of the function or phenotype of cells of the engineered T cell composition is below the threshold value of the factor, (a) one or more unit doses of cells of the engineered cell composition and an agent capable of increasing the expansion, proliferation or efficacy of the T cells of the engineered cell composition in the subject, (b) an increased dose of cells of the engineered cell composition, optionally increased compared to one or more unit doses administered to a similarly situated subject having the same disease or condition but exhibiting at or above the threshold value of the factor; or (c) administering an alternative therapy for treating the disease or condition other than the dose of the engineered cell composition. and / or administering a therapy selected from the group consisting of a therapeutic treatment, wherein the threshold factor is (i) at or about 40%, 50%, or about 50%, 60%, or about 60%, 70%, or about 70%, 80%, or about 80%, 85%, or about 85%, 90%, or about 90%, 91%, or about 91%, 92%, or about 92%, 93%, or about 93%, 94%, or about 94%, 95%, or about 95%, 96%, or about 96%, 97%, or about 97%, 98%, or about 98%, 99%, or about 99%, or 100% or about 100% of the total number of T cells in the composition or of the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27; and / or (ii) at or about 100% of the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27; 70% or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of T cells are capable of producing interleukin-2 (IL-2) or TNF-alpha after stimulation with a stimulant;and / or (iii) 10% or about 10%, 15% or about 15%, 20% or about 20%, 25% or about 25%, 30% or about 30%, 40% or about 40%, 50% or about 50%, 60% or about 60% or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are selected from being polyfunctional for the production of two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha after stimulation with a stimulatory agent.

[0149] In some embodiments, the cells of the cell composition are autologous to the subject. In some embodiments, at least 35%, at least 40%, at least 50%, at least 60%, or at least 70%, or at least 75% of the subjects treated according to the method achieve progression-free survival for at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 9 months, or at least 12 months after administration of one or more unit doses. In some embodiments, at least 35%, at least 40%, at least 50%, at least 60%, or at least 70%, or at least 75% of the subjects treated according to the method do not experience disease progression for at least 3 months or at least about 3 months, at least 4 months or at least about 4 months, at least 5 months or at least about 5 months, at least 6 months or at least about 6 months, at least 9 months or at least about 9 months, or at least 12 months or at least about 12 months after administration of one or more unit doses. In some embodiments, subjects treated according to the methods achieve, on average, improved pharmacokinetic properties of the cells of the cell composition in a biological sample, optionally a blood or serum sample, compared to a group of similarly situated subjects with the same disease or condition who are administered a similar dose of an autologous cell composition comprising T cells expressing a recombinant receptor, but whose factors indicative of cellular function or phenotype are below a threshold. In some embodiments, the pharmacokinetic property is total exposure over time (AUC) or peak number of cells in the biological sample.

[0150] 1. A method of predicting likelihood of response to a therapeutic T cell composition, comprising: (a) assaying an engineered cell composition containing T cells expressing a recombinant receptor for a factor indicative of the function or phenotype of the engineered cells in the composition selected from: (i) the percentage of T cells expressing a recombinant receptor that is surface positive for CCR7 and / or CD27, and optionally surface negative for CD45RA; and / or (ii) the percentage of CD4+ T cells expressing a recombinant receptor that is capable of producing IL-2, TNF-alpha, and / or IFN-gamma after stimulation with a stimulator; and (b) determining the likelihood of response following administration of a dose of a cell therapy containing the engineered cells, wherein if the factor is at or above a threshold value, the subject is considered likely to achieve a sustainable response or progression-free survival, optionally for at least three months after administration of the therapy; or if the factor is below a threshold value, the subject is considered unlikely to exhibit a sustainable response or progression-free survival to the therapy, the threshold value for the factor being selected from: (i) the total number of T cells in the composition; or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the number or total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27;and / or (ii) at or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing interleukin 2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or (iii) at or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing interleukin 2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; Provided herein are methods comprising the steps of: selecting at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or more of the total number of T cells as being polyfunctional for the production of two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha after stimulation with a stimulator;

[0151] In some embodiments, the cells of the cell composition are autologous to the subject. In some embodiments, when the subject is deemed likely to achieve a sustainable response or progression-free survival to the therapy, one or more unit doses of cells of the engineered T cell composition are administered to the subject. In some embodiments, the one or more unit doses are 1×10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 106 and 1×10 7 Or about 1 x 10 7 Total T cells expressing recombinant receptors between + In some embodiments, one or more unit doses contain about 5 x 10 T cells (i.e., 1 x 10 T cells) or total T cells. 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below +In some embodiments, the threshold factor is that 40%, or about 40%, 50%, or about 50%, 60%, or about 60%, 70%, or about 70%, 80%, or about 80%, 85%, or about 85%, 90%, or about 90%, 91%, or about 91%, 92%, or about 92%, 93%, or about 93%, 94%, or about 94%, 95%, or about 95%, 96%, or about 96%, 97%, or about 97%, 98%, or about 98%, 99%, or about 99%, or 100% or about 100% of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27. In some embodiments, the threshold for the factor is that 40% or about 40%, 50% or about 50%, 60% or about 60%, 70% or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of T cells in the composition or the total number of T cells in the composition expressing the recombinant receptor are surface positive for CCR7 and / or CD27. In some embodiments, the threshold factor is that 70% or about 70%, 80% or about 80%, 85% or about 85%, 90% or about 90%, 91% or about 91%, 92% or about 92%, 93% or about 93%, 94% or about 94%, 95% or about 95%, 96% or about 96%, 97% or about 97%, 98% or about 98%, 99% or about 99%, or 100% or about 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha.In some embodiments, the threshold factor is that 10% or about 10%, 15% or about 15%, 20% or about 20%, 25% or about 25%, 30% or about 30%, 40% or about 40%, 50% or about 50%, 60% or about 60% or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are polyfunctional for the production of two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha. In some embodiments, the threshold factor is that 10% or about 10%, 15% or about 15%, 20% or about 20%, 25% or about 25%, 30% or about 30%, 40% or about 40%, 50% or about 50%, 60% or about 60% or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are polyfunctional for the production of the cytokines interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

[0152] In some embodiments, if a subject is deemed unlikely to achieve a durable response or progression-free survival, the subject is administered a therapy selected from: (a) a dose of cells of the engineered cell composition and an agent capable of increasing the expansion, proliferation, or efficacy of T cells of the engineered cell composition in the subject; (b) an increased dose of cells of the engineered cell composition, optionally increased compared to the dose administered to a similarly situated subject with the same disease or condition but who exhibits a likelihood of achieving progression-free survival or a durable response to the cell therapy; or (c) an alternative therapeutic treatment for treating the disease or condition other than the dose of the engineered cell composition.

[0153] In some embodiments, the methods involve administering a therapy comprising a dose of cells of the engineered cell composition and an agent capable of increasing the expansion, proliferation, or efficacy of T cells of the engineered cell composition in a subject, wherein the agent is an anti-idiotypic antibody or antigen-binding fragment thereof specific for a CAR, an immune checkpoint inhibitor, a modulator of a metabolic pathway, an adenosine receptor antagonist, a kinase inhibitor, an anti-TGFβ antibody or an anti-TGFβR antibody, or a cytokine. In some embodiments, the agent is administered before, simultaneously with, or after administration of the therapeutic T cell composition.

[0154] In some embodiments, the subject is considered to have or is known to have a high tumor burden at the time of the assay and / or prior to administration. In some embodiments, the sum of two-way products (SPD) of tumors in the subject is greater than 30 cm 2 Or about 30cm 2 , 40cm 2 Or about 40cm 2 , 50cm 2 Or about 50cm 2 , 60cm 2 Or about 60cm 2 or 70cm 2 Or about 70cm 2 and / or a subject has a high tumor burden if C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 or about 5 milligrams / liter, 10 or about 10 milligrams / liter, 15 or about 15 milligrams / liter, 20 or about 20 milligrams / liter, 25 or about 25 milligrams / liter, 30 or about 30 milligrams / liter, 40 or about 40 milligrams / liter, or 50 or about 50 milligrams / liter. In some embodiments, the SPD of a tumor in a subject is greater than 50 cm 2 or about 50cm 2 In some embodiments, a subject has high tumor burden if CRP is greater than or about 20 milligrams / liter in a biological sample.

[0155] In some embodiments, the stimulator is a non-specific or non-antigen-dependent T cell stimulator. In some embodiments, the non-specific or non-antigen-dependent T cell stimulator is a polyclonal stimulator. In some embodiments, the non-specific or non-antigen-dependent T cell stimulator comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA). In some embodiments, the non-specific or non-antigen-dependent T cell stimulator comprises PMA / ionomycin. In some embodiments, the stimulator specifically binds to the recombinant receptor, and optionally, the stimulator is an antigen-specific stimulating reagent and / or comprises an antigen or a portion thereof that is specifically recognized by the recombinant receptor. In some embodiments, cytokine production is measured in an intracellular cytokine assay.

[0156] In some embodiments, the recombinant receptor is or contains a chimeric receptor and / or a recombinant antigen receptor. In some embodiments, the recombinant receptor is capable of binding to a target antigen associated with, specific to, and / or expressed on a cell or tissue of a disease, disorder, or condition. In some embodiments, the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer. In some embodiments, the target antigen is a tumor antigen. In some embodiments, the target antigen is αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (also known as CA9, CAIX, or G250), cancer testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, CD123, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutant (EGFR), or EGFR fusion protein (EGFR fusion protein). vIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (g p100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPRC5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Rα),IL-13 receptor alpha 2 (IL-13Rα2), kinase insert domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor The antigen is selected from among receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta-isomerase or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigens, or antigens associated with universal tags, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV, or other pathogens.

[0157] In some embodiments, the recombinant receptor is or comprises a functional non-TCR antigen receptor or TCR, or an antigen-binding fragment thereof. In some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR). In some embodiments, the recombinant receptor comprises an extracellular domain containing an antigen-binding domain and an intracellular signaling region containing an intracellular signaling domain. In some embodiments, the antigen-binding domain is or comprises an antibody or antibody fragment thereof (optionally a single-chain fragment). In some embodiments, the fragment comprises antibody variable regions linked by a flexible linker. In some embodiments, the fragment comprises an scFv. In some embodiments, the intracellular signaling domain is or comprises a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain containing an immunoreceptor tyrosine-based activation motif (ITAM). In some embodiments, the intracellular signaling domain is or comprises the intracellular signaling domain of a CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof. In some embodiments, the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region. In some embodiments, the intracellular signaling region further comprises a costimulatory signaling domain. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof. In some embodiments, the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

[0158] In some embodiments, the T cells are primary T cells obtained from the subject. In some embodiments, the T cells are autologous to the subject.

[0159] A method for producing a therapeutic T cell composition, comprising: (a) selecting T cells that do not express apoptotic markers (apoptotic marker negative ( - (b) incubating an input composition containing a defined number of T cells under stimulatory conditions, thereby producing a stimulated composition; and (b) introducing a recombinant receptor into cells from the stimulated composition, thereby producing an engineered cell composition, wherein the introducing comprises contacting cells of the stimulated composition with an agent comprising a polynucleotide encoding the recombinant receptor. In some embodiments, the apoptosis marker-negative T cells are apoptosis marker-negative CD3 + Cells, apoptosis marker negative CD8 + In some embodiments, the apoptosis marker is annexin V. In some embodiments, the apoptosis marker is activated caspase 3. In some embodiments, the defined number of T cells is at least 50 x 10 6 In some embodiments, the defined number of T cells comprises at least 100 x 10 apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells. 6 In some embodiments, the defined number of T cells comprises 100 x 10 apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells. In some embodiments, the defined number of T cells comprises 100 x 10 6 or about 100 x 10 6 and 500 x 10 6 or approximately 500 x 10 6 In some embodiments, the defined number of T cells comprises between 300 x 10 and 300 x 10 apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells. 6 Or about 300 x 10 6of apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells.

[0160] In some embodiments, the percentage of apoptosis marker-negative T cells as a percentage of total T cells in the input composition is greater than or about 70%, greater than or about 80%, greater than or about 90%, or greater than or about 90%, or greater than or about 95%. In some embodiments, apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells, are isolated, selected, or enriched from the biological sample prior to incubating.

[0161] A method for producing a therapeutic T cell composition, comprising: (a) selecting from a biological sample T cells that do not express apoptotic markers (apoptotic marker negative ( - Provided herein are methods comprising: (a) isolating, selecting, or enriching a population of T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells, thereby obtaining an input composition; (b) incubating the input composition under stimulatory conditions, thereby producing a stimulated composition; and (c) introducing a recombinant receptor into cells from the stimulated composition, thereby producing an engineered cell composition, wherein the introducing comprises contacting cells of the stimulated composition with an agent comprising a polynucleotide encoding the recombinant receptor.

[0162] In some embodiments, the biological sample is primary T cells obtained from a subject. In some embodiments, the subject is a human subject. In some embodiments, the biological sample is or contains a whole blood sample, a buffy coat sample, a peripheral blood mononuclear cell (PBMC) sample, an unfractionated T cell sample, a lymphocyte sample, a leukocyte sample, an apheresis product, or a leukapheresis product. In some embodiments, the biological sample contains an apheresis product or a leukapheresis product. In some embodiments, T cells are isolated, selected, or enriched from the biological sample within 36 hours after being obtained from the subject. In some embodiments, the biological sample is deep-frozen in the presence of a cryoprotectant within 36 hours after being obtained from the subject and thawed before isolating, selecting, or enriching the cells; optionally, the deep-frozen sample is thawed immediately before, or within 6 hours, 4 hours, 2 hours, or 1 hour before isolating, selecting, or enriching the cells.

[0163] In some embodiments, the incubation is performed in the presence of one or more cytokines, optionally in serum-free medium. In some embodiments, the one or more cytokines are selected from recombinant IL-2, recombinant IL-7, and / or recombinant IL-15. In some embodiments, the one or more cytokines include 10-200 IU / mL recombinant IL-2; 100 IU / mL-1,000 IU / mL recombinant IL-7; and / or 10-200 IU / mL recombinant IL-15. In some embodiments, the stimulatory reagent contains a primary agent that specifically binds to a member of the TCR complex, optionally specifically binds to CD3, and optionally the primary agent is an antibody or antigen-binding fragment thereof. In some embodiments, the stimulatory reagent further contains a secondary agent that specifically binds to a T cell costimulatory molecule, optionally the costimulatory molecule is selected from CD28, CD137 (4-1-BB), OX40, or ICOS, and optionally the secondary agent is an antibody or antigen-binding fragment thereof. In some embodiments, the stimulation reagent comprises incubation with an anti-CD3 antibody and an anti-CD28 antibody, or an antigen-binding fragment thereof. In some embodiments, the primary agent and / or the secondary agent are present on the surface of a solid support. In some embodiments, the solid support is or comprises beads. In some embodiments, the primary agent and the secondary agent are immobilized or reversibly bound to the surface of an oligomeric particle reagent comprising a plurality of streptavidin or streptavidin mutein molecules.

[0164] In some embodiments, the input composition is incubated under stimulatory conditions for at least 12 hours and up to 36 hours, optionally at or about 24 hours. In some embodiments, the contacting is performed by viral transduction, optionally using a retroviral vector, optionally a lentiviral vector.

[0165] In some embodiments, the method further comprises culturing the engineered composition under conditions that promote the growth and / or expansion of the engineered cells, thereby producing an output composition comprising the engineered T cells. In some embodiments, the culturing is carried out in the presence of one or more cytokines, optionally in serum-free medium. In some embodiments, the one or more cytokines are selected from recombinant IL-2, recombinant IL-7, and / or recombinant IL-15. In some embodiments, the one or more cytokines comprise 50-400 IU / mL recombinant IL-2; 100 IU / mL-2,000 IU / mL recombinant IL-7; and / or 50-400 IU / mL recombinant IL-15.

[0166] In some embodiments, the input composition contains CD4+ T cells and CD8+ T cells at a ratio of 2:1 to 1:2; or the input composition contains apoptosis marker-negative CD4+ T cells and apoptosis marker-negative CD8+ T cells at a ratio of 2:1 to 1:2. In some embodiments, the CD4+ T cells and CD8+ T cells are separately selected, isolated, or enriched from the same biological sample or a sample derived therefrom and combined before incubation. In some embodiments, the recombinant receptor is a chimeric antigen receptor (CAR).

[0167] In some embodiments, the output composition contains at least 40%, at or about 40%, at least 50%, at or about 50%, at least 60%, at or about 60%, at least 70%, at or about 70%, at least 80%, at or about 80%, at least 85%, at or about 85%, at least 90%, at or about 90%, at least 91%, at or about 91%, at least 92%, at or about 92%, at least 93%, at or about 93%, at least 94%, at or about 94%, at least 95%, at or about 95%, at least 96%, at or about 96%, at least 97%, at or about 97%, at least 98%, at or about 98%, at least 99%, or at least 100% or about 100% of the total number of T cells in the composition or of the total number of T cells in the composition expressing the recombinant receptor, which are surface positive for CCR7 and / or CD27; and / or CD4+ T cells in the composition expressing the recombinant receptor. at least 70%, or at least about 70%, at least 80%, or at least about 80%, at least 85%, or at least about 85%, at least 90%, or at least about 90%, at least 91%, or at least about 91%, at least 92%, or at least about 92%, at least 93%, or at least about 93%, at least 94%, or at least about 94%, at least 95%, or at least about 95%, at least 96%, or at least about 96%, at least 97%, or at least about 97%, at least 98%, or at least about 98%, at least 99%, or at least about 99%, or at least 100% or at least about 100% of the total number of T cells are capable of producing interleukin 2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent;and / or at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are polyfunctional for the production of two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha after stimulation with a stimulator. In some embodiments, the stimulator is a non-specific or non-antigen-dependent T cell stimulator. In some embodiments, the non-specific or non-antigen-dependent T cell stimulator is a polyclonal stimulator. In some embodiments, the non-specific or non-antigen-dependent stimulator comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA). In some embodiments, the non-specific or non-antigen-dependent T cell stimulator contains PMA / ionomycin. In some embodiments, the stimulator specifically binds to the recombinant receptor, and optionally, the stimulator is an antigen-specific stimulating reagent and / or contains an antigen or portion thereof that is specifically recognized by the recombinant receptor. In some embodiments, cytokine production is measured in an intracellular cytokine assay. [Brief explanation of the drawings]

[0168] [Figure 1A] Figure 1A shows the number of CD3+ CAR+ T cells present in the CAR T cell composition for administration in DL1 and DL2. [Figure 1B] Figure 1B shows the numbers of CD4+CAR+ and CD8+CAR+ cells, CD4+CAR+TNF-α+ cells and CD8+CAR+TNF-α+ cells, CD4+CAR+CCR7+ cells and CD8+CAR+CCR7+ cells present in the CAR T cell compositions for administration in DL1 and DL2. [Figure 1C] Figure 1C shows the numbers of CD4+CAR+ and CD8+CAR+ cells, CD4+CAR+CCR7+ cells and CD8+CAR+CCR7+ cells present in the CAR T cell compositions for administration in DL1 and DL2. [Figure 2] Figure 1 depicts the median (±quartile) number of CAR-expressing CD3+ cells / μL of blood assessed by flow cytometry using an antibody specific for the truncated receptor (CD3, circles; N=87) in blood samples from 87 subjects receiving anti-CD19 CAR-expressing cells; or the median (±quartile) number of integrated CAR transgene copies / μg of genomic DNA assessed by quantitative polymerase chain reaction (qPCR) using primers specific for the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) present in the CAR-encoding vector (qPCR, squares; N=85). The cutoff for CAR+ cell detection in flow cytometry was set at ≥25 events in the CAR+ gate, and the detection limit for qPCR was ≥12.5 CAR transgene copies per μg of genomic DNA. [Figure 3A]Figure 3A shows plots of the percentage of CCR7+ cells (specifically, the CD4+ CAR+ and CD8+ CAR+ cell populations) against the pharmacokinetic parameters of CD4+ CAR+ cells and CD8+ CAR+ cells, respectively (as indicated by the AUC0-28 of CD4+ CAR+ cells or the AUC0-28 of CD8+ CAR+ cells in the blood for subjects administered cell therapy). Each dot represents an individual patient, and the shading of the individual dot indicates whether the patient received dose level 1 (DL1) or dose level 2 (DL2). Figure 3B shows similar plots separately for subjects observed to have high (indicated in this assessment by a two-way sum of products (SPD) value greater than or equal to 50 cm2) or low (SPD value less than 50 cm2) pretreatment tumor burden (assessed at the time of pretreatment lymphodepleting chemotherapy). Figure 3C depicts plots of the number of CD4+CAR+CCR7+ cells and CD8+CAR+CCR7+ cells against the AUC0-28 of CD4+ CAR+ cells or the AUC0-28 of CD8+ CAR+ cells in the blood for subjects administered cell therapy, along with the correlation coefficient and associated p-value. Figures 3D-3J depict plots of the percentage of CCR7+ or CCR7+CD27+ CD4+ or CD8+ CAR+ T cells against the pharmacokinetic parameters (AUC0-28 and Cmax) of CD4+ CAR+ T cells (Figures 3A, 3D-3F) and CD8+ CAR+ T cells (Figures 3G-3J), along with the correlation coefficient and associated p-value. [Figure 3B] See legend to Figure 3A. [Figure 3C] See legend to Figure 3A. [Figure 3D] See legend to Figure 3A. [Figure 3E] See legend to Figure 3A. [Figure 3F] See legend to Figure 3A. [Figure 3G] See legend to Figure 3A. [Figure 3H] See legend to Figure 3A. [Figure 3I] See legend to Figure 3A. [Figure 3J]See legend to Figure 3A. [Figure 4] Figures 4A-4B depict plots of the percentage of CCR7+CD27+ cells among CD4+CAR+ cells versus IFNγ (Figure 4A) and IL-2 (Figure 4B) secretion by CD4+ CAR+ compositions upon stimulation with the antigen recognized by the CAR (CD19). [Figure 5] Figures 5A-5B depict plots of the percentage of CCR7+CD27+ cells among CD8+CAR+ cells versus IL-5 (Figure 5A) and IL-13 (Figure 5B) secretion by CD4+ CAR+ compositions upon stimulation with the antigen recognized by the CAR (CD19). [Figure 6A] Figures 6A-6D show Kaplan-Meier survival curves for subjects who received CAR+ T cell compositions divided into groups who received compositions containing a percentage of CCR7+CD27+ CAR+ T cells above or below certain threshold levels among CD4+ CAR+ T cells (Figure 6A for progression-free survival, Figure 6C for duration of response) and among CD8+ CAR+ T cells (Figure 6B for progression-free survival, Figure 6D for duration of response). [Figure 6B] See legend to Figure 6A. [Figure 6C] See legend to Figure 6A. [Figure 6D] See legend to Figure 6A. [Figure 7-1] Figures 7A-7B show box plots depicting the percentage of CCR7+CD27+ cells among CD4+CAR+ cells (Figure 7A) or IFNγ secretion by CD4+ CAR+ cells (Figure 7B) in a cell composition administered to a subject as a function of whether the subject subsequently developed cytokine release syndrome (CRS; Grade 0 vs. Grade 1 or higher). Figures 7C-7D show box plots depicting the percentage of CCR7+CD27+ cells among CD4+CAR+ cells (Figure 7C) or IFNγ secretion by CD4+ CAR+ cells (Figure 7D) in a cell composition administered to a subject as a function of whether the subject subsequently developed neurotoxicity (NT; Grade 0-2 vs. Grade 3 or higher). [Figure 7-2] See description of Figure 7-1. [Figure 8] Figure 8A shows an exemplary multiparameter phenotypic analysis based on t-distributed stochastic neighbor embedding (tSNE), and Figure 8B shows exemplary flow cytometry plots (within the outline boundaries drawn on Figure 8A) of cell subtypes identified from the analysis of CCR7 and CD27 expression. [Figure 9-1] Figures 9A-9C show exemplary partial least squares discriminant analysis (PLS-DA) assessment of phenotypic and functional attributes (Figure 9A) and transcriptomic (Figure 9B) and epigenomic analyses (Figure 9C) to identify variables that distinguish between subjects who experienced a complete response (CR) and those who experienced progressive disease (PD) or partial response (PR) 6 months after administration of a CAR+ T cell composition. [Figure 9-2] See description of Figure 9-1. [Figure 10A] Figure 10A shows a plot of the frequency of CD4+CAR+CCR7+CD27+ cells in the engineered cell composition versus the frequency of CD4+CCR7+CD27+ cells in the composition before engineering (Spearman's rank correlation coefficient: 0.591). [Figure 10B] Figure 10B shows a plot of the frequency of CD8+CCR7+CD27+ cells in the pre-manipulation composition versus the frequency of CD8+CAR+CCR7+CD27+ cells in the engineered cell composition (Spearman's rank correlation coefficient: 0.357). [Figure 11A]Figures 11A-11D depict results from CAR+ T cell compositions generated from exemplary expansion and non-expansion processes involving bead-based stimulation reagents (Beads) and incubation in basal media (Bead-Basal Media) or oligomeric stimulation reagents (Oligomers). Figure 11A depicts the percentage of CD4+ and CD8+ T cells positive for both CCR7 and CD27. Figure 11B depicts the percentage of CCR7+CD27+ cells relative to CD4+ CAR+ T cells. Figure 11C depicts the percentage of CCR7+CD27+ cells relative to CD8+ CAR+ T cells. FIG. 11D displays the percentage of CCR7+ CD27+ cells generated from a representative donor from an exemplary expansion process on various days during the manufacturing process, including activation on day 1 (activation d1), transduction on day 2 (transduction d2), and various time points after initiation of culture (d4 INOC+2, d6 INOC+4, d7 INOC+5). [Figure 11B] See legend to Figure 11A. [Figure 11C] See legend to Figure 11A. [Figure 11D] See legend to Figure 11A. [Figure 12] Shown is T cell clonality before manipulation of isolated CD4+ and CD8+ T cell compositions and after manipulation of CD4+ and CD8+ therapeutic CAR+ T cell compositions (Shannon index applied). [Figure 13A] Figures 13A and 13B show progression-free survival curves (Cox proportional hazards (top) and optimal split (bottom)) for patients whose therapeutic composition had a "high" or "low" percentage of IL-2- or TNFα-producing cells among their CD4+ / CAR+ T cells, respectively, as determined using maximum selection rank statistics. [Figure 13B] See legend to Figure 13A. [Figure 14]Figure 1 shows the relationship between clinical outcomes and baseline sum of two-way products (SPD) and baseline C-reactive protein (CRP) levels in subjects with complete response (CR) (triangles) or loss of response / no response (circles) at 6 months. [Figure 15] 1 shows the percentage of IL-2 / CD4+ / CAR+ cells in therapeutic T cell compositions generated from high baseline SPD / CRP in subjects showing complete response (CR) (triangle; 1) or loss of response / no response (circle; 0) at 6 months. [Figure 16] 1 shows progression-free survival curves for exemplary tertile cutoff values ​​for cells positive for the apoptotic marker activated caspase 3 in the input composition. [Figure 17] Figure 1 shows the percentage of cells positive for the apoptotic marker activated caspase 3 in the enriched input T cell composition as a function of the age of the starting apheresis sample, i.e., the length of time (e.g., in hours (h)) between apheresis and its further processing prior to enrichment of T cells. The left plot shows the percentage of CD4+ / activated caspase 3+ cells, and the right plot shows the percentage of CD8+ / activated caspase 3+ cells. DETAILED DESCRIPTION OF THE INVENTION

[0169] Detailed Description Provided herein are methods, compositions, and articles of manufacture for use in connection with cell therapy, such as engineered T cell therapy, for the treatment of various diseases and conditions, including tumors. Provided embodiments relate to therapeutic T cell compositions containing engineered T cells, e.g., T cells engineered to express recombinant receptors, e.g., recombinant proteins, designed to recognize and / or specifically bind to molecules associated with a disease or condition, and to elicit a response, such as an immune response, against such molecules upon binding. The receptors can include chimeric receptors, e.g., chimeric antigen receptors (CARs), and other transgenic antigen receptors, including transgenic T cell receptors (TCRs).

[0170] In some aspects, provided embodiments relate to aspects in which a cell phenotype, e.g., the presence and / or expression of a surface marker and / or the absence or lack of expression of a surface marker, is related to the function or activity of a therapeutic T cell composition, pharmacokinetic parameters, e.g., exposure or maximum cell concentration, the risk of potential toxicity such as cytokine release syndrome (CRS) or neurotoxicity (NT), and / or the outcome of a cell therapy in a subject to which the T cell composition is administered, e.g., response to cell therapy, and / or may provide information regarding the efficacy of a therapeutic T cell composition. In some aspects, provided embodiments are based on the observation that certain cell phenotypes, e.g., expression of one or more surface markers, production of one or more cytokines, are associated with pharmacokinetic parameters, likelihood of response, and / or likelihood of toxicity. In some embodiments, the expression and / or absence of expression of cell surface markers, such as CC chemokine receptor type 7 (CCR7), CD27, and CD45RA, or a combination thereof, in a T cell composition for administration, e.g., a drug composition, correlates positively or negatively with pharmacokinetic parameters and / or response or toxicity outcomes. In some aspects, it is observed herein that phenotypic and functional attributes associated with less differentiated T cell products or products enriched for naive-like or central memory T cell subsets correlate with or show a relationship to improved pharmacokinetic properties or response, e.g., duration of response and / or progression-free survival, following administration to a subject.

[0171] Aspects of the provided embodiments also relate to administering or providing unit doses containing numbers, ratios, percentages, and / or proportions of cells that correlate with the number of cells of a particular phenotype, such as a phenotype indicative of a cell population, including uses related to dosing of therapeutic T cell compositions. In some aspects, the provided embodiments allow for consistent dosing and / or administration. In some aspects, the provided embodiments may be used to assess the likelihood of a particular outcome and / or the value of a particular pharmacokinetic parameter, the risk of toxicity, and / or the response, efficacy, or effectiveness of a cell therapy. In some aspects, the provided methods can control, ameliorate, or reduce the risk of toxicity in a subject administered a dose of a therapeutic T cell composition while ensuring the efficacy of the T cell composition. Also provided are articles of manufacture containing the cells and designed for administration according to such dosing regimens.

[0172] In some embodiments, the provided methods, doses, unit doses, compositions, and articles of manufacture are based on the observation that it may be advantageous to consider certain phenotypes, e.g., expression of surface markers, and combinations thereof, when determining the appropriate dose of cell therapy and / or when releasing or creating cell compositions for therapy. In certain available methods, the dose is based on the number of specific cell types, e.g., cell types engineered to exhibit a particular activity, e.g., cell types positive for an engineered receptor. For example, in certain available methods and doses, the dose is based on the number of viable cytotoxic (e.g., CD8 +These metrics are based on observed or suspected relationships between the number (or number per patient weight) of viable engineered T cells or subsets thereof, such as engineered T cells (e.g., T cells from a cell line). In various circumstances, such numbers may correlate with efficacy and / or safety outcomes (e.g., response) and / or risk of toxicity (e.g., neurotoxicity, cerebral edema, and CRS). Nevertheless, provided herein are embodiments based on the observation that such metrics, in some circumstances, do not consistently and adequately correlate with pharmacological parameters, toxicity, CRS, or NT risk in a subject, particularly without taking other variables into account. Thus, approaches to defining doses and evaluating products for release that rely solely on such metrics may not be fully satisfactory. For example, such approaches may, in some circumstances, fall short of certain therapeutic ranges or windows for safe and effective cell therapy. Provided herein are methods (including treatments, dosing, dose determinations, and assays), therapeutic compositions, and articles of manufacture that address such shortcomings.

[0173] In some aspects, provided embodiments enable the administration of controlled and consistent doses of cells, thereby minimizing variability in efficacy and / or safety outcomes in subjects. In some aspects, controlling the dose of cells based on a defined number, ratio, percentage, and / or proportion of a particular cell subset, e.g., based on cell phenotype, allows for understanding the impact of a cell subset having a particular phenotype on the health, potency, and / or efficacy of cells contained in a therapeutic composition. Such an approach can be used to determine and / or calculate a consistent and accurate effective dose of cells in a cell therapy and / or to control the pharmacokinetic parameters of the cell therapy. Methods for determining such doses, including unit doses for administration, based on the number, ratio, percentage, and / or proportion of cells expressing specific surface markers and / or having a specific phenotype are provided, as well as articles of manufacture and kits containing doses determined using such methods and instructions for administration, and related methods of treatment. In some aspects, provided embodiments provide for the identification of CCR7 in a therapeutic composition for administration to a subject, the attributes of cells in the engineered cell composition and their potential correlation with pharmacokinetics (PK) and clinical outcomes, such as response and toxicity. + , CD27 + and / or CD45RA-, CD4 + and CD8 + CAR + This allows for consistent and tightly controlled identification of T cells and, optionally, their subsets in the cell composition for administration. + Such consistency and tight control of total cell numbers can provide the basis for strategies to evaluate and control other aspects of cells in engineered cell compositions for administration.

[0174] In some aspects, provided embodiments provide CAR antibodies that exhibit low variability, are pure, and contain precise doses. +This allows for the administration of T cell compositions, thereby enabling the identification of relationships between various cellular attributes and clinical outcomes, such as response and toxicity. In some aspects, the provided embodiments are based on the observation that cell populations expressing markers associated with certain phenotypes, such as less differentiated phenotypes, exhibit certain functional profiles (e.g., cytokine production). In some aspects, CCR7 + CD27 + CD4 + CAR T cells produce reduced IFNγ and increased IL-2, and in some aspects, CCR7 + CD27 + CD8 + The CAR T cells produce reduced IL-5 and IL-13, for example, upon antigen-specific stimulation of the CAR. In some embodiments, a certain percentage of the cells in the composition exhibit a less differentiated state and / or express CCR7 + CD27 + CAR + Administered cell compositions containing T cells are associated with desirable or improved pharmacokinetic parameters, safety events, and increased sustainable responses, for example, compared to compositions containing a lower percentage of such cell populations.

[0175] All publications referenced in this application, including patent documents, scientific papers, and databases, are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication were individually incorporated by reference. To the extent that a definition set forth herein contradicts or is otherwise inconsistent with a definition set forth in a patent, patent application, published patent application, or other publication incorporated herein by reference, the definition set forth herein shall take precedence over the definition incorporated herein by reference.

[0176] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0177] I. Dosage and Dose Determination for Therapeutic T Cell Compositions Provided herein are methods, compositions, and articles of manufacture for use in connection with cell therapy, such as engineered T cell therapy, for the treatment of various diseases and conditions, including tumors. Provided embodiments relate to methods for determining a dose to administer to a subject, related methods of treatment, compositions containing doses of cells, and related articles of manufacture and kits. In some aspects, provided embodiments relate to aspects in which the cell phenotype of the cells in the composition for administration, e.g., the presence and / or expression of surface markers, the absence or lack of expression of surface markers, and / or the ability to produce one or more cytokines (e.g., IL-2), is related to, correlated with, or associated with a pharmacokinetic parameter, e.g., the value or measurement of exposure or maximum cell concentration. In some aspects, the phenotype and / or parameter also relates to, correlates with, or is associated with the activity and / or function of the cells and / or the potential for toxicity, such as cytokine release syndrome (CRS) or neurotoxicity (NT), and / or the outcome of the cell therapy in a subject to whom the T cell composition is administered, e.g., response to the cell therapy, e.g., duration of response and progression-free survival.

[0178] In some aspects, provided embodiments involve determining and / or administering a dose of a cell composition, e.g., one or more unit doses of a cell composition and / or an amount corresponding to such a unit dose. In some aspects, the cell dose is enriched for, e.g., has a high or relatively high percentage of, biologically active engineered T cells with a particular phenotype. In some aspects, the dose is based on the number of biologically active engineered T cells with a particular phenotype. In some aspects, "biologically active" refers to the property of cells that are not programmed to undergo cell death, e.g., non-apoptotic cells or cells that do not show signs of entry into the apoptotic pathway. In some aspects, the dose is based on the number of biologically active engineered CD8 T cells with a particular phenotype. +The dose is based on the number of T cells and / or the total number of biologically active engineered T cells having a particular phenotype.

[0179] In some embodiments, the methods ensure that a unit dose contains a relatively consistent number, ratio, ratio, and / or percentage of engineered cells having a particular phenotype in one or more particular compositions. In some aspects, the consistency is associated with or relates to a relatively consistent activity, function, pharmacokinetic parameters, toxicity outcome, and / or response outcome.In some embodiments, the percentage of engineered cells that enable consistent activity, function, pharmacokinetic parameters, toxicity outcomes, and / or response outcomes (e.g., sustainable response, sustainable progression-free survival) is at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60%, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85% of the total number of engineered cells (e.g., T cells) in the composition or the total number of engineered cells (e.g., T cells) in the composition that are surface positive for CCR7 and / or CD27, optionally, and further negative for surface expression of CD45RA. is about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least 94% or at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or At or about 95%, at least 96%, or at least about 96%, or 96% or about 96%, at least 97%, or at least about 97%, or 97% or about 97%, at least 98%, or at least about 98%, or 98% or about 98%, at least 99%, or at least about 99%, or 99% or about 99%, or at least 100%, or at least about 100%, or 100% or about 100%. In some embodiments, the engineered cells (e.g., T cells) are further negative for apoptotic markers such as activated caspase 3 or annexin V.In some embodiments, the percentage of engineered cells that enable consistent activity, function, pharmacokinetic parameters, toxicity outcomes, and / or response outcomes (e.g., sustainable response, sustainable progression-free survival) is at least 75%, or at least about 75%, or 75%, or at least about 75%, or at least 80%, or at least about 80%, or 80% or about 80% of the total number of engineered cells (e.g., T cells) in a composition that are negative for markers of apoptosis (e.g., caspase-3, annexin V), or at least 80%, or at least about 80%, or at least about 80% of the total number of engineered cells (e.g., T cells) in a composition that express a recombinant receptor. at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, or at least 99% or at least about 99%, or 99% or about 99%.In some embodiments, the percentage of engineered cells that enable consistent activity, function, pharmacokinetic parameters, toxicity outcomes, and / or response outcomes (e.g., sustainable response, sustainable progression-free survival) is at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, of the total number of engineered CD4+ T cells in the composition or the total number of engineered CD4+ T cells expressing a recombinant receptor capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha. at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100%.In some embodiments, the percentage of engineered cells that enable consistent activity, function, pharmacokinetic parameters, toxicity outcomes, and / or response outcomes (e.g., sustainable response, sustainable progression-free survival) is determined based on the total number of engineered CD4+ T cells in the composition or the percentage of engineered CD4+ T cells expressing a recombinant receptor that are multifunctional for the production of two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha. At least or at least about 10%, or 10% or about 10%, at least 15%, or 15% or about 15%, at least 20%, or at least about 20%, or 20% or about 20%, at least 25%, or at least about 25%, or 25% or about 25%, at least 30%, or at least about 30%, or 30% or about 30%, at least 40%, or at least about 40%, or 40% or about 40%, at least 50%, or at least about 50%, or 50% or about 50%, at least 60%, or at least about 60%, or 60% or about 60%, or more of the total number of T cells. In some embodiments, the engineered cells of the composition that enable consistent activity, function, pharmacokinetic parameters, toxicity outcomes, and / or response outcomes (e.g., sustainable response, sustainable progression-free survival) are or comprise one or more phenotypes described herein. Thus, in some embodiments, the percentage of engineered cells that allows for consistent activity, function, pharmacokinetic parameters, toxicity outcome, and / or response outcome (e.g., sustainable response, sustainable progression-free survival) is or includes a combination of phenotypic percentages. In some embodiments, a composition to be used as a therapeutic composition is selected based on the percentage of cells comprising one or more phenotypes in the composition (e.g., output composition).

[0180] In some aspects, the number, ratio, proportion and / or percentage of cells in a plurality of subjects, compositions and / or doses is relatively consistent, e.g., has a particular phenotype, e.g., expresses CCR7 (CCR7 + ) and / or the number or ratio of cells capable of producing one or more cytokines varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%. In some aspects, the number or ratio of cells in a composition or unit dose that have a particular phenotype, for example, expressing CCR7 (CCR7 + ) and / or the number or ratio of cells capable of producing one or more cytokines varies by no more than 20%, or no more than 10%, or no more than 5% from the mean of said numbers or ratios among a plurality of T cell compositions produced by the process and / or varies by no more than one standard deviation from such mean, or varies by no more than 20%, or no more than 10%, or no more than 5% among a plurality of T cell compositions or doses determined.

[0181] In some embodiments, the methods involve determining and / or administering one or more unit doses of cells, e.g., engineered T cells, that express a recombinant receptor such as a CAR. In some aspects, the methods involve administering one or more unit doses of cells that are enriched for and / or contain a high or relatively high percentage of cells that have a particular phenotype, such as a particular surface marker phenotype, and / or exhibit a functional activity, such as the ability to produce one or more cytokines. In some aspects, the unit dose is determined based on the number, percentage, ratio, frequency, and / or proportion of engineered T cells, e.g., a subset of cells that have a particular phenotype, such as a particular surface marker phenotype, and / or exhibit a functional activity, such as the ability to produce one or more cytokines. In some embodiments, engineered or therapeutic cell compositions can be produced by methods that result in an output composition that contains or is enriched for cells with such phenotype or functional attribute, e.g., compared to the starting material or input composition used to generate the engineered output composition. Exemplary phenotypic and functional attributes of cells of engineered or therapeutic cell compositions, such as output compositions, produced according to methods for making engineered cells (including in one or more unit doses of any such composition) are described herein.

[0182] In some aspects, the cellular markers include markers indicative of cell health, viability, and / or the apoptotic state of a cell. In some aspects, exemplary markers include CD3, CD4, CD8, CCR7, CD27, CD45RA, Annexin V, or activated caspase 3. In some aspects, exemplary markers include CCR7, CD27, and / or CD45RA. In some aspects, exemplary markers include CCR7 and / or CD27. In some aspects, an exemplary marker is CCR7. In some aspects, an exemplary marker is CD27.

[0183] In some aspects, the provided methods also involve assessing the therapeutic composition for cell phenotype, e.g., expression of markers such as CD3, CD4, CD8, CCR7, CD27, CD45RA, Annexin V, or activated caspase 3. In some embodiments, the phenotype comprises antigen-specific function. In some embodiments, the phenotype comprises cytokine production or secretion, e.g., upon stimulation with an antigen that the recombinant receptor (e.g., CAR) specifically binds to and / or recognizes. In some embodiments, the phenotype comprises production of cytokines associated with a particular cell type, such as cytokines associated with Th1, Th2, Th17, and / or Treg subtypes.

[0184] In some aspects, provided methods involve determining a suitable dose, e.g., one or more unit doses, of a therapeutic composition containing engineered T cells for administration to a subject with a disease or condition. In some aspects, the methods also involve assessing the number, percentage, ratio, frequency, and / or proportion of engineered T cells, e.g., a particular cell subset having a particular phenotype, such as a particular surface marker phenotype, in a therapeutic composition containing engineered T cells. In some embodiments, provided methods ... + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and CCR7 are expressed (receptor + / CCR7 + ) involves assessing the number, percentage, or ratio of T cells.

[0185] In some embodiments, the methods provided involve administering to a subject a plurality of CD8 + and / or CD4 +In a therapeutic composition comprising T cells, the method involves assessing the number, percentage, or ratio of T cells that express or do not express CD27 and / or CD45RA. Other exemplary phenotypic markers for assessment and / or methods for assessment are described below, e.g., in Section IA. In some aspects, a unit dose can be determined based on any assessment. In some aspects, the method also involves assessing the number, percentage, ratio, frequency, and / or proportion of cells that express a recombinant receptor (e.g., a CAR) or a surrogate marker for the recombinant receptor. A dose, e.g., one or more unit doses, can be determined based on an assessment of the number, percentage, ratio, frequency, and / or proportion of any of the phenotypic markers described herein.

[0186] A. Cell phenotyping In some embodiments, a cell phenotype, or the level or percentage of cells of such a phenotype in a composition, correlates with or is associated with the activity and / or function of the cells and / or the likelihood of causing toxicity, such as cytokine release syndrome (CRS) or neurotoxicity (NT), and / or the outcome of the cell therapy in a subject to which the T cell composition is administered, e.g., response to the cell therapy, e.g., duration of response and progression-free survival. In some embodiments, a composition comprising cells with a specific phenotype, more particularly the percentage of cells with such a specific phenotype, correlates with a clinical outcome, such as a durable response and / or progression-free survival. In some embodiments, the phenotype is determined by assessing the presence or absence of one or more specific molecules, including surface molecules and / or molecules that may accumulate or be produced by cells or subpopulations of cells within the T cell composition. In some embodiments, the phenotype directly or inversely indicates or is indicative of the biological activity of a cell or population of cells within the T cell composition. In some embodiments, the phenotype may include a cellular activity, such as the production of a factor (e.g., a cytokine) in response to a stimulus. In certain embodiments, the assessment of cellular composition is performed to identify, detect, or quantify the phenotype of the cellular composition, hi certain embodiments, the measurement of cellular composition is performed to identify, detect, or quantify the presence, absence, degree, or level of expression of a specific molecule.

[0187] It is contemplated that the phenotype of a cell composition (e.g., an engineered T cell composition) may, in some cases, depend on many factors, including, but not limited to, the phenotype of the starting cellular material (e.g., an apheresis product or leukapheresis product) used to generate the cell composition and the phenotype of the cellular material that undergoes processing to generate the cell composition. Thus, in some embodiments, phenotype is assessed in the starting material and / or material that undergoes processing to generate the cell composition, as well as in the cells of the final cell composition.

[0188] In some embodiments, the phenotype is indicative of cell viability. In some embodiments, the phenotype is indicative of the absence of apoptosis, the absence of early stages of apoptosis, or the absence of late stages of apoptosis. In some embodiments, the phenotype is the absence of a factor ... recombinant receptor-expressing T cells (e.g., CAR + T cells), CD8 + T cells or CD4 + The phenotype is a phenotype of a subpopulation or subset of T cells, such as T cells. In some embodiments, the phenotype is that of non-activated cells and / or cells lacking or having reduced or low expression of one or more activation markers. In some embodiments, the phenotype is that of non-depleted cells and / or cells lacking or having reduced or low expression of one or more depletion markers.

[0189] In some embodiments, a phenotype is indicated by the presence, absence, or level of expression in cells of one or more specific molecules, such as a specific surface marker that indicates the phenotype, for example, a surface protein, an intracellular marker that indicates the phenotype, or a nucleic acid that indicates the phenotype, or other molecules or factors that indicate the phenotype. In some embodiments, a phenotype is or includes one or more positive or negative expression of specific molecules. In some embodiments, specific molecules include, but are not limited to, surface markers, such as membrane glycoproteins or receptors; markers associated with apoptosis or viability; or specific molecules that indicate the state of immune cells, such as markers associated with activated, exhausted, or mature, or naive phenotypes. In some embodiments, the number of cells of a phenotype can be determined using any known method for evaluating or measuring, counting, and / or quantifying cells based on specific molecules.

[0190] In some embodiments, the phenotype is or includes positive or negative expression of one or more specific molecules in a cell. In some embodiments, positive expression is indicated by a detectable amount of a specific molecule in a cell. In certain embodiments, a detectable amount is any detected amount of a specific molecule in a cell. In certain embodiments, a detectable amount is an amount greater than background, e.g., background staining, signal, etc., in a cell. In certain embodiments, positive expression is an amount of a specific molecule greater than a threshold, e.g., a predetermined threshold. Similarly, in certain embodiments, a cell having negative expression of a specific molecule can be any cell not determined to have positive expression, or a cell lacking a detectable amount of the specific molecule or a detectable amount of the specific molecule above background. In some embodiments, a cell has negative expression of a specific molecule if the amount of the specific molecule is below a threshold. Those skilled in the art will understand how to define a threshold for defining positive and / or negative expression for a specific molecule as a routine technique, and the threshold can be defined according to, for example, but not limited to, the assay or method of detection, the identity of the specific molecule, the reagents used for detection, and the specific parameters of the instrument.

[0191] Examples of methods that can be used to detect specific molecules and / or analyze cellular phenotypes include, but are not limited to, biochemical analysis; immunochemical analysis; image analysis; cell morphological analysis; molecular analysis such as PCR, sequencing, high-throughput sequencing, and DNA methylation determination; proteomic analysis such as protein glycosylation and / or phosphorylation pattern determination; genomic analysis; epigenomic analysis (e.g., ChIP-seq or ATAC-seq); transcriptomic analysis (e.g., RNA-seq); and any combination thereof. In some embodiments, the method can include evaluation of immune receptor repertoires, such as T cell receptor (TCR) repertoires. In some aspects, any phenotypic determination can be evaluated by high-throughput automated methods and / or single-cell-based methods. In some aspects, large-scale or genome-wide methods can be used to identify one or more molecular signatures. In some aspects, large-scale or genome-wide methods can be used to identify molecular signatures associated with therapy outcomes, such as efficacy and safety, or pharmacokinetic parameters. In some aspects, one or more molecular signatures can be determined, for example, the expression of specific RNA or protein in cells.In some embodiments, the molecular characteristics of phenotype are analyzed by image analysis, PCR (including standard PCR and all variants of PCR), microarray (including but not limited to DNA microarray, MM chip for microRNA, protein microarray, cell microarray, antibody microarray and carbohydrate array), sequencing, biomarker detection, or the method for determining DNA methylation or protein glycosylation pattern.In certain embodiments, specific molecule is polypeptide, i.e., protein.In some embodiments, specific molecule is polynucleotide.

[0192] In some embodiments, positive or negative expression of a specific molecule identifies cells as positively or negatively selected for one or more surface markers (markers) expressed on the selected cells, respectively. +) or one or more surface markers expressed at relatively high levels (markers high Expression is determined by incubating with one or more antibodies or other binding agents that specifically bind to the specific marker. In certain embodiments, positive or negative expression is determined by flow cytometry, immunohistochemistry, or any other suitable method for detecting specific markers.

[0193] In certain embodiments, the expression of specific molecules is assessed by flow cytometry, a laser- or impedance-based biophysical technique used in cell counting, cell sorting, biomarker detection, and protein engineering by suspending cells in a fluid stream and passing them past an electronic detection device. This allows for simultaneous multiparametric analysis of the physicochemical properties of up to thousands of particles per second.

[0194] Data generated by a flow cytometer can be plotted in one dimension to create histograms, or in two-dimensional dot plots, or even in three dimensions. Regions on these plots can be sequentially separated based on fluorescence intensity by creating a series of subset extracts called "gates." Specific gating protocols exist for diagnostic and clinical purposes, particularly in the context of immunology. Plots are often generated on a logarithmic scale. Because the emission spectra of different fluorochromes overlap, the signals at the detector must be corrected electronically and computationally. Data accumulated using a flow cytometer can be analyzed using software such as JMP (statistical software), WinMDI, Flow Software, and the web-based Cytobank, Cellcion, FCS Express, FlowJo, FACSDiva, CytoPaint (aka Paint-A-Gate), VenturiOne, CellQuest Pro, Infinicyt, or Cytospec.

[0195] Flow cytometry is a standard technique in the art, and one of skill in the art would readily understand how to design or adapt a protocol to detect one or more specific molecules and to analyze the data to determine the expression of the one or more specific molecules in a population of cells. Standard protocols and techniques for flow cytometry can be found in Lloyd "Flow Cytometry in Microbiology; Practical Flow Cytometry by Howard M. Shapiro; Flow Cytometry for Biotechnology by Larry A. Sklar; Handbook of Flow Cytometry Methods by J. Paul Robinson, et al., Current Protocols in Cytometry, Wiley-Liss Pub; Flow Cytometry in Clinical Diagnosis, v4, (Carey, McCoy, and Keren, eds), ASCP Press, 2007; Ormerod, MG (ed.) (2000) Flow Cytometry - A practical approach. 3rd edition. Oxford University Press, Oxford, UK; Ormerod, MG (1999) Flow Cytometry. 2nd edition. BIOS Scientific Publishers, Oxford.; and Flow Cytometry - A basic introduction. Michael G. Ormerod, 2008.

[0196] In some embodiments, cells are sorted by phenotype for further analysis. In some embodiments, cells of different phenotypes in the same cell composition are sorted by fluorescence-activated cell sorting (FACS). FACS is a specialized form of flow cytometry that allows heterogeneous cell mixtures to be sorted into two or more containers, one cell at a time, based on the specific light scattering and fluorescence properties of each cell. This is a useful scientific instrument because it provides fast, objective, and quantitative recording of the fluorescent signals from individual cells and physical separation of specific cells of interest.

[0197] In some embodiments, the phenotype is determined by measuring the total T cell count or the total CD3 + The phenotype may include the number of T cells. In certain embodiments, a T cell composition, e.g., a therapeutic T cell composition, containing cells expressing a recombinant receptor or CAR may include one or more different subtypes of T cells. In some embodiments, the phenotype is or includes the identity of a T cell subtype. Different populations or subtypes of T cells include effector T cells, helper T cells, memory T cells, regulatory T cells, naive T cells, CD4 + cells and CD8 + In certain embodiments, T cell subtypes can be identified by detecting the presence or absence of specific molecules. In certain embodiments, the specific molecules are surface markers that can be used to identify T cell subtypes.

[0198] In some embodiments, the phenotype is a positive or high level expression of one or more specific molecules that are surface markers, e.g., CD3, CD4, CD8, CD28, CD62L, CCR7, CD27, CD127, CD4, CD8, CD45RA, and / or CD45RO. In certain embodiments, the phenotype is a surface marker of a T cell or of a subpopulation or subset of T cells, e.g., based on positive surface marker expression of one or more surface markers, e.g., CD3 + , CD4 + , CD8 + , CD28 + , CD62L+ , CCR7 + , CD27 + , CD127 + , CD4 + , CD8 + , CD45RA + and / or CD45RO + In some embodiments, the phenotype is positive or high level expression of one or more specific molecules that are surface markers, e.g., CC chemokine receptor type 7 (CCR7), cluster of differentiation 27 (CD27), cluster of differentiation 28 (CD28), and cluster of differentiation 45RA (CD45RA). In certain embodiments, the phenotypic markers include CCR7, CD27, CD28, CD44, CD45RA, CD62L, and L-selectin. In some embodiments, the phenotype is negative or absent expression of one or more specific molecules that are surface markers, e.g., CD3, CD4, CD8, CD28, CD62L, CCR7, CD27, CD127, CD4, CD8, CD45RA, and / or CD45RO. In certain embodiments, the phenotype is negative or absent expression of a surface marker of a T cell or of a subpopulation or subset of T cells, e.g., based on the absence of surface marker expression of one or more surface markers, e.g., CD3 - , CD4 - , CD8-, CD28 - , CD62L - , CCR7 - , CD27 - , CD127 - , CD4 - , CD8 - , CD45RA - and / or CD45RO - In some embodiments, the phenotype is negative or absent expression of one or more specific surface markers, such as CC chemokine receptor type 7 (CCR7), cluster of differentiation 27 (CD27), cluster of differentiation 28 (CD28), and cluster of differentiation 45RA (CD45RA). In certain embodiments, the phenotypic markers include CCR7, CD27, CD28, CD44, CD45RA, CD62L, and L-selectin.

[0199] In certain embodiments, the phenotype is or includes positive or negative expression of CD27, CCR7 and / or CD45RA. + In some embodiments, the phenotype is CD27 + In some embodiments, the phenotype is CCR7 + / CD27 + In some embodiments, the phenotype is CD45RA - In some embodiments, the phenotype is CCR7 + / CD45RA - In some embodiments, the phenotype is CD27 + / CD45RA - In some embodiments, the phenotype is CCR7 + / CD27 + / CD45RA - is.

[0200] In certain embodiments, the surface marker indicates the expression of a recombinant receptor, e.g., a CAR. In certain embodiments, the surface marker is the expression of a recombinant receptor, e.g., a CAR, which in some aspects can be determined using an antibody such as an anti-idiotype antibody. In some embodiments, the surface marker indicating the expression of a recombinant receptor is a surrogate marker. In certain embodiments, such a surrogate marker is a surface protein that has been modified to have little activity. In certain embodiments, the surrogate marker is encoded on the same polynucleotide that encodes the recombinant receptor. In some embodiments, the nucleic acid sequence encoding the recombinant receptor is operably linked to the nucleic acid sequence encoding the marker, optionally separated by an internal ribosome entry site (IRES) or a 2A sequence such as a nucleic acid encoding a self-cleaving peptide or a peptide that causes ribosome skipping, e.g., T2A (e.g., SEQ ID NOs:6 and 17), P2A (e.g., SEQ ID NO:18 and 19), E2A (e.g., SEQ ID NO:20), or F2A (e.g., SEQ ID NO:21). In some cases, exogenous marker genes may be utilized in conjunction with engineered cells to allow for cell detection or selection, and in some cases to promote cell suicide.

[0201] Exemplary surrogate markers can include truncated cell surface polypeptides, such as truncated human epidermal growth factor receptor 2 (tHER2), truncated epidermal growth factor receptor (EGFRt, exemplary EGFRt sequences shown in SEQ ID NO: 7 or 16), or prostate-specific membrane antigen (PSMA) or modified forms thereof. EGFRt may contain an epitope recognized by the antibody cetuximab (Erbitux®) or other therapeutic anti-EGFR antibodies or binding molecules, which can be used to identify or select cells engineered with EGFRt constructs and recombinant receptors, such as chimeric antigen receptors (CARs), and / or to eliminate or isolate cells expressing the receptor. See U.S. Patent No. 8,802,374 and Liu et al., Nature Biotech. 2016 April; 34(4): 430-434. In some aspects, markers, e.g., surrogate markers, include all or a portion (e.g., a truncated form) of CD34, NGFR, or epidermal growth factor receptor (e.g., tEGFR). In some embodiments, the nucleic acid encoding the marker is operably linked to a linker sequence, e.g., a cleavable linker sequence, e.g., a polynucleotide encoding T2A. For example, the marker, and optionally the linker sequence, can be any as disclosed in PCT Publication No. WO2014031687. For example, the marker can be a truncated EGFR (tEGFR), optionally linked to a linker sequence, such as a T2A cleavable linker sequence. Exemplary polypeptides for truncated EGFR (e.g., tEGFR) include the sequence of amino acids set forth in SEQ ID NO: 7 or 16, or a sequence of amino acids that exhibits at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to SEQ ID NO: 7 or 16.

[0202] In some embodiments, the marker is or comprises a fluorescent protein such as green fluorescent protein (GFP), enhanced green fluorescent protein (EGFP), e.g., superfold GFP, red fluorescent protein (RFP), e.g., tdTomato, mCherry, mStrawberry, AsRed2, DsRed or DsRed2, cyan fluorescent protein (CFP), blue-green fluorescent protein (BFP), enhanced blue fluorescent protein (EBFP), and yellow fluorescent protein (YFP), and variants thereof (including species variants, monomeric variants, and codon-optimized and / or enhanced variants of fluorescent proteins). In some embodiments, the marker is or comprises an enzyme such as luciferase, the lacZ gene from Escherichia coli (E. coli), alkaline phosphatase, secreted embryonic alkaline phosphatase (SEAP), chloramphenicol acetyltransferase (CAT), etc. Exemplary luminescent reporter genes include luciferase (luc), β-galactosidase, chloramphenicol acetyltransferase (CAT), β-glucuronidase (GUS), or variants thereof.

[0203] In certain embodiments, the phenotype includes expression, e.g., surface expression, of one or more surface markers CD3, CD4, CD8, and / or a recombinant receptor (e.g., a CAR) or a surrogate marker thereof that is indicative of or correlates with expression of the recombinant receptor (e.g., a CAR).

[0204] In certain embodiments, the phenotype is determined by the expression of one or more specific molecules that are surface markers. In certain embodiments, the phenotype is or includes the positive or negative expression of CD3, CD4, CD8 and / or recombinant receptors, such as CAR. In certain embodiments, the recombinant receptor is CAR. In certain embodiments, the phenotype is determined by the expression of CD3 + / CAR + , CD4 + / CAR + and / or CD8 + / CAR + Includes.

[0205] In certain embodiments, the phenotype is or comprises positive or negative expression of CD27, CCR7 and / or CD45RA, and / or recombinant receptors, such as CAR. In some embodiments, the phenotype is or comprises positive or negative expression of CCR7 + / CAR + In some embodiments, the phenotype is CD27 + / CAR + In some embodiments, the phenotype is CCR7 + / CD27 + / CAR + In some embodiments, the phenotype is CD45RA - / CAR + In some embodiments, the phenotype is CCR7 + / CD45RA - / CAR + In some embodiments, the phenotype is CD27 + / CD45RA - / CAR + In some embodiments, the phenotype is CCR7 + / CD27 + / CD45RA - / CAR + is.

[0206] In some embodiments, the phenotype is viability. In certain embodiments, the phenotype is the positive expression of a marker indicating that the cell is undergoing normal functional cellular processes and / or is not undergoing necrosis or programmed cell death or is not in the process of undergoing necrosis or programmed cell death. In some embodiments, viability can be assessed by the redox potential of the cell, the integrity of the cell membrane, or the activity or function of mitochondria. In some embodiments, viability is the absence of specific molecules associated with cell death in the assay, or the absence of signs of cell death.

[0207] In some embodiments, the phenotype is or includes cell viability. In certain embodiments, cell viability can be detected, measured, and / or assessed by a number of means routine in the art. Non-limiting examples of such viability assays include, but are not limited to, dye uptake assays (e.g., calcein AM assay), XTT cell viability assays, and dye exclusion assays (e.g., trypan blue, eosin, or propidium dye exclusion assays). Viability assays are useful for determining cell dose, cell composition, and / or the number or percentage (e.g., frequency) of viable cells in a cell sample. In certain embodiments, the phenotype includes cell viability along with other characteristics, such as recombinant receptor expression.

[0208] In certain embodiments, the phenotype is cell viability, viable CD3 + , viable CD4 + , viable CD8 + , viable CD3 + / CAR + , viable CD4 + / CAR + , viable CD8 + / CAR + , viable CD4 + / CCR7 + / CAR + , viable CD8 + / CD27 + / CAR + , viable CD4 + / CD27 + / CAR + , viable CD8 + / CCR7 + / CD27 + / CAR + , viable CD4 + / CCR7 + / CD27 + / CAR + , viable CD8 + / CCR7 + / CD45RA - / CAR + or viable CD4 + / CCR7 + / CD45RA - or a combination thereof.

[0209] In certain embodiments, the phenotype is the absence or inclusion of apoptosis and / or signs that the cell is undergoing the apoptotic process. Apoptosis is a process of programmed cell death that involves a series of typical morphological and biochemical events that lead to characteristic cellular changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, chromosomal DNA fragmentation, and extensive mRNA degradation. Apoptosis is a well-characterized process, and specific molecules associated with various stages are well known in the art.

[0210] In some embodiments, the phenotype is the absence of early stages of apoptosis and / or the absence of indicators and / or specific molecules associated with the early stages of apoptosis. The early stages of apoptosis are characterized by changes in the cell membrane and mitochondrial membrane. Biochemical changes also occur in the cell cytoplasm and nucleus. For example, the early stages of apoptosis can be indicated by the activation of certain caspases, such as 2, 8, 9, and 10. In certain embodiments, the phenotype is the absence of late stages of apoptosis and / or the absence of indicators and / or specific molecules associated with the late stages of apoptosis. The mid-to-late stages of apoptosis are characterized by further loss of membrane integrity, chromatin condensation, and DNA fragmentation, and include biochemical events such as the activation of caspases 3, 6, and 7.

[0211] In certain embodiments, the phenotype is negative expression of one or more factors associated with apoptosis, including pro-apoptotic factors known to initiate apoptosis, e.g., members of the death receptor pathway, activated members of the mitochondrial (intrinsic) pathway, e.g., Bcl-2 family members (e.g., Bax, Bad, and Bid), and caspases. In some embodiments, the phenotype is negative or low levels of a marker of apoptosis. In certain embodiments, the phenotype is negative expression of a marker of apoptosis. In certain embodiments, the phenotype is the absence of an indicator, e.g., Annexin V molecule, that preferentially binds to cells undergoing apoptosis when incubated or contacted with the cell composition. In some embodiments, the phenotype is or includes expression of one or more markers indicative of an apoptotic state in a cell.

[0212] In some embodiments, the phenotype is negative (or low) expression of specific molecules that are markers for apoptosis.Various apoptosis markers are known to those skilled in the art, and include, but are not limited to, the increase in the activity of one or more caspases, i.e., activated caspases (e.g., activated caspases), the increase in PARP cleavage, the activation and / or translocation of Bcl-2 family proteins that are members of cell death pathways, such as Fas and FADD, the presence of nuclear shrinkage (e.g., monitored by microscopy) and the presence of chromosomal DNA fragmentation (e.g., the presence of chromosomal DNA ladders), or by apoptosis assays including TUNEL staining and Annexin V staining.

[0213] Caspases are enzymes that cleave proteins after aspartic acid residues, and the term derives from "cysteine-aspartic acid protease." Caspases are involved in apoptosis, and thus activation of caspases, such as caspase-3, is an indicator of increased or resurrected apoptosis. In certain embodiments, caspase activation can be detected by methods known to those skilled in the art. In some embodiments, caspase activation can be detected using antibodies that specifically bind to activated caspases (i.e., specifically bind to the cleaved polypeptide). In another example, a fluorochrome inhibitor of caspase activity (FLICA) assay can be used to detect caspase-3 activation by detecting the hydrolysis of acetylAsp-Glu-Val-Asp 7-amido-4-methylcoumarin (Ac-DEVD-AMC) by caspase-3 (i.e., detecting the release of fluorescent 7-amino-4-methylcoumarin (AMC)). FLICA assays can be used to determine caspase activation by detecting the products of substrates processed by multiple caspases (e.g., FAM-VAD-FMK FLICA). Other techniques include the CASPASE-GLO® Caspase Assay (PROMEGA), which uses luminogenic caspase-8 tetrapeptide substrates (Z-LETD-aminoluciferin), caspase-9 tetrapeptide substrates (Z-LEHD-aminoluciferin), caspase-3 / 7 substrates (Z-DEVD-aminoluciferin), caspase-6 substrates (Z-VEID-aminoluciferin), or caspase-2 substrates (Z-VDVAD-aminoluciferin).

[0214] In certain embodiments, the phenotype is or comprises negative expression in the cell of activated caspase-1, activated caspase-2, activated caspase-3, activated caspase-7, activated caspase-8, activated caspase-9, activated caspase-10, and / or activated caspase-13. -In some embodiments, surrogate (cleaved zymogen) forms of caspases (e.g., any of those listed above) are also markers that indicate the presence of apoptosis. In some embodiments, the phenotype is or includes the absence or negative expression of surrogate forms of caspases, such as surrogate forms of caspase-3.

[0215] In some embodiments, the marker of apoptosis is cleaved by poly ADP-ribose polymerase 1 (PARP). PARP is cleaved by caspases during the early stages of apoptosis. Therefore, detection of cleaved PARP peptide is a marker for apoptosis. In certain embodiments, the phenotype is or includes the positive or negative expression of cleaved PARP.

[0216] In some embodiments, the apoptosis marker is a reagent that detects a cellular feature associated with apoptosis. In certain embodiments, the reagent is an annexin V molecule. During the early stages of apoptosis, the lipid phosphatidylserine (PS) translocates from the inner to the outer leaflet of the plasma membrane. PS is usually confined to the inner membrane in healthy and / or non-apoptotic cells. Annexin V is a protein that preferentially binds to phosphatidylserine (PS) with high affinity. When conjugated to a fluorescent tag or other reporter, annexin V can be used to rapidly detect this early cell surface indicator of apoptosis. In some embodiments, the presence of PS on the outer membrane persists until the later stages of apoptosis. Thus, in some embodiments, annexin V staining is an indication of both the early and late stages of apoptosis. In certain embodiments, annexin, such as annexin V, is tagged with a detectable label and incubated with, exposed to, and / or contacted with cells of a cell composition to detect cells undergoing apoptosis, for example, by flow cytometry. In some embodiments, fluorescently tagged annexins, e.g., annexin V, are used to tag cells for flow cytometry analysis, e.g., annexin V. - V / 7 -Staining is performed using the AAD assay. Alternative protocols suitable for detecting apoptosis using annexins include techniques and assays that utilize radiolabeled annexin V. In certain embodiments, the phenotype is determined by negative staining with annexins, e.g., annexin V. - In certain embodiments, the phenotype is or comprises the absence of PS on the outer plasma membrane. In certain embodiments, the phenotype is or comprises cells to which annexin, e.g., annexin V, is not bound. In certain embodiments, cells lacking detectable PS on the outer membrane are characterized by the absence of annexin V. - In certain embodiments, after incubation with labeled annexin V, annexin V is detected in an assay, e.g., flow cytometry. - Cells that do not bind annexin V - is.

[0217] In certain embodiments, the phenotype is determined by Annexin V - , annexin V - CD3 + , annexin V - CD4 + , annexin V - CD8 + , annexin V - CD3 + / CAR + , annexin V - CD4 + / CAR + , annexin V - CD8 + / CAR + , activated caspase 3 - , activated caspase 3 - / CD3 + , activated caspase 3 - / CD4 + , activated caspase 3 - / CD8 + , activated caspase 3 - / CD3 + / CAR + , activated caspase 3 - / CD4 + / CAR + , activated caspase 3- / CD8 + / CAR + , annexin V - / CD4 + / CCR7 + / CAR + , annexin V - / CD8 + / CD27 + / CAR + , Annexin V / CD4 + / CD27 + / CAR + , annexin V - / CD8 + / CCR7 + / CD27 + / CAR + , Annexin V / CD4 + / CCR7 + / CD27 + / CAR + , annexin V - / CD8 + / CCR7 + / CD45RA - / CAR + or Annexin V- / CD4 + / CCR7 + / CD45RA - ; Activated caspase 3 - / CD4 + / CCR7 + / CAR + , activated caspase-3 / CD8 + / CD27 + / CAR + , activated caspase 3 - / CD4 + / CD27 + / CAR + , activated caspase 3 - / CD8 + / CCR7 + / CD27 + / CAR + , activated caspase 3 - / CD4 + / CCR7 + / CD27 + / CAR + , activated caspase 3 - / CD8+ / CCR7 + / CD45RA - / CAR + or activated caspase 3 - / CD4 + / CCR7 + / CD45RA - , or a combination thereof.

[0218] Certain embodiments contemplate that cells positive for expression of markers for apoptosis are undergoing programmed cell death, exhibit reduced or absent immune function, and have a diminished ability, if any, to undergo activation, expansion, and / or bind antigens to initiate, carry out, or contribute to an immune response or activity. In certain embodiments, the phenotype is defined by negative expression for activated caspases and / or negative staining with Annexin V.

[0219] In certain embodiments, the phenotype is activated caspase 3 - (Caspase 3 - ) and / or annexin V - is or contains

[0220] Among other things, a phenotype is one or more subtypes or subpopulations of T cells, or the expression or surface expression of one or more markers generally associated with those phenotypes. T cell subtypes and subpopulations include naive T (T N ) cells, effector T cells (T EFF ), memory T cells and their subtypes, such as stem cell memory T (T SCM ), Central Memory T(T CM ), Effector Memory T(T EM ), T EMRACD4 T cells or terminally differentiated effector memory T cells, tumor infiltrating lymphocytes (TIL), immature T cells, mature T cells, helper T cells, cytotoxic T cells, mucosal-associated invariant T (MAIT) cells, naturally occurring adaptive regulatory T (Treg) cells, helper T cells, such as TH1 cells, TH2 cells, TH3 cells, TH17 cells, TH9 cells, TH22 cells, follicular helper T cells, alpha / beta T cells, and delta / gamma T cells. + and / or CD8 + It may include T cells as well as their subtypes.

[0221] In some aspects, the phenotype includes expression or markers or functions, e.g., antigen-specific functions, such as cytokine secretion, associated with less differentiated or more differentiated cell subsets. + , CD27 + and interleukin-2 (IL-2) production. In some aspects, the less differentiated subset may also be associated with therapeutic efficacy, self-renewal, survival function, or graft-versus-host disease. In some embodiments, the phenotype is associated with the more differentiated subset, such as one or more of interferon-gamma (IFN-γ) or IL-13 production. In some aspects, the more differentiated subset may also be associated with senescence and effector function.

[0222] In some embodiments, the phenotype is or comprises the phenotype of memory T cells or memory T cell subsets exposed to their cognate antigen. CM cell, T EM Cells or T EMRA cell, T SCMThe phenotype is or comprises a phenotype of a memory T cell (or one or more markers associated therewith), such as a memory T cell, a T cell, or a combination thereof. In certain embodiments, the phenotype is or comprises the expression of one or more specific molecules that are markers for memory and / or T cell or subtypes thereof. In some aspects, the T CM An exemplary phenotype associated with cells is CD45RA - , CD62L + , CCR7 + and CD95 + In some aspects, the compound may include one or more of the following: EM An exemplary phenotype associated with cells is CD45RA - , CD62L - , CCR7 - and CD95 + may include one or more of:

[0223] In certain embodiments, the phenotype is or comprises the expression of one or more specific molecules that are markers for naive T cells.

[0224] In some embodiments, the phenotype is or comprises a memory T cell or a naive T cell. In certain embodiments, the phenotype is the positive or negative expression of one or more specific molecules that are markers for memory. In some embodiments, memory markers are specific molecules that can be used to define a memory T cell population.

[0225] In some embodiments, the phenotype is or includes one or more markers associated with non-memory T cells or subtypes thereof; in some aspects, it is or includes a phenotype or marker associated with naive cells. In some embodiments, the phenotype is CCR7 + / CD27 + / CD28 + / CD45RA + In certain embodiments, the phenotype is CCR7 + / CD45RA +In some embodiments, the phenotype is or comprises a central memory T cell phenotype. In certain embodiments, the phenotype is or comprises a CCR7 + / CD27 + / CD28 + / CD45RA - In some embodiments, the phenotype is or comprises CCR7 - / CD27 + / CD28 + / CD45RA - In certain embodiments, the phenotype is or comprises T EMRA Cells or T SCM In certain embodiments, the phenotype is of or comprises a cell. + In certain embodiments, the phenotype is or comprises CCR7 - / CD27 - / CD28 - / CD45RA + In some embodiments, the phenotype is or comprises CD27 + / CD28 + , CD27 - / CD28 + , CD27 + / CD28 - or CD27 - / CD28 - In some embodiments, the phenotype is or comprises one of: CCR7 + / CD27 + / CD45RA + In certain embodiments, the phenotype is CCR7 + / CD45RA + In certain embodiments, the phenotype is or comprises CCR7 + / CD27 + / CD45RA - In some embodiments, the phenotype is or comprises CCR7 - / CD27 + / CD45RA - In certain embodiments, the phenotype is or comprises CD45RA +In certain embodiments, the phenotype is or comprises CCR7 - / CD27 - / CD45RA + is or contains

[0226] In some embodiments, the phenotype is or comprises any of the aforementioned phenotypic characteristics and further comprises expression of a recombinant receptor, such as a phenotype associated with a memory T cell or memory subtype and expressing a CAR, or a phenotype associated with a naive cell expressing a CAR. In certain embodiments, the phenotype is that of a central memory T cell or stem central memory T cell expressing a CAR. In certain embodiments, the phenotype is that of an effector memory cell expressing a CAR. In some embodiments, the phenotype is that of a T cell expressing a CAR. EMRA In certain embodiments, the phenotype is of or comprises a CAR + / CCR7 + / CD27 + / CD28 + / CD45RA - ;CAR + / CCR7 - / CD27 + / CD28 + / CD45RA - ;CAR + / CCR7 - / CD27 - / CD28 - / CD45RA + ;CAR + / CD27 + / CD28 + ;CAR + / CD27 - / CD28 + ;CAR + / CD27 + / CD28 - ; or CAR + / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises a CAR + / CCR7 + / CD27+ / CD45RA - ;CAR + / CCR7 - / CD27 + / CD45RA - ;CAR + / CCR7 - / CD27 - / CD28 - / CD45RA + ;CAR + / CD27 + ;CAR + / CD27 - ;CAR + / CD27 + / CD28 - ; or CAR + / CD27 - / CD28 - is or contains

[0227] In certain embodiments, the phenotype is or comprises a T cell phenotype that is negative for a marker of apoptosis. In certain embodiments, the phenotype is or comprises a naive cell phenotype that is negative for a marker of apoptosis. In some embodiments, the marker of apoptosis is activated caspase 3. In some embodiments, the marker of apoptosis is positive staining with Annexin V.

[0228] In certain embodiments, the phenotype is of or comprises memory T cells or subtypes thereof that are negative for markers of apoptosis that express a CAR. In certain embodiments, the phenotype is of or comprises memory T cells or particular subtypes that are negative for markers of apoptosis that express a CAR. In certain embodiments, the phenotype is of or comprises naive cells that are negative for markers of apoptosis that express a CAR. In certain embodiments, the phenotype is of central memory T cells or T that are negative for markers of apoptosis that express a CAR. SCMThe phenotype is of or comprises an effector memory cell that expresses a CAR and is negative for a marker of apoptosis. In certain embodiments, the ... - / CAR + / CCR7 + / CD27 + / CD28 + / CD45RA - ;Annexin V - / CAR + / CCR7 - / CD27 + / CD28 + / CD45RA - ;Annexin V - / CAR + / CCR7 - / CD27 - / CD28 - / CD45RA + ;Annexin V - / CAR + / CD27 + / CD28 + ;Annexin V - / CAR + / CD27 - / CD28 + ;Annexin V - / CAR + / CD27 + / CD28 - or Annexin V - / CAR + / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises activated caspase 3 - / CAR + / CCR7 + / CD27 + / CD28 + / CD45RA - ; Activated caspase 3 - / CAR + / CCR7 - / CD27 + / CD28 + / CD45RA -; Activated caspase 3 - / CAR + / CCR7 - / CD27 - / CD28 - / CD45RA + ; Activated caspase 3 - / CAR + / CD27 + / CD28 + ; Activated caspase 3 - / CAR + / CD27 - / CD28 + ; Activated caspase 3 - / CAR + / CD27 + / CD28 - or activated caspase 3 - / CAR + / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises annexin V. - / CAR + / CCR7 + / CD27 + / CD45RA - ;Annexin V - / CAR + / CCR7 - / CD27 + / CD45RA - ;Annexin V - / CAR + / CCR7 - / CD27 - / CD45RA + ;Annexin V - / CAR + / CD27 + / CD28 + ;Annexin V - / CAR + / CD27 - / CD28 + ;Annexin V - / CAR + / CD27 + or Annexin V - / CAR + / CD27 -In certain embodiments, the phenotype is or comprises activated caspase 3 - / CAR + / CCR7 + / CD27 + / CD45RA - ; Activated caspase 3 - / CAR + / CCR7 - / CD27 + / CD45RA - ; Activated caspase 3 - / CAR + / CCR7 - / CD27 - / CD45RA + ; Activated caspase 3 - / CAR + / CD27 + / CD28 + ; Activated caspase 3 - / CAR + / CD27 - / CD28 + ; Activated caspase 3 - / CAR + / CD27 + or activated caspase 3 - / CAR + / CD27 - is or contains

[0229] In certain embodiments, the phenotype is CD27 + / CD28 + , CD27 - / CD28 + , CD27 + / CD28 - , CD27 - / CD28 - In some embodiments, the phenotype is or comprises a CAR + / CD27 + / CD28 + , CAR + / CD27 - / CD28 + , CAR + / CD27 + / CD28 - , CAR+ / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises activated caspase 3, or a combination thereof. - / CAR + / CD27 + / CD28 + , activated caspase 3 - / CAR + / CD27 - / CD28 + , activated caspase 3 - / CAR + / CD27 + / CD28 - , activated caspase 3 - / CAR + / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises annexin V, - / CAR + / CD27 + / CD28 + , annexin V - / CAR + / CD27 - / CD28 + , annexin V - / CAR + / CD27 + / CD28 - , annexin V - / CAR + / CD27 - / CD28 - In certain embodiments, the phenotype is or comprises CD27 + , CD27 - , CD27 + , CD27 - In some embodiments, the phenotype is or comprises a CAR + / CD27 + , CAR + / CD27 - , CAR + / CD27 + , CAR + / CD27- In certain embodiments, the phenotype is or comprises activated caspase 3, or a combination thereof. - / CAR + / CD27 + , activated caspase 3 - / CAR + / CD27 - , activated caspase 3 - / CAR + / CD27 + , activated caspase 3 - / CAR + / CD27 - In certain embodiments, the phenotype is or comprises annexin V, - / CAR + / CD27 + , annexin V - / CAR + / CD27 - , annexin V - / CAR + / CD27 + , annexin V - / CAR + / CD27 - or a combination thereof.

[0230] In certain embodiments, the phenotype is CCR7 + / CD28 + , CCR7 - / CD28 + , CCR7 + / CD28 - , CCR7 - / CD28 - In some embodiments, the phenotype is or comprises a CAR + / CCR7 + / CD28 + , CAR + / CCR7 - / CD28 + , CAR + / CCR7 + / CD28 - , CAR + / CCR7 - / CD28- In certain embodiments, the phenotype is or comprises activated caspase 3, or a combination thereof. - / CAR + / CCR7 + / CD28 + , activated caspase 3 - / CAR + / CCR7 - / CD28 + , activated caspase 3 - / CAR + / CCR7 + / CD28 - , activated caspase 3 - / CAR + / CCR7 - / CD28 - In certain embodiments, the phenotype is or comprises annexin V, - / CAR + / CCR7 + / CD28 + , annexin V - / CAR + / CCR7 - / CD28 + , annexin V - / CAR + / CCR7 + / CD28 - , annexin V - / CAR + / CCR7 - / CD28 - In certain embodiments, the phenotype is or comprises CCR7 + , CCR7 - , CCR7 + , CCR7 - In some embodiments, the phenotype is or comprises a CAR + / CCR7 + , CAR + / CCR7 - , CAR + / CCR7 + , CAR + / CCR7 -In certain embodiments, the phenotype is or comprises activated caspase 3, or a combination thereof. - / CAR + / CCR7 + , activated caspase 3 - / CAR + / CCR7 - , activated caspase 3 - / CAR + / CCR7 + , activated caspase 3 - / CAR + / CCR7 - In certain embodiments, the phenotype is or comprises annexin V, - / CAR + / CCR7 + , annexin V - / CAR + / CCR7 - , annexin V - / CAR + / CCR7 + , annexin V - / CAR + / CCR7 - or a combination thereof.

[0231] In some embodiments, the phenotype is or includes the positive or negative expression of a marker of depletion. In certain embodiments, the phenotype is or includes the positive or negative expression of a specific molecule associated with depletion. In certain embodiments, the specific molecule is any molecule associated with depletion or a quality associated with depletion, such as insufficient effector function or inhibitory receptor expression. In certain embodiments, the phenotype is the positive or negative expression of an immune checkpoint inhibitor. In certain embodiments, the marker of depletion is CTLA-4, FOXP3, PD-1, TIGIT, LAB-3, 2B4, BTLA, TIM3, VISTA, or CD96, or a combination thereof. In certain embodiments, the phenotype is the positive or negative expression of CTLA-4, FOXP3, PD-1, TIGIT, LAB-3, 2B4, BTLA, TIM3, VISTA, or CD96, or a combination thereof. In certain embodiments, the phenotype is the positive or negative expression of PD1 and / or FOXP3.

[0232] In some embodiments, the phenotype is a CD3 + In certain embodiments, the phenotype is a positive or negative expression of a depletion marker in the cells. + In some embodiments, the phenotype is or comprises positive or negative expression of depletion markers in the cells. In some embodiments, the phenotype is or comprises positive or negative expression of depletion markers and CD3 + and positive expression of a recombinant receptor or CAR. In certain embodiments, the phenotype is or comprises a CD4 + In some embodiments, the phenotype is a CD8+ / CD9+ marker expressing a recombinant receptor or CAR. + The depletion markers may be or include positive or negative expression of depletion markers in the cells. In certain embodiments, the depletion markers are one or more of CTLA-4, FOXP3, PD-1, TIGIT, LAB-3, 2B4, BTLA, TIM3, VISTA, or CD96. In certain embodiments, the depletion markers are PD1 and / or FOXP3.

[0233] In certain embodiments, the phenotype is PD1 - / CD3 + , PD1 - / CD4 + , PD1 - / CD8 + , PD1 - / CD3 + / CAR + , PD1 - / CD4 + / CAR + , PD1 - / CD8 + / CAR + , PD1 - / Annexin V - , PD1 - / Annexin V - / CD3 + , PD1 - / Annexin V - / CD4 + , PD1 - / Annexin V - / CD8 + , PD1 - / Annexin V / - CD3 + / CAR + , PD1 - / Annexin V - / CD4 + / CAR + , PD1 - / Annexin V - / CD8 + / CAR + , PD1 - / activated caspase 3 - , PD1 - / activated caspase 3 - / CD3 + , PD1 - / activated caspase 3 - / CD4 + , PD1 - / activated caspase 3 - / CD8 + , PD1 - / activated caspase 3 - / CD3 + / CAR +, PD1 - / activated caspase 3 - / CD4 + / CAR + , PD1 - / activated caspase 3 - / CD8 + / CAR + or a combination thereof.

[0234] In certain embodiments, the phenotype is FOXP3 - / CD3 + , FOXP3 - CD4 + , FOXP3 - / CD8 + , FOXP3 - / CD3 + / CAR + , FOXP3 - / CD4 + / CAR + , FOXP3 - / CD8 + / CAR + , FOXP3 - / Annexin V - , FOXP3 - / Annexin V - / CD3 + , FOXP3 - / Annexin V - / CD4 + , FOXP3 - / Annexin V - / CD8 + , FOXP3 - / Annexin V / - CD3 + / CAR + , FOXP3 - / Annexin V - / CD4 + / CAR + , FOXP3 - / Annexin V - / CD8 + / CAR + , FOXP3 - / activated caspase 3 - , FOXP3 - / activated caspase 3 - / CD3 + , FOXP3 - / activated caspase 3 - / CD4 + , FOXP3 - / activated caspase 3 - / CD8 + , FOXP3 - / activated caspase 3 - / CD3 + / CAR + , FOXP3 - / activated caspase 3 - / CD4 + / CAR + , FOXP3 - / activated caspase 3 - / CD8 + / CAR + or a combination thereof.

[0235] In certain embodiments, the phenotype is the negative expression of a specific molecule associated with T cell activation. In some embodiments, the phenotype is or includes the negative expression of one or more specific molecules that are activation markers. Generally, T cell activation requires two simultaneous signals. The first is the binding of the T cell receptor complex (TCR) to a major histocompatibility complex (MHC) molecule carrying a peptide antigen. The second is provided by the binding of the costimulatory receptor CD28 to a protein in the surface of an APC, such as B7-2 or B7-1. In certain embodiments, the specific molecule is associated with TCR activation, e.g., activated, altered, or expressed as a result of T cell activation. In some embodiments, the specific molecule is associated with activating the CD28 receptor, e.g., a molecule that is activated, altered, or expressed as a result of T cell activation.

[0236] In certain embodiments, the phenotype is or comprises negative expression of one or more specific molecules that are activation markers. In one or more certain embodiments, the activation marker is one or more of CD25, CD26, CD27, CD28, CD30, CD71, CD154, CD40L, CD127, LAG3, Ki67, or a combination thereof. In certain embodiments, the phenotype is negative or positive expression of one or more of CD25, CD26, CD27, CD28, CD30, CD71, CD154, CD40L, CD127, LAG3, or Ki67. In certain embodiments, the phenotype is or comprises expression of CD25, CD127, LAG3, Ki67, or a combination thereof.

[0237] In some embodiments, the phenotype is a CD3 + The phenotype is or includes positive or negative expression of activation markers in cells. In certain embodiments, the phenotype is a CD4 + In some embodiments, the phenotype is a CD8+ / CD9+ phenotype that expresses a recombinant receptor or CAR. + The positive or negative expression of an activation marker in the cells or the like is or comprises the positive or negative expression of an activation marker in the cells. In certain embodiments, the activation marker is one or more of CD25, CD26, CD27, CD28, CD30, CD71, CD154, CD40L, CD127, LAG3, or Ki67. In certain embodiments, the activation marker is CD25, CD127, LAG3, Ki67, or a combination thereof.

[0238] In certain embodiments, the phenotype is determined by positive or negative expression of activation markers, as well as CD3 + , CD4 + , CD8 + , CD3 + / CAR + , CD4 + / CAR + , CD8 + / CAR + , annexin V - , annexin V - / CD3+ , annexin V - / CD4 + , annexin V - / CD8 + , Annexin V / - CD3 + / CAR + , annexin V - / CD4 + / CAR + , annexin V - / CD8 + / CAR + , activated caspase 3 - , activated caspase 3 - / CD3 + , activated caspase 3 - / CD4 + , activated caspase 3 - / CD8 + , activated caspase 3 - / CD3 + / CAR + , activated caspase 3 - / CD4 + / CAR + , activated caspase 3 - / CD8 + / CAR + or a combination thereof.

[0239] In some embodiments, the phenotype is assessed by response to a stimulus, for example, a stimulus that induces, induces, stimulates, or prolongs immune cell function. In certain embodiments, cells are incubated under stimulatory conditions or in the presence of a stimulus, and the phenotype is or includes a response to the stimulus. In certain embodiments, the phenotype is or includes the production or secretion of a soluble factor in response to one or more stimuli. In some embodiments, the phenotype is or includes the absence, production, or secretion of a soluble factor in response to one or more stimuli. In certain embodiments, the soluble factor is a cytokine. In some embodiments, the cytokine is IL-2. In some embodiments, the cytokine is TNFα.

[0240] The conditions can include one or more of a specific medium, temperature, oxygen content, carbon dioxide content, time, agents such as nutrients, amino acids, antibiotics, ions, and / or stimulatory factors such as cytokines, chemokines, antigens, binding partners, fusion proteins, recombinant soluble receptors, and any other agents designed to activate cells. In some embodiments, the cells are stimulated, and the phenotype is determined by whether a soluble factor, such as a cytokine or chemokine, is produced or secreted. In some embodiments, the stimulus is non-specific, i.e., not an antigen-specific stimulus. In some embodiments, the stimulus includes PMA and ionomycin. In some embodiments, the cells are incubated in the presence of stimulatory conditions or stimulatory agents for a period of about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, about 11 hours, about 12 hours, about 18 hours, about 24 hours, about 48 hours, or between 1 hour and 4 hours, between 1 hour and 12 hours, between 12 hours and 24 hours, or for a period of more than 24 hours.

[0241] In some embodiments, the cells are stimulated with an agent that is an antigen or epitope specific for the recombinant receptor, or an antibody or fragment thereof that binds to and / or recognizes the recombinant receptor, or a combination thereof. In some embodiments, the recombinant receptor is a CAR, and the agent is an antigen or epitope specific for the CAR, or an antibody or fragment thereof that binds to and / or recognizes the CAR, or a combination thereof. In certain embodiments, the cells are stimulated by incubating the cells in the presence of target cells that have surface expression of the antigen recognized by the CAR. In certain embodiments, the recombinant receptor is a CAR, and the agent is an antibody, or an active fragment, variant, or portion thereof, that binds to the CAR. In certain embodiments, the antibody, or an active fragment, variant, or portion thereof, that binds to the CAR is an anti-idiotypic (anti-ID) antibody.

[0242] In some embodiments, the stimulatory conditions or agents include one or more agents, e.g., ligands, that can stimulate / activate the intracellular signaling domain of the TCR complex. In some aspects, the agents enable or initiate the TCR / CD3 intracellular signaling cascade in T cells. Such agents can include antibodies, e.g., those specific for TCR components and / or costimulatory receptors, e.g., anti-CD3, anti-CD28 (e.g., bound to a solid support such as beads), and / or one or more cytokines. In some embodiments, the one or more agents are PMA and ionomycin.

[0243] In certain embodiments, the phenotype is or includes the production or secretion of cytokines in response to one or more stimuli. Cytokine production and / or secretion contribute to immune responses and are involved in different processes, including the induction of antiviral proteins and the induction of T cell proliferation. Cytokines are not preformed factors, but are rapidly produced and secreted in response to cell activation. Cytokine production or secretion can be measured, detected, and / or quantified by any suitable technique known in the art.

[0244] In certain embodiments, the phenotype is the production of one or more cytokines. In some embodiments, the production of two or more cytokines from the same cell can be indicative of the multifunctional characteristics of such cells. In certain embodiments, the production of one or more cytokines is measured, detected, and / or quantified by intracellular cytokine staining. Intracellular cytokine staining (ICS) by flow cytometry is a well-adapted technique for studying cytokine production at the single-cell level. It detects the production and accumulation of cytokines in the endoplasmic reticulum after cell stimulation, thereby enabling the identification of cell populations that are positive or negative for the production of a particular cytokine or the separation of high- and low-producing cells based on a threshold. In some embodiments, as described above, stimulation can be performed using a non-specific stimulus (e.g., not an antigen-specific stimulus). For example, PMA / ionomycin can be used for non-specific cell stimulation. In some embodiments, stimulation can be performed with an agent that is an antigen or epitope specific to a recombinant receptor (e.g., a CAR), or an antibody or fragment thereof that binds to and / or recognizes the recombinant receptor, or a combination thereof. ICS can also be used in combination with other flow cytometry protocols for immunophenotyping using cell surface markers or MHC multimers to access cytokine production in specific cell subpopulations, making ICS a highly flexible and versatile method. Other single-cell techniques for measuring or detecting cytokine production include, but are not limited to, ELISPOT, limiting dilution, and T cell cloning.

[0245] In some embodiments, the phenotype is the production of a cytokine, such as after stimulation of the recombinant receptor with an antigen specific for and / or recognized by the recombinant receptor. In certain embodiments, the phenotype is the lack of cytokine production, such as after stimulation of the recombinant receptor with an antigen specific for and / or recognized by the recombinant receptor. In certain embodiments, the phenotype is positive for or high-level production of a cytokine. In certain embodiments, the phenotype is negative for or low-level production of a cytokine. Cytokines may include, but are not limited to, interleukin-1 (IL-1), IL-1β, IL-2, sIL-2Ra, IL-3, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-27, IL-33, IL-35, TNF, tumor necrosis factor alpha (TNF-α), CXCL2, CCL2, CCL3, CCL5, CCL17, CCL24, PGD2, LTB4, interferon gamma (IFN-γ), granulocyte-macrophage colony-stimulating factor (GM-CSF), macrophage inflammatory protein (MIP)-1α, MIP-1β, Flt-3L, fractalkine, and / or IL-5. In some embodiments, the phenotype includes production of cytokines associated with a particular cell type, such as cytokines associated with Th1, Th2, Th17, and / or Treg subtypes. In some embodiments, exemplary Th1-associated cytokines include IL-2, IFN-γ, and transforming growth factor beta (TGF-β), and in some cases are involved in cellular immune responses. In some embodiments, exemplary Th2-associated cytokines include IL-4, IL-5, IL-6, IL-10, and IL-13, and in some cases are associated with humoral immunity and anti-inflammatory properties. In some embodiments, exemplary Th17-associated cytokines include IL-17A and IL-17F, and in some cases are involved in the recruitment of neutrophils and macrophages, for example, during inflammatory responses.

[0246] In some embodiments, the phenotype is or comprises cytokine production. In certain embodiments, the phenotype is or comprises production of more than one cytokine (e.g., multifunctional). In certain embodiments, the phenotype is or comprises lack of production of one or more cytokines. In certain embodiments, the phenotype is or comprises production of or lack of one or more of IL-2, IL-5, IL-13, IFN-gamma, or TNF-alpha. In certain embodiments, the phenotype is or comprises production of or lack of one or more of IL-2, IL-13, IFN-gamma, or TNF-alpha. In some embodiments, the phenotype is the presence of cytokine production, and / or the presence of high levels of cytokine production. In some embodiments, the phenotype is low cytokine production, reduced cytokine production, or absence of cytokine production.

[0247] In some embodiments, the phenotype is or includes the endogenous (intracellular) production of a cytokine, e.g., as measured in the presence of a stimulator or under stimulating conditions when secretion is blocked or inhibited. In some embodiments, the stimulator is a non-specific stimulator, e.g., a stimulator that does not bind to an antigen-binding domain (e.g., on a recombinant receptor (e.g., a CAR)). In some embodiments, the stimulator is PMA / ionomycin, which can act as a non-specific stimulator. In some embodiments, the stimulator is a specific stimulator, e.g., an antigen or epitope thereof specific for a recombinant receptor (e.g., a CAR), or an antibody or fragment thereof that binds to and / or recognizes the recombinant receptor, or a combination thereof. In certain embodiments, the phenotype is or includes the lack or absence of endogenous production of a cytokine. In certain embodiments, the phenotype is or includes the endogenous amount of one or more cytokines upon production of more than one cytokine, as measured by an ICS assay. In certain embodiments, the phenotype is or includes an internal amount of one or more of IL-2, IL-5, IL-13, IFN-gamma, or TNF-alpha, as measured by an ICS assay. In some embodiments, the phenotype is or includes a low internal amount or lack of detectable amounts of one or more cytokines, as measured by an ICS assay. In certain embodiments, the phenotype is or includes a low internal amount or lack of detectable amounts of IL-2, IL-5, IL-13, IFN-gamma, or TNF-alpha, as measured by an ICS assay. In some embodiments, the phenotype includes assessment of multiple cytokines, e.g., by a multiplexed assay or an assay that assesses multifunctionality (see, e.g., Xue et al., (2017) Journal for ImmunoTherapy of Cancer 5:85). In some embodiments, the lack of cytokine expression inversely correlates or is associated with cellular activity and / or function and / or duration of response and progression-free survival.In some embodiments, cells that have reduced, minimal, or no cytokine production, as assessed according to any known method or the methods described herein, are reduced in a cell composition (e.g., an output composition, a therapeutic cell composition).

[0248] Certain embodiments contemplate that the phenotype may include cytokine production or the absence or low production of a cytokine. This may depend on several factors, including, but not limited to, the identity of the cytokine, the assay performed to detect the cytokine, and the stimuli or conditions used in conjunction with the assay. For example, in some embodiments, it is contemplated that the phenotype is or includes the absence or low levels of IL-13 production as indicated by ICS, while in some embodiments, the phenotype is or includes the production of IFN-gamma as indicated by ICS.

[0249] In some embodiments, the phenotype is determined by the production of one or more cytokines, as well as CD3 + , CD4 + , CD8 + , CD3 + / CAR + , CD4 + / CAR + , CD8 + / CAR + , annexin V - , annexin V - CD3 + , annexin V - CD4 + , annexin V - CD8 + , annexin V - CD3 + / CAR + , annexin V - CD4 + / CAR + , annexin V - CD8 + / CAR + , activated caspase 3 - , activated caspase 3 - / CD3+ , activated caspase 3 - / CD4 + , activated caspase 3 - / CD8 + , activated caspase 3 - / CD3 + / CAR + , activated caspase 3 - / CD4 + / CAR + , or activated caspase 3 - / CD8 + / CAR + In certain embodiments, the phenotype is or comprises any one of, or a combination thereof. + / CAR + and / or CD8 + / CAR + In some embodiments, the phenotype is or comprises the production of one or more cytokines in the CD4 + / CAR + In some embodiments, the phenotype is or comprises the production of IL-2 in CD4 cells. + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in CD4 cells. + / CAR + In some embodiments, the phenotype is or comprises the production of IL-2 and TNF-alpha in CD4 cells. + / CAR + In some embodiments, the phenotype is or comprises the production of IL-2 and IFN-gamma in CD8 cells. + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in CD8 cells. + / CAR + In some embodiments, the phenotype is or comprises the production of IFN-gamma and TNF-alpha in the cell. - / CD4 + / CAR+ In some embodiments, the phenotype is or comprises the production of IL-2 in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or includes the production of IL-2 and TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or includes the production of IL-2 and IFN-gamma in the cell. - / CD8 + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in the cell. - / CD8 + / CAR + In some embodiments, the phenotype is or comprises the production of IFN-gamma and TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises the production of IL-2 and TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises the production of IL-2 and IFN-gamma in the cell. - / CD8 + / CAR + In some embodiments, the phenotype is or comprises the production of TNF-alpha in the cell. - / CD8 + / CAR +The phenotypes described in this paragraph are or include the production of IFN-gamma and TNF-alpha in the cells. In some embodiments, the phenotypes described in this paragraph positively correlate with durable responses and progression-free survival. Thus, in some embodiments, cells comprising these phenotypes are maximized or expanded in a cell composition (e.g., output composition, therapeutic cell composition). In some embodiments, the cell composition comprises CD4+ T cells, e.g., engineered CD4+ T cells expressing a recombinant receptor (e.g., CAR), capable of producing cytokines selected from interleukin 2 (IL-2) and / or TNF-alpha. At least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least at least 94% or at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or 95% or about 95%, at least 96% or at least about 96%, or 96% or about 96%, at least 97% or at least about 97%, or 97% or about 97%, at least 98% or at least about 98%, or 98% or about 98%, at least 99% or at least about 99%, or 99% or about 99%, or at least 100% or at least about 100%, or 100% or about 100%.In some embodiments, the cell composition comprises at least or at least about 10%, or 10%, or at least 15%, or at least about 15%, or 15%, or at least 20%, or at least about 20%, or 20% or about 20%, or at least 25%, or 25%, or at least 30%, or at least about 30%, or 30% or about 30%, or at least 40%, or 40% or about 40%, or more, of the total number of CD4+ T cells, e.g., engineered CD4 T cells expressing a recombinant receptor (e.g., a CAR), in the composition that are multifunctional for the production of two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

[0250] In some embodiments, the phenotype is or includes a lack of production of one or more cytokines. In certain embodiments, the phenotype includes a lack of production of one or more cytokines, as well as a lack of CD3 + , CD4 + , CD8 + , CD3 + / CAR + , CD4 + / CAR + , CD8 + / CAR + , annexin V - , annexin V - CD3 + , annexin V - CD4 + , annexin V - CD8 + , annexin V - CD3 + / CAR + , annexin V - CD4 + / CAR + , annexin V - CD8 + / CAR + , activated caspase 3 - , activated caspase 3 - / CD3+ , activated caspase 3 - / CD4 + , activated caspase 3 - / CD8 + , activated caspase 3 - / CD3 + / CAR + , activated caspase 3 - / CD4 + / CAR + , or activated caspase 3 - / CD8 + / CAR + In some embodiments, the phenotype is or comprises one or more of the following: IL-2, IFN-gamma, and / or TNF-alpha; or a combination thereof. In some embodiments, the phenotype is or comprises activated caspase 3; - / CD4 + / CAR + In some embodiments, the phenotype is or comprises a lack of production of IL-2 in the cells. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises a lack of production of TNF-alpha in the cell. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises a lack of production of IL-2 and TNF-alpha in the cells. - / CD4 + / CAR + In some embodiments, the phenotype is or comprises a lack of production of IL-2 and IFN-gamma in the cells. - / CD8 + / CAR + In some embodiments, the phenotype is or comprises a lack of production of TNF-alpha in the cell. - / CD8 + / CAR +The phenotypes described in this paragraph are or include the lack of production of INF-gamma and TNF-alpha in cells. In some embodiments, the phenotypes described in this paragraph are negatively correlated with sustainable response and progression-free survival. Thus, in some embodiments, cells comprising these phenotypes are minimized or reduced in the cell composition (e.g., output composition, therapeutic cell composition). For example, in some embodiments, cells comprising the phenotypes described in this paragraph comprise less than 2%, 5%, 10%, 15%, 20%, or 25% of the total cells in the cell composition (e.g., output composition, therapeutic cell composition).

[0251] In certain embodiments, the phenotype is or includes the internal amount of one or more of IL-2, IL-13, IFN-gamma, or TNF-alpha as measured by an ICS assay, and the presence or absence of one or more specific markers for cell subsets or cells of a particular cell type. In some embodiments, the phenotype is or includes the production or lack thereof of one or more of IL-2, IL-13, IFN-gamma, or TNF-alpha, as well as CD4 + / CAR + and / or CD8 + / CAR + In certain embodiments, the phenotype is or comprises IL-2 production and CD4 + / CAR + and / or CD8 + / CAR + In some embodiments, the phenotype is or comprises a lack of or low production of IL-2 and CD4 + / CAR + and / or CD8 + / CAR + In some embodiments, the phenotype is or comprises IL-13 production as well as CD4 + / CAR + and / or CD8 + / CAR + In some embodiments, the phenotype is or comprises IL-13 production as well as CD4 + / CAR + and / or CD8 + / CAR + In certain embodiments, the phenotype is or comprises a lack of or low production of IL-13 and CD4 + / CAR + and / or CD8 + / CAR + In some embodiments, the phenotype is or comprises IFN-gamma production and CD4 + / CAR + and / or CD8 + / CAR + In certain embodiments, the phenotype is or comprises TNF-alpha production and CD4 + / CAR + and / or CD8 + / CAR + In certain embodiments, the phenotype is or comprises a lack of or low production of TNF-alpha and CD4 + / CAR + and / or CD8 + / CAR + is or contains

[0252] Any one or more phenotypes, alone or in combination, can be assessed or determined according to the methods provided. In some embodiments, the phenotype is CD3 + , CD3 + / CAR + , CD4 + / CAR + , CD8 + / CAR + , or a combination thereof.

[0253] In certain embodiments, the phenotype is CD3 + In certain embodiments, the phenotype is or comprises CD3 + / CAR + In some embodiments, the phenotype is or comprises CD8 + / CAR + In certain embodiments, the phenotype is or comprises CD4+ / CAR+.

[0254] In certain aspects, the phenotype is annexin - / CD3 + / CAR + In some embodiments, the phenotype is or comprises annexin - / CD4 + / CAR + In certain aspects, the phenotype is or comprises annexin - / CD8 + / CAR.

[0255] In certain embodiments, the phenotype is characterized by the absence or low amount of intracellular IL-2 and CD4 + / CAR + In certain embodiments, the phenotype is characterized by the absence or low amount of intracellular IL-13 and CD4 + / CAR + In some embodiments, the phenotype is characterized by the absence or low amount of intracellular expression of IL-13 and CD8 + / CAR + In certain embodiments, the phenotype is characterized by the absence or low amount of intracellular TNF-alpha, CD4 + / CAR + is.

[0256] In certain embodiments, the phenotype is CD8 + / CAR + In certain aspects, the phenotype is or comprises annexin - / CD8 + / CAR + is or contains

[0257] In some embodiments, the phenotype includes indicators of production of one or a combination of cytokines (wherein the one or more cytokines are IL-2, IL-13, IL-17, IFN-gamma, or TNF-alpha), optionally non-specifically and / or polyclonally produced to an antigen or recombinant receptor. In some embodiments, the indicators of production are measured in an assay comprising incubating a sample of the T cell composition with a polyclonal agent, an antigen-specific agent, or an agent that binds to the recombinant receptor (optionally a CAR), optionally in an intracellular cytokine staining assay. In some embodiments, the agent is or comprises PMA and ionomycin, or is or comprises a T cell receptor or T cell receptor complex agonist. In some embodiments, the phenotype includes negative expression of an activation marker selected from among CD25, CD127, LAG3, Ki67, and combinations thereof. In some embodiments, the phenotype includes negative expression of an exhaustion marker, where the exhaustion marker is a PD1 or FOXP3 gene product, or a combination thereof. In some embodiments, the phenotype comprises a naive phenotype or a memory phenotype, and optionally, the memory phenotype comprises a T effector memory phenotype, a T central memory phenotype, or a T effector memory phenotype expressing CD45RA (Temra).

[0258] In some embodiments, the recombinant receptor (e.g., CAR)-dependent activity is a measure of the production or accumulation of a pro-inflammatory cytokine, optionally one or a combination of TNF-alpha, IFN-gamma, and IL-2. In some embodiments, the reference measure is an average of the measures among a plurality, optionally at least 10, at least 15, or at least 20, reference therapeutic T cell compositions comprising the recombinant receptor (e.g., CAR), where (i) each of the reference therapeutic T cell compositions has been observed or determined to result in an acceptable safety profile following administration to a subject (optionally, the subject expresses the antigen or has a disease or condition associated therewith); and / or (ii) each of the reference therapeutic T cell compositions has been observed or determined to result in a desired efficacy following administration to a subject (optionally, the subject expresses the antigen or has a disease or condition associated therewith).

[0259] In certain embodiments, the number, multiple, or fraction of cells of a particular phenotype in a cell composition is determined, measured, obtained, detected, observed, and / or identified. In some embodiments, the cell composition is a T cell composition. In certain embodiments, the cell composition contains cells expressing a recombinant receptor, e.g., a CAR. In certain embodiments, the cell composition is a therapeutic T cell composition containing cells expressing a recombinant receptor that can be administered to a subject to treat a disease or condition. In certain embodiments, the number of cells of the phenotype is the total number of cells of the phenotype in the cell composition. In certain embodiments, the number of cells of the phenotype is the total number of cells of the phenotype present in a dose of the cell composition. In certain embodiments, the number of cells of the phenotype is the number of cells of the phenotype present in a sample of the cell composition. In some embodiments, the number of cells of the phenotype can be expressed as the frequency, ratio, and / or percentage of cells of the phenotype present in the cell composition, or the dose or sample thereof.

[0260] In certain embodiments, the number, multiple, or fraction of cells of a phenotype is transformed, e.g., to compress the range of relevant values ​​of the number, multiple, or fraction. In some embodiments, a transformation is any application of a deterministic mathematical function to each point in a data set, e.g., each data point x is replaced by its transformed value y=f(x), where f is a function. Generally, a transformation can be applied so that the data more closely meets the assumptions of the statistical inference procedure to be applied or to improve the interpretation or appearance of a graph. In most cases, the functions used to transform data are invertible and generally continuous. Transformations are usually applied to sets of comparable measurements. Examples of suitable transformations include, but are not limited to, log and square root transformations, reciprocal transformations, and power transformations. In certain embodiments, the number, multiple, or fraction of cells of a phenotype is transformed by a logarithmic transformation. In certain embodiments, a logarithmic transformation is performed using a common logarithm (log 10 (x)), natural logarithm (ln(x)), or binary logarithm (log2(x)).

[0261] B. Medication and Administration Decisions A therapeutic composition or dose thereof, in some embodiments, contains an amount of cells effective to treat or prevent a disease or condition, such as a therapeutically effective amount or a prophylactically effective amount. In some embodiments, the composition includes an amount of cells effective to reduce the burden of a disease or condition. In some embodiments, the composition includes an amount of cells that provides a more consistent outcome, e.g., response and / or safety outcome, and / or more consistent pharmacokinetic parameters, among a group of subjects to whom the composition is administered. In some embodiments, the composition includes an amount of cells effective to promote a durable response and / or progression-free survival. In some aspects, provided methods involve evaluating a therapeutic composition containing T cells for cell phenotype and determining a dose based on such outcome.

[0262] In some embodiments, doses are determined to include a relatively consistent number, ratio, ratio, and / or percentage of engineered cells having a particular phenotype in one or more particular compositions. In some aspects, consistency is associated with or relates to a relatively consistent activity, function, pharmacokinetic parameters, toxicity outcome, and / or response outcome. In some aspects, across multiple subjects, compositions, and / or doses, the number, ratio, ratio, and / or percentage is relatively consistent, e.g., has a particular phenotype, e.g., expresses CCR7 (CCR7), in a composition or unit dose. + The number or ratio of cells that produce cytokines, such as IL-2, TNF-alpha, or IFN-gamma, varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%. In some aspects, the number or ratio of cells in a composition or unit dose that have a particular phenotype, such as expressing CCR7 (CCR7 + ) cells vary by no more than 20%, or no more than 10%, or no more than 5% from the mean of said numbers or ratios among a plurality of T cell compositions produced by the process, and / or vary by no more than one standard deviation from such mean, or vary by no more than 20%, or no more than 10%, or no more than 5% among a plurality of T cell compositions or doses determined. In some embodiments, the plurality of subjects comprises at least 10 subjects, e.g., at least 15, at least 20, at least 25, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100 or more subjects.

[0263] In some aspects, the dose, e.g., one or more unit doses, is determined based on the number, percentage, ratio, frequency, and / or proportion of engineered T cells, e.g., a particular cell subset having a particular phenotype, such as a particular surface marker phenotype. In some aspects, the cell phenotype is determined based on the expression and / or absence of a particular cell marker, e.g., a surface marker. In some aspects, the cell marker includes a marker indicative of cell viability and / or apoptotic state. In some aspects, exemplary markers include CD3, CD4, CD8, CCR7, CD27, CD45RA, Annexin V, or activated caspase 3. In some aspects, an exemplary marker is CCR7. In some aspects, an exemplary marker is CD27. In some aspects, exemplary markers include CCR7 and / or CD27. In some aspects, exemplary markers include CCR7, CD27, and / or CD45RA.

[0264] In some embodiments, methods are provided that involve administering to a subject one or more unit doses of a therapeutic T cell composition, e.g., any unit dose described herein and / or determined by the methods provided herein.

[0265] In some embodiments, methods involve administering to a subject having a disease or condition a unit dose of a T cell composition comprising cells comprising a recombinant receptor, such as a chimeric antigen receptor (CAR), that specifically binds to an antigen associated with the disease or condition, wherein the therapeutic composition is administered at a dose that is in a range of 100-1500 mg / mL, and / or 100-2000 mg / mL. + ) cells, total CD8 + Recombinant receptor expression (receptor + / CD8 + ) cells are administered, and / or certain phenotypes of such cells, e.g., CCR7 + / CD4 + , CCR7 + / CD8 + , CD27 + / CD4 + , CD27 + / CD8+ , CD45RA + / CD4 + , CD45RA + / CD8 + , CCR7 - / CD4 + , CCR7 - / CD8 + , CD27 - / CD4 + , CD27 - / CD8 + , CD45RA - / CD4 + , CD45RA - / CD8 + , CCR7 + / CD27 + / CD4 + , CCR7 + / CD27 + / CD8 + , CCR7 + / CD45RA - / CD4 + , CCR7 + / CD45RA - / CD8 + , CCR7 - / CD45RA - / CD4 + , CCR7 - / CD45RA - / CD8 + , CCR7 - / CD27 - / CD4 + , CCR7 - / CD27 - / CD8 + In some embodiments, a unit dose is administered containing a defined number, percentage, ratio, frequency and / or proportion of cells having:

[0266] In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 C chemokine receptor type 7 (CCR7) ... + T cells (receptors + / CD8 + / CCR7 + cells) and / or a defined number of recombinant receptor-expressing CD4+ T cells (receptors + / CD4 + / CCR7 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + Cells and / or receptors + / CD4 + / CCR7 + In some embodiments, a unit dose of cells comprises a defined number of CD8 + / CCR7 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + In some embodiments, the defined number or ratio is further based on the expression or absence of expression on the cells of CD27 and / or CD45RA.

[0267] In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CD27 +Cells and / or receptors + / CD4 + / CD27 + In some embodiments, a unit dose of cells comprises a defined number of CD8 + / CD27 + In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CD27 + In some embodiments, the defined number or ratio is further based on the expression or absence of expression on the cells of CCR7 and / or CD45RA.

[0268] In some embodiments, the unit dose of cells comprises a defined number of recombinant receptor-expressing CD8 + T cells (receptors + / CD8 + / CCR7 + / CD27 + cells) and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptors + / CD4 + / CCR7 + / CD27 + cells) and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + Cells and / or receptors in defined ratios + / CD8 + / CCR7 + / CD27 + Cells and / or receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, a unit dose of cells comprises a defined number of CD8 + / CCR7 + / CD27+ In some embodiments, the unit dose of cells comprises a defined number of CD4 + / CCR7 + / CD27 + In some embodiments, the defined number or ratio is further based on the expression or absence of expression on the cells of CD45RA.

[0269] In some embodiments, the number of cells in a unit dose is the number of cells or the number of recombinant receptor-expressing or CAR-expressing cells, or the number, percentage, ratio, frequency and / or proportion of such cells of a certain phenotype, e.g., cells that express or do not express one or more markers selected from CD3, CD4, CD8, CCR7, CD27, CD45RA, Annexin V or activated caspase 3, that it is desired to administer in a dose to a particular subject, such as the subject from whom the cells were derived. In some embodiments, the number of cells in a unit dose is the number of cells or the number of recombinant receptor-expressing or CAR-expressing cells, or the number, percentage, ratio, frequency and / or proportion of such cells of a certain phenotype, e.g., cells that express or do not express one or more markers selected from CD3, CD4, CD8, CCR7, CD27, CD45RA, Annexin V or activated caspase 3. + , CD27 + , CD45RA + , CD45RA - , CD4 + , CD8 + , CD3 + , apoptosis marker negative (e.g., Annexin V - or caspase 3 - ) cells, or the number, percentage, ratio, frequency and / or proportion of cells that are positive or negative for any one or more of the foregoing.

[0270] In some embodiments, the number of cells in a unit dose is determined based on the number of cells or recombinant receptor- or CAR-expressing cells, or the number of such cells of a particular phenotype, e.g., CCR7, desired to be administered in a dose to a particular subject, such as the subject from whom the cells were derived. + / CD4 + , CCR7 + / CD8 + , CD27 + / CD4 + , CD27+ / CD8 + , CD45RA + / CD4 + , CD45RA + / CD8 + , CCR7 - / CD4 + , CCR7 - / CD8 + , CD27 - / CD4 + , CD27 - / CD8 + , CD45RA - / CD4 + , CD45RA - / CD8 + , CCR7 + / CD27 + / CD4 + , CCR7 + / CD27 + / CD8 + , CCR7 + / CD45RA - / CD4 + , CCR7 + / CD45RA - / CD8 + , CCR7 - / CD45RA - / CD4 + , CCR7 - / CD45RA - / CD8 + , CCR7 - / CD27 - / CD4 + , CCR7 - / CD27 - / CD8 + and apoptosis marker negative (e.g., Annexin V - or caspase 3 - ) cell number, percentage, ratio and / or proportion. In some embodiments, the unit dose is a defined number of cells, or a number of recombinant receptor- or CAR-expressing cells, or such cells of a certain phenotype, e.g., CCR7 + / CD4 + , CCR7 + / CD8 + , CD27 + / CD4 + , CD27 + / CD8 + , CD45RA + / CD4 + , CD45RA + / CD8 + , CCR7 - / CD4 + , CCR7 - / CD8 + , CD27 - / CD4 + , CD27 - / CD8 + , CD45RA - / CD4 + , CD45RA - / CD8 + , CCR7 + / CD27 + / CD4 + , CCR7 + / CD27 + / CD8 + , CCR7 + / CD45RA - / CD4 + , CCR7 + / CD45RA - / CD8 + , CCR7 - / CD45RA - / CD4 + , CCR7 - / CD45RA - / CD8 + , CCR7 - / CD27 - / CD4 + , CCR7 - / CD27 - / CD8 + and apoptosis marker negative (e.g., Annexin V - or caspase 3 - ) cells, and / or any subset thereof.

[0271] In some embodiments, the unit dose comprises annexin V - / CCR7 + / CAR + ;Annexin V- / CCR7 + / CAR + / CD4 + ;Annexin V - / CCR7 + / CAR + / CD8 + ;Annexin V - / CD27 + / CAR + ;Annexin V - / CD27 + / CAR + / CD4 + ;Annexin V - / CD27 + / CAR + / CD8 + ;Annexin V - / CCR7 + / CD27 + / CAR + ;Annexin V - / CCR7 + / CD27 + / CAR + / CD4 + ;Annexin V - / CCR7 + / CD27 + / CAR + / CD8 + ;Annexin V - / CCR7 + / CD45RA - / CAR + ;Annexin V - / CCR7 + / CD45RA - / CAR + / CD4 + ;Annexin V - / CCR7 + / CD45RA - / CAR + / CD8 + ;Annexin V - / CCR7 - / CD45RA - / CAR + ;Annexin V - / CCR7 - / CD45RA - / CAR + / CD4+ ;Annexin V - / CCR7 - / CD45RA - / CAR + / CD8 + ;Annexin V - / CCR7 - / CD27 - / CAR + , annexin V - / CCR7 - / CD27 - / CAR + / CD4 + ;Annexin V - / CCR7 - / CD27 - / CAR + / CD8 + ; Activated caspase 3 - / CCR7 + / CAR + ; Activated caspase 3 - / CCR7 + / CAR + / CD4 + ; Activated caspase 3 - / CCR7 + / CAR + / CD8 + ; Activated caspase 3 - / CD27 + / CAR + ; Activated caspase 3 - / CD27 + / CAR + / CD4 + ; Activated caspase 3 - / CD27 + / CAR + / CD8 + ; Activated caspase 3 - / CCR7 + / CD27 + / CAR + ; Activated caspase 3 - / CCR7 + / CD27 + / CAR + / CD4 + ; Activated caspase 3 - / CCR7 + / CD27 + / CAR + / CD8 + ; Activated caspase 3 - / CCR7 + / CD45RA - / CAR + ; Activated caspase 3 - / CCR7 + / CD45RA - / CAR + / CD4 + ; Activated caspase 3 - / CCR7 + / CD45RA - / CAR + / CD8 + ; Activated caspase 3 - / CCR7 - / CD45RA - / CAR + ; Activated caspase 3 - / CCR7 - / CD45RA - / CAR + / CD4 + ; Activated caspase 3 - / CCR7 - / CD45RA - / CAR + / CD8 + ; Activated caspase 3 - / CCR7 - / CD27 - / CAR + ; Activated caspase 3 - / CCR7 - / CD27 - / CAR + / CD4 + and / or activated caspase 3 - / CCR7 - / CD27 - / CAR + / CD8 + or a combination thereof, such as the number of cells or cell types and / or the frequency, ratio and / or percentage of cells or cell types, e.g., individual populations, phenotypes, or subtypes, in the cell composition.

[0272] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises about 1 x 10 cells. 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8+ / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells.

[0273] In some embodiments, the unit dose is 5×10 inclusive. 5 Or about 5 x 10 5 and 5×10 7 Or about 5 x 10 7 Between 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cells or Receptors + / CD4 + / CCR7 + In some embodiments, the unit dose comprises at least 5 x 10 cells. 7 Or at least about 5 x 10 7 , at least 1 x 10 7 Or at least about 1 × 10 7 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 1 x 10 6 Or at least about 1 × 10 6 , or at least about 5 × 10 5 Total receptors + / CD8 + / CCR7 + Cells or Receptors + / CD4 + / CCR7 + Contains cells.

[0274] In some embodiments, the unit dose is 5×10 inclusive. 5 Or about 5 x 10 5 and 5×107 Or about 5 x 10 7 Between 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CD27 + Cells or Receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises at least 5 x 10 cells. 7 Or at least about 5 x 10 7 , at least 1 x 10 7 Or at least about 1 × 10 7 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 1 x 10 6 Or at least about 1 × 10 6 , or at least about 5 × 10 5 Total receptors + / CD8 + / CD27 + Cells or Receptors + / CD4 + / CD27 + Contains cells.

[0275] In some embodiments, the unit dose is at least 1 x 10, inclusive. 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 x 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 106 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + cells, and / or at least 1 x 10 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 x 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + cells, and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Contains cells.

[0276] In some embodiments, the unit dose is at least 1 x 10, inclusive. 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 x 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 106 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD8 + / CD27 + cells, and / or at least 1 x 10 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 × 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7Total receptors between + / CD8 + / CD27 + cells, and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CD27 + The unit dose includes cells.

[0277] In some embodiments, the unit dose is 5×10 inclusive. 5 Or about 5 x 10 5 and 5×10 7 Or about 5 x 10 7 Between 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + Cells or Receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises at least 5 x 10 cells. 7 Or at least 5x10 7 Or at least about 5 x 10 7 , at least 1 x 10 7 Or at least 1×10 7Or at least about 1 × 10 7 , at least 5 × 10 6 Or at least 5x10 6 Or at least about 5 x 10 6 , at least 1 x 10 6 Or at least 1×10 6 Or at least about 1 × 10 6 , or at least 5 × 10 5 Or at least about 5 x 10 5 Total receptors + / CD8 + / CCR7 + / CD27 + Cells or Receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0278] In some embodiments, the unit dose is at least 1 x 10, inclusive. 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 x 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells, and / or at least 1 x 10 6 Or at least about 1 × 10 6 , at least 2 × 10 6 Or at least about 2 × 10 6 , at least 3 × 10 6 Or at least about 3 x 10 6 , at least 4 × 10 6 Or at least about 4×10 6 , at least 5 × 10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , or at least 1 × 10 7 Or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 3 x 10 cells, inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells, and / or 3 x 10 6 Or about 3 x 106 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + Contains cells.

[0279] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0280] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CD27 + Cell vs. Receptor + / CD4 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0281] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + In some embodiments, the unit dose comprises 1 x 10 cells. 8 Or about 1 x 10 8 Below, 5 x 10 7 Or about 5 x 10 7 Below, 1×10 7 Or about 1 x 10 7 Below, 5 x 10 6 Or about 5 x 10 6 Below, 1×10 6 Or about 1 x 10 6 or less, or 5 x 10 5 Or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0282] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor+ / CD4 + / CCR7 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0283] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD3 expressing the recombinant receptor between + Cells (receptors + / CD3 + cells) or total CD3 + In some embodiments, the unit dose comprises 5 x 10 cells. 8 Or about 5 x 10 8 Below, 1×10 8 Or about 1 x 10 8 Below, 5 x 10 7 Or about 5 x 10 7 Below, 1×10 7 Or about 1 x 10 7 Below, 5 x 10 6 Or about 5 x 10 6 Below, 1×10 6 Or about 1 x 10 6 or less, or 5 x 10 5 Or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + Contains cells.

[0284] In some embodiments, CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that are alive or viable. + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 +Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The total number of cells is the total number of such cells that do not express the apoptotic marker and / or are apoptotic marker negative ( - ) is the total number of such cells, where the apoptotic marker is annexin V or activated caspase 3.

[0285] In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, at least 15% or at least about 15%, or 15% or about 15%, at least 20% or at least about 20%, or 20% or about 20%, at least 30% or at least about 30%, or 30% or about 30%, at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60%, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% of the total number of T cells in the composition (or of the total number of T cells in the composition expressing a recombinant receptor). or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or 93% or about 93%, at least 94% or at least about 94%, or 94% or about 94%, at least 95% or at least about 95%, or 95% or about 95%, or at least At least 96%, or at least about 96%, or 96% or about 96%, at least 97%, or at least about 97%, or 97% or about 97%, at least 98%, or at least about 98%, or 98% or about 98%, at least 99%, or at least about 99%, or 99% or about 99%, or at least 100%, or at least about 100%, or 100% or about 100% are surface positive for CCR7 and / or CD27.

[0286] In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, at least 85% or at least about 85%, or 85% or about 85%, at least 90% or at least about 90%, or 90% or about 90%, at least 91% or at least about 91%, or 91% or about 91%, at least 92% or at least about 92%, or 92% or about 92%, at least 93% or at least about 93%, or at least about 93%, at least 94%, or at least about 94%, or 94% or about 94%, at least 95%, or at least about 95%, or 95% or about 95%, at least 96%, or at least about 96%, or 96% or about 96%, at least 97%, or at least about 97%, or 97% or about 97%, at least 98%, or at least about 98%, or 98% or about 98%, at least 99%, or at least about 99%, or 99% or about 99%, or at least 100%, or at least about 100%, or 100% or about 100% are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha. In some embodiments, the T cells are capable of producing IL-2 and / or TNF-alpha and are CD4+ T cells.

[0287] In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, the total receptor in a unit dose +at least 15% or at least about 15%, or 15% or about 15%, at least 20% or at least about 20%, or 20% or about 20%, at least 30% or at least about 30%, or 30% or about 30%, at least 40% or at least about 40%, or 40% or about 40%, at least 50% or at least about 50%, or 50% or about 50%, at least 60% or at least about 60%, or 60% or about 60%, at least 70% or at least about 70%, or 70% or about 70%, at least 80% or at least about 80%, or 80% or about 80%, or at least 90% or at least about 90%, or 90% or about 90% of the cells, or the total receptors in the unit dose + Between 15% or about 15% and 90% or about 90%, between 20% or about 20% and 80% or about 80%, between 30% or about 30% and 70% or about 70%, or between 40% or about 40% and 60% or about 60%, inclusive, of the cells are receptor + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, the total receptor in a unit dose is + at least 15% or at least about 15%, at least 20% or at least about 20%, at least 30% or at least about 30%, at least 40% or at least about 40%, at least 50% or at least about 50%, at least 60% or at least about 60%, at least 70% or at least about 70%, at least 80% or at least about 80%, or at least 90% or at least about 90% of the cells or total receptors in the unit dose +Between 15% or about 15% and 90% or about 90%, between 20% or about 20% and 80% or about 80%, between 30% or about 30% and 70% or about 70%, or between 40% or about 40% and 60% or about 60%, inclusive, of the cells are receptor + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + In some embodiments, the total receptor in the unit dose is + at least 15% or at least about 15%, at least 20% or at least about 20%, at least 30% or at least about 30%, at least 40% or at least about 40%, at least 50% or at least about 50%, at least 60% or at least about 60%, at least 70% or at least about 70%, at least 80% or at least about 80%, or at least 90% or at least about 90% of the cells or total receptors in the unit dose + Between 15% or about 15% and 90% or about 90%, between 20% or about 20% and 80% or about 80%, between 30% or about 30% and 70% or about 70%, or between 40% or about 40% and 60% or about 60%, inclusive, of the cells are receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - , receptor + / CD4 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD45RA - is.

[0288] In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, the total receptor in the composition or unit dose + / CD8 + At least 50% or at least about 50%, at least 60% or at least about 60%, at least 70% or at least about 70%, at least 80% or at least about 80%, or at least 90% or at least about 90% of the cells or total receptors in the composition or unit dose + / CD8 + Between 50% or about 50% and 90% or about 90%, between 60% or about 60% and 90% or about 90%, and between 70% or about 70% and 80% or about 80% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or receptor + / CD8 + / CCR7 + / CD27 + In some embodiments, in any of the compositions comprising T cells expressing a recombinant receptor provided herein, the total receptor in the composition or unit dose is + / CD4 + At least 50% or at least about 50%, at least 60% or at least about 60%, at least 70% or at least about 70%, at least 80% or at least about 80%, or at least 90% or at least about 90% of the cells or total receptors in the composition or unit dose + / CD4 + Between 50% or about 50% and 90% or about 90%, between 60% or about 60% and 90% or about 90%, and between 70% or about 70% and 80% or about 80% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27+ or receptor + / CD4 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - , receptor + / CD4 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD45RA - In some embodiments, the total receptors in the composition + / CD8 + At least 50% or at least about 50%, at least 60% or at least about 60%, at least 70% or at least about 70%, at least 80% or at least about 80%, or at least 90% or at least about 90% of the cells + / CD8 + / CCR7 + / CD27 + or the total receptors in the composition + / CD4 + At least 15% or at least about 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least about 80%, or at least 90% of the cells are receptor + / CD4 + / CCR7 + / CD27 + is.

[0289] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + In some embodiments, the unit dose comprises 1 x 10 cells. 8 Or about 1 x 10 8 Below, 5 x 10 7 Or about 5 x 10 7 Below, 1×10 7 Or about 1 x 10 7 Below, 5 x 10 6 Or about 5 x 10 6 Below, 1×10 6 Or about 1 x 10 6 or less, or 5 x 10 5 Or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors+ / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells.

[0290] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + Cell vs. Receptor + / CD4 + / CCR7 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0291] In some embodiments, the unit dose is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total CD8 expressing the recombinant receptor between + Cells (receptors + / CD8 + cells) or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4+ / CCR7 + / CD27 + In some embodiments, the unit dose comprises 1 x 10 cells. 8 Or about 1 x 10 8 Below, 5 x 10 7 Or about 5 x 10 7 Below, 1×10 7 Or about 1 x 10 7 Below, 5 x 10 6 Or about 5 x 10 6 Below, 1×10 6 Or about 1 x 10 6 or less, or 5 x 10 5 Or about 5 x 10 5 Total receptors below + / CD8 + Cell or total receptor + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells.

[0292] In some embodiments, the unit dose of cells comprises a defined ratio of receptors. + / CD8 + / CCR7 + / CD27 + Cell vs. Receptor + / CD4 + / CCR7 + / CD27 + cells, optionally at a ratio of 1:1 or about 1:1, or from about 1:3 to about 3:1.

[0293] In some embodiments, the methods provided involve administering a dose containing a defined number of cells. + / CD4 + , CCR7 + / CD8 + , CD27 + / CD4+ , CD27 + / CD8 + , CD45RA + / CD4 + , CD45RA + / CD8 + , CCR7 - / CD4 + , CCR7 - / CD8 + , CD27 - / CD4 + , CD27 - / CD8 + , CD45RA - / CD4 + , CD45RA - / CD8 + , CCR7 + / CD27 + / CD4 + , CCR7 + / CD27 + / CD8 + , CCR7 + / CD45RA - / CD4 + , CCR7 + / CD45RA - / CD8 + , CCR7 - / CD45RA - / CD4 + , CCR7 - / CD45RA - / CD8 + , CCR7 - / CD27 - / CD4 + , or CCR7 - / CD27 - / CD8 + A defined number of cells, e.g., a defined number of CARs, + The cells were 5.0 × 10 6 or approximately 5.0 x 10 6 and 2.25 x 10 7 Or about 2.25 x 10 7 Between 5.0 x 10 6 or approximately 5.0 x 10 6 and 2.0 x 10 7 Or about 2.0 x 10 7Between 5.0 x 10 6 or approximately 5.0 x 10 6 and 1.5 x 10 7 Or about 1.5 x 10 7 Between 5.0 x 10 6 or approximately 5.0 x 10 6 and 1.0×10 7 or approximately 1.0 x 10 7 Between 5.0 x 10 6 or approximately 5.0 x 10 6 and 7.5 x 10 6 Or about 7.5 x 10 6 Between 7.5 x 10 6 Or about 7.5 x 10 6 and 2.25 x 10 7 Or about 2.25 x 10 7 Between 7.5 x 10 6 Or about 7.5 x 10 6 and 2.0 x 10 7 Or about 2.0 x 10 7 Between 7.5 x 10 6 Or about 7.5 x 10 6 and 1.5 x 10 7 Or about 1.5 x 10 7 Between 7.5 x 10 6 Or about 7.5 x 10 6 and 1.0×10 7 or approximately 1.0 x 10 7 Between 1.0 x 10 7 or approximately 1.0 x 10 7 and 2.25 x 10 7 Or about 2.25 x 10 7 Between 1.0 x 10 7 or approximately 1.0 x 10 7 and 2.0 x 10 7 Or about 2.0 x 10 7 Between 1.0 x 10 7 or approximately 1.0 x 10 7 and 1.5 x 10 7 Or about 1.5 x 10 7 Between 1.5 x 10 7 Or about 1.5 x 10 7 and 2.25 x 10 7 Or about 2.25 x 107 Between 1.5 x 10 7 Or about 1.5 x 10 7 and 2.0 x 10 7 Or about 2.0 x 10 7 Between 2.0 x 10 7 Or about 2.0 x 10 7 and 2.25 x 10 7 Or about 2.25 x 10 7 In some embodiments, such a dose, e.g., such a specified number of cells, refers to the total recombinant receptor-expressing cells in the administered composition. In some aspects, the specified number of recombinant receptor-expressing cells administered are cells that are apoptosis marker-negative (-), and optionally the apoptosis marker is annexin V or activated caspase 3.

[0294] In some embodiments, the unit dose of cells comprises a number of cells, e.g., a defined number of cells, at least 5×10 6 Or at least about 5 x 10 6 , at least 6 × 10 6 Or at least about 6 x 10 6 , at least 7 × 10 6 Or at least about 7 x 10 6 , at least 8 x 10 6 Or at least about 8 x 10 6 , at least 9 x 10 6 Or at least about 9 x 10 6 , at least ...

Claims

1. A therapeutic composition comprising T cells expressing a recombinant receptor, wherein the T cells expressing the recombinant receptor are at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%, or at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 85%, at least about 90%, at least or at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% are surface positive for CCR7 and / or CD27.

2. 2. The therapeutic composition of claim 1, wherein at least 50%, or at least about 50%, or 50%, or about 50% of the total number of T cells in the composition or the total number of T cells expressing the recombinant receptor in the composition are surface positive for CCR7 and / or CD27.

3. 2. The therapeutic composition of claim 1, wherein at least 60%, or at least about 60%, or 60%, or about 60% of the total number of T cells in the composition or the total number of T cells expressing the recombinant receptor in the composition are surface positive for CCR7 and / or CD27.

4. 2. The therapeutic composition of claim 1, wherein at least 70%, or at least about 70%, or 70%, or about 70% of the total number of T cells in the composition or the total number of T cells expressing the recombinant receptor in the composition are surface positive for CCR7 and / or CD27.

5. 2. The therapeutic composition of claim 1, wherein at least 80%, or at least about 80%, or 80%, or about 80% of the total number of T cells in the composition or the total number of T cells expressing the recombinant receptor in the composition are surface positive for CCR7 and / or CD27.

6. T cells in the composition and / or expressing the recombinant receptor are CD8 + comprising or consisting of T cells; T cells in the composition and / or expressing the recombinant receptor are CD4 + comprising or consisting of T cells; T cells in the composition and / or expressing the recombinant receptor are CD8 + and CD4 + comprising or consisting of T cells of and / or at least about 30%, at least about 50%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, or at least about 100% CD8 T cells in the composition and / or expressing the recombinant receptor. + comprising or consisting of T cells; and / or at least about 30%, at least 50%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or at least 100%, of the T cells in the composition and / or expressing the recombinant receptor are at least about 30%, at least about 50%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, or at least about 100%, CD4 + comprising or consisting of T cells; and / or The T cells in the composition and / or expressing the recombinant receptor are in a ratio of about 1:1, or about 1:3 to 3:1, or about 1:2 to 2:1 CD4 + T cells and CD8 + comprising or consisting of T cells, A therapeutic composition according to any one of claims 1 to 5.

7. A therapeutic composition comprising T cells expressing a recombinant receptor, wherein the T cells express at least about 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%, or at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least 100% of the total number of T cells in the composition or of the total number of T cells expressing the recombinant receptor in the composition. at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% are surface positive for CCR7 and / or CD27, and optionally T cells in the composition and / or expressing the recombinant receptor are CD8 + comprising or consisting of T cells; T cells in the composition and / or expressing the recombinant receptor are CD4 + comprising or consisting of T cells; T cells in the composition and / or expressing the recombinant receptor are CD8 + and CD4 + comprising or consisting of T cells of and / or at least about 30%, at least 50%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or at least 100% of the T cells in the composition and / or expressing the recombinant receptor are CD8 + comprising or consisting of T cells; and / or at least about 30%, at least about 50%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, or at least about 100% of the T cells in the composition and / or expressing the recombinant receptor are CD4 + comprising or consisting of T cells; and / or The T cells in the composition and / or expressing the recombinant receptor are in a ratio of about 1:1, or about 1:3 to 3:1, or about 1:2 to 2:1 CD4 + T cells and CD8 + comprising or consisting of T cells, The therapeutic composition.

8. 8. The therapeutic composition of any one of claims 1 to 7, wherein the total number of T cells in said composition or the total number of T cells expressing the recombinant receptor in said composition are surface positive for CCR7.

9. 8. The therapeutic composition of any one of claims 1 to 7, wherein the total number of T cells in said composition or the total number of T cells expressing the recombinant receptor in said composition are surface positive for CD27.

10. 8. The therapeutic composition of any one of claims 1 to 7, wherein the total number of T cells in said composition or the total number of T cells expressing the recombinant receptor in said composition are surface positive for CCR7 and CD27.

11. 11. The therapeutic composition of any one of claims 1 to 10, comprising one or more unit doses of cells.

12. The unit dose or composition is 1 x 10 inclusive 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD8 expressing recombinant receptors + Cells (receptors + / CD8 + cells), or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + and / or The unit dose or composition is about 1 x 10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cells, or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + including cells, A therapeutic composition according to any one of claims 1 to 11.

13. The composition or the unit dose is at least about 3×10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells; and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + 13. The therapeutic composition of any one of claims 1 to 12, comprising:

14. The unit dose or composition is 3×10 inclusive 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cellular or total receptors + / CD8 + / CD27 + cells, and / or 3 × 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cellular or total receptors + / CD4 + / CD27 + 14. The therapeutic composition of any one of claims 1 to 13, comprising:

15. the total receptors in the unit dose or the composition + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose or said composition + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptor in said unit dose or said composition + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose or said composition + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + 15. The therapeutic composition of any one of claims 1 to 14, wherein

16. The unit dose or the composition contains a predetermined number of receptors. + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + 16. The therapeutic composition of any one of claims 1 to 15, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

17. A defined number or ratio of cells may also be expressed as CD27 + or CD45RA - 17. The therapeutic composition of claim 16, wherein the cell is a cell of the

18. The unit dose or the composition contains a predetermined number of receptors. + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + 16. The therapeutic composition of any one of claims 1 to 15, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

19. A defined number or ratio of cells may further comprise CCR7 + or CD45RA - 20. The therapeutic composition of claim 18, wherein the cell is a cell of the present invention.

20. The unit dose or composition contains a predetermined number of receptors. + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD8 + / CCR7 + / CD45RA - and / or the unit dose or composition contains a predetermined number of receptor cells. + / CD4 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD45RA - 20. The therapeutic composition of any one of claims 1 to 19, comprising cells.

21. the total receptors in the unit dose or the composition + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose or said composition + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - , receptor + / CD4 + / CCR7 + / CD27 + , or receptor + / CD4 + / CCR7 + / CD45RA - 21. The therapeutic composition of any one of claims 1 to 20, wherein

22. The unit dose or number of cells expressing the recombinant receptor is 1 x 10, inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD3 expressing recombinant receptors + Cells (receptors + / CD3 + cells) or total CD3 + cells; and / or the unit dose or number of cells expressing the recombinant receptor is about 5 x 10 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + 22. The therapeutic composition of any one of claims 1 to 21, comprising cells.

23. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - 23. The therapeutic composition of any one of claims 1 to 22, wherein the cells are the total number of living or viable cells.

24. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The cells are the total number of cells that do not express apoptotic markers and / or are apoptotic marker negative ( - 24. The therapeutic composition of any one of claims 1 to 23, wherein the apoptosis marker is annexin V or activated caspase 3.

25. 30. The therapeutic composition of any one of claims 1 to 29, wherein the recombinant receptor is capable of binding to a target antigen associated with, specific for, and / or expressed in a cell or tissue of a disease, disorder, or condition.

26. 26. The therapeutic composition of claim 25, wherein the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer.

27. Target antigens include αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, also known as CAIX or G250), cancer-testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, and CD12. 3, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutated (EGFRvIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OG D2), ganglioside GD3, glycoprotein 100 (gp100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPCR5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), quinine kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM),27. The therapeutic composition of claim 25 or 26, wherein the antigen is selected from the group consisting of oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigen, or universal tag-related antigen, and / or biotinylated molecules, and / or molecules expressed by HIV, HCV, HBV, or other pathogens.

28. 28. The therapeutic composition of any one of claims 1 to 27, wherein the recombinant receptor is a chimeric antigen receptor (CAR).

29. 29. The therapeutic composition of any one of claims 1 to 28, wherein the recombinant receptor comprises an extracellular domain comprising an antigen-binding domain, and an intracellular signaling region comprising an intracellular signaling domain, and further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region.

30. 29. The therapeutic composition of claim 28, wherein the antigen-binding domain is or comprises an antibody or an antibody fragment thereof, optionally a single-chain fragment.

31. 31. The therapeutic composition of claim 30, wherein the fragment comprises an scFv.

32. The intracellular signaling domain is the intracellular signaling domain of the CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof 32. The therapeutic composition of any one of claims 29 to 31, which is or comprises:

33. 33. The therapeutic composition of any one of claims 29-32, wherein the intracellular signaling region further comprises a costimulatory signaling domain.

34. 34. The therapeutic composition of claim 33, wherein the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule or a signaling portion thereof.

35. 35. The therapeutic composition of claim 33 or 34, wherein the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof.

36. 36. The therapeutic composition of any one of claims 33-35, wherein the costimulatory signaling domain is located between the transmembrane domain and the intracellular signaling domain.

37. The therapeutic composition of any one of claims 1 to 36, wherein the T cells are primary T cells obtained from the subject.

38. The therapeutic composition of any one of claims 1 to 37, wherein the T cells are autologous to the subject.

39. 38. The therapeutic composition of any one of claims 1 to 37, wherein the T cells are allogeneic to the subject.

40. The therapeutic cell composition of any one of claims 37 to 39, wherein the subject has cancer, and optionally, the cancer is leukemia or lymphoma.

41. 41. The therapeutic cell composition of claim 40, wherein the subject is identified or known to have a high tumor burden at the time the cells are obtained.

42. The sum of two-way product (SPD) of the tumor in the subject is 30cm 2 Super, 40cm 2 Super, 50cm 2 Super, 60cm 2 Over or 70cm 2 Over or about 30 cm 2 Super, about 40cm 2 Super, about 50cm 2 Super, about 60cm 2 Over or about 70cm 2 if greater than 100, the subject has a high tumor burden; and / or 42. The therapeutic cell composition of claim 41, wherein a subject has a high tumor burden when C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 milligrams / liter, greater than 10 milligrams / liter, greater than 15 milligrams / liter, greater than 20 milligrams / liter, greater than 25 milligrams / liter, greater than 30 milligrams / liter, greater than 40 milligrams / liter, or greater than 50 milligrams / liter, or greater than about 5 milligrams / liter, greater than about 10 milligrams / liter, greater than about 15 milligrams / liter, greater than about 20 milligrams / liter, greater than about 25 milligrams / liter, greater than about 30 milligrams / liter, greater than about 40 milligrams / liter, or greater than about 50 milligrams / liter.

43. The SPD of the tumor in the subject is 50 cm 2 Over or about 50cm 2 43. The therapeutic cell composition of claim 42, wherein the subject has a high tumor burden.

44. 44. The therapeutic cell composition of claim 42 or 43, wherein the subject has a high tumor burden when CRP is greater than or about greater than 20 milligrams / liter in the biological sample.

45. A container containing the composition of any one of claims 1 to 44; instructions for administering the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses, to a subject having a disease or condition; Manufactured goods including.

46. A container containing one or more unit doses of cells present in one or more therapeutic compositions, the unit doses expressing (and optionally engineered to express) a plurality of CD8 + and / or CD4 + T cells, wherein a unit dose of cells contains a defined number of recombinant receptor-expressing CD8 T cells expressing (optionally surface-expressing) CC chemokine receptor type 7 (CCR7). + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + a container containing the cells and another subset of cells in the composition; instructions for administering to a subject having a disease or condition the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses; Manufactured goods including.

47. A container containing one or more unit doses of cells present in one or more therapeutic compositions, the unit doses expressing (and optionally engineered to express) a plurality of CD8 + and / or CD4 + T cells, wherein a unit dose of cells contains a defined number of recombinant cluster of differentiation 27 (CD27)-expressing (optionally surface-expressing) CD8 + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + a container containing the cells and another subset of cells in the composition; instructions for administering to a subject having a disease or condition the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses; Manufactured goods including.

48. 48. The article of manufacture of claim 46 or 47, wherein a unit dose of cells comprises the composition of any one of claims 1-44.

49. administering to a subject having a disease or condition one or more unit doses of the therapeutic composition of claim 1 or 7, or a plurality of CD8 + and / or CD4 + A therapeutic composition comprising T cells, and / or a method of treatment comprising administering an amount corresponding to such a unit dose, optionally comprising: A unit dose of cells contains a defined number of recombinant receptor-expressing CD8 + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + and / or comprising a subset of cells in the composition; Among the group of subjects treated according to the method, cells expressing CCR7 (CCR7 + The method of treatment, wherein the number or ratio of cells (e.g., serotype, phenotype, or phenotype) varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

50. A unit dose of cells produces a defined number of CD8 + / CCR7 + cells and / or a unit dose of cells contains a defined number of CD4 + / CCR7 + 50. The method of claim 49, comprising a cell.

51. administering to a subject having a disease or condition one or more unit doses of the therapeutic composition of claim 1 or 7, or a plurality of CD8 + and / or CD4 + A therapeutic composition comprising T cells, and / or a method of treatment comprising administering an amount corresponding to such a unit dose, optionally comprising: A unit dose of cells contains a defined number of recombinant cluster of differentiation 27 (CD27) receptor-expressing CD8 + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + and / or comprising a subset of cells in the composition; Among the group of subjects treated according to the method, cells expressing CD27 (CD27 + The method of treatment, wherein the number or ratio of cells (e.g., serotype, phenotype, or phenotype) varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

52. A unit dose of cells produces a defined number of CD8 + / CD27 + cells and / or a unit dose of cells contains a defined number of CD4 + / CD27 + 52. The method of claim 51, comprising a cell.

53. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD8 expressing recombinant receptors + Cells (receptors + / CD8 + cells), or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells; The unit dose is about 1 x 10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cells, or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + containing cells; and / or The unit dose is at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells, and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + Including, 53. The method of any one of claims 49 to 52.

54. The unit dose is 3 x 10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cellular or total receptors + / CD8 + / CD27 + cells, and / or 3 × 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cellular or total receptors + / CD4 + / CD27 + 54. The method of any one of claims 49 to 53, comprising:

55. Total receptors in the unit dose + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptors in said unit dose + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + The method of any one of claims 49 to 54, wherein

56. A unit dose of cells contains a defined number of receptors. + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + 56. The method of any one of claims 49, 50, and 53-55, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

57. A unit dose of cells contains a defined number of receptors. + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + 56. The method of any one of claims 51 to 55, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

58. A unit dose of cells contains a defined number of receptors. + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD8 + / CCR7 + / CD45RA - and / or a unit dose of cells contains a predetermined number of receptors + / CD4 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD45RA - 58. The method of claim 56 or 57, comprising a cell.

59. Total receptors in the unit dose + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD8 + / CCR7 + / CD45RA - and / or the total receptors in said unit dose + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD45RA - 59. The method of claim 58, wherein:

60. 10. A method of treatment comprising administering to a subject having a disease or condition one or more unit doses of the composition of claim 1 or 7.

61. A unit dose of cells contains a defined number of recombinant receptor-expressing CD8 + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + comprising a subset of cells in the composition; and Among the group of subjects treated according to the method, the number of cells expressing CCR7 (CCR7 + the number or ratio of cells) varies by 40% or less, 30% or less, 20% or less, 10% or less, or 5% or less; 61. The method of claim 60.

62. administering to a subject having a disease or condition a plurality of CD8 + and / or CD4 + A therapeutic composition comprising T cells and / or a method of treatment comprising administering an amount corresponding to such a unit dose, A unit dose of cells contains a defined number of recombinant receptor-expressing CD8 + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + cells and another subset of cells in the composition; and Among the group of subjects treated according to the method, cells expressing CCR7 (CCR7 + the number or ratio of cells) varies by 40% or less, 30% or less, 20% or less, 10% or less, or 5% or less; The treatment method.

63. A unit dose of cells produces a defined number of CD8 + / CCR7 + / CD27 + cells and / or a unit dose of cells contains a defined number of CD4 + / CCR7 + / CD27 + 63. The method of any one of claims 58 to 62, comprising a cell.

64. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD8 expressing recombinant receptors + Cells (receptors + / CD8 + cells), or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + Contains cells; The unit dose is about 1 x 10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cells, or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + containing cells; and / or The unit dose is at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells; and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + including cells, 64. The method of any one of claims 58 to 63.

65. The unit dose is 3 x 10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells; and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + 65. The method of any one of claims 58 to 64, comprising a cell.

66. Total receptors in the unit dose + at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + 66. The method of any one of claims 58 to 65, wherein

67. A unit dose of cells is mixed with a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + 67. The method of any one of claims 58-66, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

68. A defined number of cells further express or do not express CD45RA, and optionally a defined number of cells further express CD45RA - The method of any one of claims 49 to 67, wherein the cell is a cell.

69. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD3 expressing recombinant receptors + Cells (receptors + / CD3 + cells) or total CD3 + 69. The method of any one of claims 49 to 68, comprising a cell.

70. The unit dose is about 5×10 8 Below, approximately 1×10 8 Below, approximately 5×10 7 Below, approximately 1×10 7 Below, approximately 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + 70. The method of any one of claims 49 to 69, comprising a cell.

71. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - 71. The method of claim 70, wherein the cells are the total number of live or viable cells.

72. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The cells are the total number of cells that do not express apoptotic markers and / or are apoptotic marker negative ( - 72. The method of any one of claims 49 to 71, wherein the apoptosis marker is annexin V or activated caspase 3.

73. 73. The method of any one of claims 49-72, comprising administering a plurality of unit doses contained in a plurality of separate compositions.

74. Multiple distinct compositions are identified as CD8 + T cells and CD4 + a first composition comprising one of CD8 T cells; + T cells and CD4 + and a second composition comprising the other of said T cells.

75. The first composition comprises a CD8 + 75. The method of claim 74, comprising T cells.

76. The first composition comprises CD4 + 75. The method of claim 74, comprising T cells.

77. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 77. The method of any one of claims 49-76, comprising administering the T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, no more than 48 hours apart.

78. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 78. The method of claim 77, wherein the T cells or one or more unit doses thereof and / or compositions containing an amount corresponding to such one or more unit doses are administered no more than 36 hours apart, no more than 24 hours apart, no more than 12 hours apart, no more than 6 hours apart, no more than 4 hours apart, no more than 2 hours apart, no more than 1 hour apart, or no more than 30 minutes apart.

79. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 79. The method of claim 77 or 78, wherein the T cells or one or more unit doses thereof and / or compositions containing an amount corresponding to such one or more unit doses are administered no more than 2 hours apart, no more than 1 hour apart, no more than 30 minutes apart, no more than 15 minutes apart, no more than 10 minutes apart, or no more than 5 minutes apart, or simultaneously.

80. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, + 80. The method of any one of claims 73 to 79, comprising administering before administration a composition containing the cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

81. CD4 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses are administered to a subject in need thereof. + 80. The method of any one of claims 73 to 79, comprising administering before administration a composition containing the cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

82. (a) Multiple CD8s engineered to express recombinant receptors + and / or CD4 + In a therapeutic composition comprising T cells, T cells expressing the recombinant receptor and CCR7 (receptor + / CCR7 + assessing the number, percentage, or ratio of (b) Receptor + / CCR7 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit doses contain a predetermined number of recombinant receptor-expressing CD8 chemokine receptor type 7 (CCR7)-expressing cells; + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + the step of including a different subset of cells in the composition. A method for determining a unit dose of engineered T cells for treating a subject, comprising:

83. (a) Multiple CD8s engineered to express recombinant receptors + and / or CD4 + In a therapeutic composition comprising T cells, T cells expressing the recombinant receptor and CD27 (receptor + / CD27 + assessing the number, percentage, or ratio of (b) Receptor + / CCR7 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit doses contain a predetermined number of recombinant cluster of differentiation 27 (CD27)-expressing receptor-expressing CD8 + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant CD27-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + the step of including a different subset of cells in the composition. A method for determining a unit dose of engineered T cells for treating a subject, comprising:

84. (a) Multiple CD8s engineered to express recombinant receptors + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and T cells expressing CCR7 and / or CD27 (receptor + / CCR7 + and / or receptors + / CD27 + assessing the number, percentage, or ratio of (b) filling a container with all or a portion of the T cell composition, and optionally another solution, to achieve a unit dose of the T cell composition, wherein the unit dose contains a predetermined number of recombinant receptor-expressing CD8 T cells expressing CC chemokine receptor type 7 (CCR7). + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + cells and another subset of cells in the composition, or a defined number of recombinant cluster of differentiation 27 (CD27) receptor-expressing CD8 + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant CD27-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + the step of comprising a cell and another subset of cells in the composition. A method of producing a composition comprising a unit dose of a T cell composition, comprising:

85. filling a container with all or a portion of the T cell composition to achieve a unit dose of the T cell composition, the T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition, the unit dose comprising a predetermined number of recombinant receptor-expressing CD8 T cells that express CC chemokine receptor type 7 (CCR7); + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + A method for producing a therapeutic composition comprising a unit dose of a T cell composition comprising cells and another subset of cells in the composition.

86. filling a container with all or a portion of the T cell composition to achieve a unit dose of the T cell composition, the T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition, the unit dose comprising a predetermined number of recombinant receptor-expressing CD8 T cells that express cluster of differentiation 27 (CD27). + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant CD27-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + A method for producing a therapeutic composition comprising a unit dose of a T cell composition comprising cells and another subset of cells in the composition.

87. A unit dose of cells produces a defined number of CD8 + / CCR7 + cells and / or a unit dose of cells contains a defined number of CD4 + / CCR7 + 87. The method of any one of claims 83 to 86, comprising a cell.

88. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD8 expressing recombinant receptors + Cells (receptors + / CD8 + cells), or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + Contains cells; The unit dose is about 1 x 10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cells, or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, or total receptors + / CD4 + / CD27 + containing cells; and / or The unit dose is at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells, and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + Including, 88. The method of any one of claims 82 to 87.

89. The unit dose is 3 x 10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cellular or total receptors + / CD8 + / CD27 + cells, and / or 3 × 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cellular or total receptors + / CD4 + / CD27 + 89. The method of any one of claims 82 to 88, comprising:

90. Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + or the total receptors in said unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + The method of any one of claims 82 to 89, wherein

91. A unit dose of cells is mixed with a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + 91. The method of any one of claims 82-90, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

92. The unit dose further comprises a predetermined number of cells, including cells that express or do not express CD27 and / or CD45, and optionally, the unit dose + or CD45RA - 92. The method of any one of claims 82-91, further comprising a defined number of cells comprising the cells.

93. In the therapeutic composition, the number, percentage, or ratio of T cells that express or do not express CD27 and / or CD45RA, optionally, + or CD45RA - The method of any one of claims 82-92, further comprising assessing the number, percentage, or ratio of T cells that are

94. A unit dose of cells contains a defined number of receptors. + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD8 + / CCR7 + / CD45RA - A unit dose of cells may contain a predetermined number of receptors, and / or cells. + / CD4 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD45RA - 94. The method of claim 92 or 93, comprising a cell.

95. Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in the unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - , receptor + / CD4 + / CCR7 + / CD27 + , or receptor + / CD4 + / CCR7 + / CD45RA - The method of any one of claims 92 to 95, wherein

96. (a) Multiple CD8s engineered to express recombinant receptors + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and T cells expressing CCR7 and CD27 (receptor + / CCR7 + / CD27 + assessing the number, percentage, or ratio of (b) Receptor + / CCR7 + / CD27 + determining one or more unit doses of cells for administration to a subject having a disease or condition based on the number, percentage, or ratio of cells, wherein the unit doses contain a predetermined number of recombinant receptor-expressing CD8 + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + the step of including a different subset of cells in the composition. A method for determining a unit dose of engineered T cells for treating a subject, comprising:

97. (a) Multiple CD8s engineered to express recombinant receptors + and / or CD4 + In a therapeutic composition comprising T cells, the recombinant receptor and T cells expressing CCR7 and CD27 (receptor + / CCR7 + / CD27 + assessing the number, percentage, or ratio of (b) filling all or a portion of the composition, and optionally another solution, into a container to achieve a unit dose of the T cell composition, the unit dose containing a predetermined number of recombinant receptor-expressing CD8 T cells expressing CCR7 and CD27. + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + the step of comprising a cell and another subset of cells in the composition. A method of producing a composition comprising a unit dose of a T cell composition, comprising:

98. filling a container with all or a portion of the T cell composition to achieve a unit dose of the T cell composition, the T cell composition comprising T cells comprising a recombinant receptor that specifically binds to an antigen associated with a disease or condition, the unit dose comprising a predetermined number of recombinant receptor-expressing CD8 T cells that express CCR7 and CD27. + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + A method for producing a therapeutic composition comprising a unit dose of a T cell composition comprising cells and another subset of cells in the composition.

99. A unit dose of cells produces a defined number of CD8 + / CCR7 + / CD27 + cells and / or a unit dose of cells contains a defined number of CD4 + / CCR7 + / CD27 + 99. The method of any one of claims 96 to 98, comprising a cell.

100. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD8 expressing recombinant receptors + Cells (receptors + / CD8 + cells), or total CD4 expressing recombinant receptors + Cells (receptors + / CD4 + cells), total receptors + / CD8 + / CCR7 + / CD27 + Cells or total receptors + / CD4 + / CCR7 + / CD27 + cells; said unit dose comprising about 1 x 10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD8 + Cells or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells or total receptors + / CD4 + / CCR7 + / CD27 + and / or said unit dose comprises at least about 3 x 10 cells; 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + 100. The method of any one of claims 96 to 99, comprising a cell.

101. The unit dose is 3 x 10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells, and / or 3 × 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + 101. The method of any one of claims 96 to 100, comprising a cell.

102. Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + The method of any one of claims 96 to 103, wherein

103. A unit dose of cells is mixed with a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + 103. The method of any one of claims 96-102, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

104. The unit dose further comprises a predetermined number of cells, including cells that express or do not express CD45, and optionally, the unit dose comprises a CD45RA - The method of any one of claims 96-103, further comprising a defined number of cells comprising the cells.

105. In the therapeutic composition, the number, percentage, or ratio of T cells that express or do not express CD45RA, optionally, - The method of any one of claims 96-104, further comprising assessing the number, percentage, or ratio of T cells that are

106. i) Identifying target percentages of CD8+ cells that express or do not express CCR7, CD27, and / or CD45RA from a biological sample from the subject. + T cells and / or target percentages of CD4 T cells that may or may not express CCR7, CD27, and / or CD45RA. + providing an input composition comprising T cells; ii) introducing a polynucleotide encoding a recombinant receptor into the input composition; and iii) stimulating the cells in the input composition before, during, and / or after said introducing, wherein the stimulating comprises incubating the cells in the presence of one or more stimulating agents, wherein the stimulating results in activation and / or proliferation of the cells.

1. A method for producing a cell composition comprising genetically engineered cells, comprising: Recombinant receptor expression (receptor expression) with or without CCR7, CD27, and / or CD45RA in defined ratios. + ) CD8 + T cells with receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios. + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + The method produces an output composition comprising T cells and any other subset of cells in the composition.

107. 1. A method for generating a cell composition comprising genetically engineered cells, comprising introducing a polynucleotide encoding a recombinant receptor into an input composition, The input composition comprises a target percentage of CD8+ cells that express or do not express CCR7, CD27, and / or CD45RA provided from a biological sample from a subject. + T cells and / or target percentages of CD4 T cells that may or may not express CCR7, CD27, and / or CD45RA. + the input composition comprises T cells, and is stimulated before, during, and / or after the introduction by incubating the cells in the presence of one or more stimulatory agents, the stimulation resulting in activation and / or proliferation of the cells; and The method comprises the step of producing recombinant receptor expression vectors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios (receptors + ) CD8 + T cells with receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios. + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + producing an output composition comprising T cells and another subset of cells in said composition; The method.

108. i) A target number of CD8+ cells expressing or not expressing CCR7, CD27, and / or CD45RA are isolated from a biological sample obtained from the subject. + T cells and / or target numbers of CD4 + isolating T cells, thereby generating an input composition; ii) introducing a polynucleotide encoding a recombinant receptor into the input composition; and iii) stimulating the cells in the input composition before, during, and / or after said introducing, wherein the stimulating comprises incubating the cells in the presence of one or more stimulating agents, wherein the stimulating results in activation and / or proliferation of the cells.

1. A method for producing a cell composition comprising genetically engineered cells, comprising: Recombinant receptor expression (receptor expression) with or without CCR7, CD27, and / or CD45RA in defined ratios. + ) CD8 + T cells with receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios. + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + The method produces an output composition comprising T cells and any other subset of cells in the composition.

109. 1. A method for generating a cell composition comprising genetically engineered cells, comprising introducing a polynucleotide encoding a recombinant receptor into an input composition, The input composition contains target numbers of CD8 + T cells and / or target numbers of CD4 + T cells are generated by isolating them from a biological sample obtained from a subject, and the input composition is stimulated before, during, and / or after the introduction by incubating the cells in the presence of one or more stimulatory agents, which stimulation results in activation and / or proliferation of the cells; and The method comprises the step of producing recombinant receptor expression vectors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios (receptors + ) CD8 + T cells with receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + T cells and / or receptors expressing or not expressing CCR7, CD27, and / or CD45RA in defined ratios. + / CD8 + T cells and / or receptors that may or may not express CCR7, CD27, and / or CD45RA + / CD4 + producing an output composition comprising T cells and another subset of cells in said composition; The method.

110. The input composition contains a target percentage of CD8+ cells that express or do not express CCR7, CD27, and / or CD45RA. + T cells and / or target percentages of CD4 T cells that may or may not express CCR7, CD27, and / or CD45RA. + The method of any one of claims 106 to 109, comprising T cells.

111. The input composition is a target percentage of CD4 + / CCR7 + , CD4 + / CD27 + , CD4 + / CCR7 + / CD27 + , CD4 + / CCR7 + / CD45RA - , CD4 + / CCR7 + / CD45RA + , CD8 + / CCR7 + , CD8 + / CD27 + , CD8 + / CCR7 + / CD27 + , CD8 + / CCR7 + / CD45RA - , and / or CD8 + / CCR7 + / CD45RA + 111. The method of any one of claims 106-110, comprising cells of

112. the output composition comprising: A defined ratio of receptors + / CD8 + / CCR7 + , receptor + / CD8 + / CD27 + , receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - and / or receptor + / CD8 + / CCR7 + / CD45RA + Cells and receptors + / CD4 + / CCR7 + , receptor + / CD4 + / CD27 + , receptor + / CD4 + / CCR7 + / CD27 + , receptor + / CD4 + / CCR7 + / CD45RA - and / or receptor + / CD4 + / CCR7 + / CD45RA + cells of; or A defined ratio of receptors + / CD8 + / CCR7 + , receptor + / CD8 + / CD27 + , receptor + / CD8 + / CCR7 + / CD27 + , receptor + / CD8 + / CCR7 + / CD45RA - and / or receptor + / CD8 + / CCR7 + / CD45RA + Cells and / or receptors of + / CD4 + / CCR7 + , receptor + / CD4 + / CD27 + , receptor + / CD4 + / CCR7 + / CD27 + , receptor + / CD4 + / CCR7 + / CD45RA - and / or receptor + / CD4 + / CCR7 + / CD45RA + and another subset of cells in the composition 112. The method of any one of claims 106 to 111, comprising:

113. determining one or more unit doses of the T cell composition for administration to a subject having a disease or condition, the unit doses comprising: A defined number of recombinant receptor-expressing CD8 cells expressing CC chemokine receptor type 7 (CCR7) + T cells (receptor + / CD8 + / CCR7 + cells), and / or a defined number of recombinant CCR7-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + Cells and / or Receptors + / CD4 + / CCR7 + cells and another subset of cells in the composition; A defined number of recombinant CD8 expressing cluster of differentiation 27 (CD27) receptors + T cells (receptor + / CD8 + / CD27 + cells), and / or a defined number of recombinant CD27-expressing receptor-expressing CD4 + T cells (receptor + / CD4 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CD27 + Cells and / or Receptors + / CD4 + / CD27 + cells and a different subset of cells in the composition; or A defined number of recombinant receptor-expressing CD8 expressing CCR7 and CD27 + T cells (receptor + / CD8 + / CCR7 + / CD27 + cells), and / or a defined number of recombinant receptor-expressing CD4 + T cells (receptor + / CD4 + / CCR7 + / CD27 + cells), and / or a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + cells, and / or receptors in a defined ratio + / CD8 + / CCR7 + / CD27 + Cells and / or Receptors + / CD4 + / CCR7 + / CD27 + cells and another subset of cells in the composition 113. The method of any one of claims 106-112, comprising all or a portion of an output composition comprising:

114. A unit dose of cells induces a defined number of CD8 + / CCR7 + cells, CD4 + / CCR7 + cells, CD8 + / CD27 + cells, CD4 + / CD27 + cells, CD8 + / CCR7 + / CD27 + cells, and / or CD4 + / CCR7 + / CD27 + 114. The method of claim 113, comprising a cell.

115. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + Cells, or total receptors + / CD4 + Cells, total receptors + / CD8 + / CCR7 + Cells, total receptors + / CD4 + / CCR7 + Cells, total receptors + / CD8 + / CD27 + Cells, total receptors + / CD4 + / CD27 + Cells, total receptors + / CD8 + / CCR7 + / CD27 + Cells, or total receptors + / CD4 + / CCR7 + / CD27 + 115. The method of claim 113 or 114, comprising a cell.

116. The unit dose is at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells; and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + and / or at least about 3×10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD8 + / CCR7 + / CD27 + cells and / or at least about 3 x 10 6 , at least about 4 × 10 6 , at least about 5 × 10 6 , at least about 6 × 10 6 , at least about 7 × 10 6 , at least about 8 × 10 6 , at least about 9 × 10 6 , or at least about 1 × 10 7 Total receptors + / CD4 + / CCR7 + / CD27 + containing cells; and / or The unit dose is 3 x 10 inclusive. 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + Cell or total receptor + / CD8 + / CD27 + cells; and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + Cell or total receptor + / CD4 + / CD27 + and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD8 + / CCR7 + / CD27 + cells; and / or 3 x 10 6 Or about 3 x 10 6 and 2.5 x 10 7 Or about 2.5 x 10 7 Between 4×10 6 Or about 4 x 10 6 and 2×10 7 Or about 2 x 10 7 Between or 5 x 10 6 Or about 5 x 10 6 and 1×10 7 Or about 1 x 10 7 Total receptors between + / CD4 + / CCR7 + / CD27 + including cells, 116. The method of any one of claims 113 to 115.

117. Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + or receptor + / CD8 + / CD27 + is; Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + or receptor + / CD4 + / CD27 + and / or Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD8 + / CCR7 + / CD27 + or receptor + / CD4 + / CCR7 + / CD27 + That is, 117. The method of any one of claims 113 to 116.

118. A unit dose of cells is mixed with a defined ratio of receptors + / CD8 + / CCR7 + Cells and Receptors + / CD4 + / CCR7 + cells, optionally in a ratio of 1:1 or about 1:1, or about 1:3 to about 3:1; and / or in a defined ratio of receptors + / CD8 + / CD27 + Cells and Receptors + / CD4 + / CD27 + cells, optionally in a ratio of 1:1 or about 1:1, or about 1:3 to about 3:1; and / or in a defined ratio of receptors + / CD8 + / CCR7 + / CD27 + Cells and Receptors + / CD4 + / CCR7 + / CD27 + 118. The method of any one of claims 113-117, comprising cells, optionally at a ratio of 1:1 or about 1:1, or about 1:3 to about 3:

1.

119. The unit dose further comprises a predetermined number of cells, including cells that express or do not express CD45, and optionally, the unit dose comprises a CD45RA - 119. The method of any one of claims 113-118, further comprising a defined number of cells comprising the cells.

120. A unit dose of cells contains a defined number of receptors + / CD4 + / CCR7 + / CD45RA - Cells and / or Receptors + / CD8 + / CCR7 + / CD45RA - 111. The method of claim 110, comprising cells.

121. Total receptors in the unit dose + At least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the cells or total receptors in said unit dose + Approximately 15% to 90%, 20% to 80%, 30% to 70%, or 40% to 60% of the cells, inclusive, are receptor-positive. + / CD4 + / CCR7 + / CD45RA - or receptor + / CD8 + / CCR7 + / CD45RA - The method of any one of claims 113 to 120, wherein

122. Prior to provision or isolation, CD8 + T cells and / or CD4 with or without CCR7, CD27, and / or CD45RA expression + 122. The method of any one of claims 106-121, comprising determining the number, number by volume, number by weight, and / or percentage of T cells.

123. One or more stimulatory agents may stimulate T cells, CD4 + T cells and / or CD8 + capable of activating T cells; capable of inducing signals through the TCR complex; and / or capable of activating T cells, CD4 + T cells and / or CD8 + The method of any one of claims 106 to 122, wherein the method is capable of inducing proliferation of T cells.

124. The method of any one of claims 106-123, wherein the one or more stimulating agents bind to a member of the TCR complex, optionally comprising a primary agent that specifically binds to CD3.

125. 125. The method of claim 124, wherein the one or more stimulatory agents further comprise a secondary agent that specifically binds to a T cell costimulatory molecule.

126. The method of claim 125, wherein the costimulatory molecule is selected from the group consisting of CD28, CD137 (4-1BB), OX40, or ICOS.

127. 127. The method of claim 125 or 126, wherein the primary and secondary agents comprise antibodies, and optionally, one or more stimulatory agents comprise incubating with the anti-CD3 antibody and the anti-CD28 antibody.

128. The method of any one of claims 106-127, wherein the one or more stimulating agents are present on the surface of a solid support, optionally a bead.

129. The method of any one of claims 106-128, wherein the one or more stimulatory agents are selected from the group consisting of a CD3-binding molecule; a CD28-binding molecule; recombinant IL-2; recombinant IL-15; and recombinant IL-7, a vaccine comprising an antigen that is specifically recognized by an antigen receptor, and an anti-idiotypic antibody that specifically binds to an antigen receptor, or a combination thereof.

130. 130. The method of any one of claims 82-129, wherein the therapeutic composition is the composition of claim 1 or 7.

131. 131. The method of any one of claims 82-130, further comprising administering to a subject having the disease or condition the therapeutic composition, optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

132. Among the group of subjects treated according to the method, the number of cells expressing CCR7 (CCR7 + 132. The method of any one of claims 82-131, wherein the number or ratio of cells (cells) varies by no more than 40%, no more than 30%, no more than 20%, no more than 10%, or no more than 5%.

133. The unit dose is 1 x 10 inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between total CD3 expressing recombinant receptors + Cells (receptors + / CD3 + cells) or total CD3 + 82- and / or unit doses of about 5 x 10 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + / CD3 + Cellular or total CD3 + The method of any one of claims 82-132, comprising a cell.

134. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The method of any one of claims 82-133, wherein the cells are the total number of live or viable cells.

135. CD3 + Total number of cells, receptors + / CD3 + Total number of cells, receptors + / CD8 + Total number of cells, receptors + / CD4 + Total number of cells, receptors + / CD8 + / CCR7 + Total number of cells, receptors + / CD4 + / CCR7 + Total number of cells, receptors + / CD8 + / CD27 + Total number of cells, receptors + / CD4 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD27 + Total number of cells, receptors + / CD4 + / CCR7 + / CD27 + Total number of cells, receptors + / CD8 + / CCR7 + / CD45RA - Total number of cells and / or receptors + / CD4 + / CCR7 + / CD45RA - The cells are the total number of cells that do not express apoptotic markers and / or are apoptotic marker negative ( - 135. The method of any one of claims 82-134, wherein the total number of cells that are apoptotic is 100% or less, and the apoptotic marker is annexin V or activated caspase 3.

136. 136. The method of any one of claims 131-135, comprising administering a plurality of unit doses contained in a plurality of separate compositions.

137. Multiple distinct compositions are identified as CD8 + T cells and CD4 + a first composition comprising one of CD8 T cells; + T cells and CD4 + and a second composition comprising the other of said T cells.

138. The first composition comprises a CD8 + The method of claim 137, comprising T cells.

139. The first composition comprises CD4 + The method of claim 137, comprising T cells.

140. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 140. The method of any one of claims 131-139, comprising administering, no more than 48 hours apart, a composition containing the T cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

141. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 141. The method of claim 140, wherein the T cells or one or more unit doses thereof and / or compositions containing an amount corresponding to such one or more unit doses are administered no more than 36 hours apart, no more than 24 hours apart, no more than 12 hours apart, no more than 6 hours apart, no more than 4 hours apart, no more than 2 hours apart, no more than 1 hour apart, or no more than 30 minutes apart.

142. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 142. The method of claim 140 or 141, wherein the T cells or one or more unit doses thereof and / or compositions containing an amount corresponding to such one or more unit doses are administered no more than 2 hours apart, no more than 1 hour apart, no more than 30 minutes apart, no more than 15 minutes apart, no more than 10 minutes apart, or no more than 5 minutes apart, or simultaneously.

143. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, + 143. The method of any one of claims 136-142, comprising administering before administering a composition containing the cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

144. CD4 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses are administered to a subject in need thereof. + 143. The method of any one of claims 136-142, comprising administering before administering a composition containing the cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

145. The method of any one of claims 49-144, wherein the recombinant receptor is capable of binding to a target antigen associated with, specific for, and / or expressed in a cell or tissue of the disease, disorder, or condition.

146. 146. The method of claim 145, wherein the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer.

147. Target antigens include αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, also known as CAIX or G250), cancer-testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, and CD12. 3, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutated (EGFRvIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OG D2), ganglioside GD3, glycoprotein 100 (gp100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPCR5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), quinine kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM),147. The method of claim 145 or 146, wherein the antigen is selected from among oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigen, or universal tag-related antigen, and / or biotinylated molecule, and / or molecule expressed by HIV, HCV, HBV or other pathogen.

148. The method of any one of claims 49-147, wherein the recombinant receptor is a chimeric antigen receptor (CAR).

149. 149. The method of any one of claims 48-148, wherein the recombinant receptor comprises an extracellular domain comprising an antigen-binding domain, and an intracellular signaling region comprising an intracellular signaling domain, and further comprises a transmembrane domain disposed between said extracellular domain and said intracellular signaling region.

150. 150. The method of claim 149, wherein the antigen-binding domain is or comprises an antibody or an antibody fragment thereof, optionally a single-chain fragment.

151. The method of claim 150, wherein the fragment comprises an scFv.

152. The intracellular signaling domain is the intracellular signaling domain of the CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof 152. The method of any one of claims 149-151, wherein said method is or comprises:

153. The method of any one of claims 149-152, wherein the intracellular signaling region further comprises a costimulatory signaling domain.

154. The method of claim 153, wherein the costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof.

155. The method of claim 153 or 154, wherein the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof.

156. 156. The method of any one of claims 153-155, wherein the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

157. The method of any one of claims 49-156, wherein the T cells are primary T cells obtained from the subject.

158. The method of any one of claims 49-157, wherein the T cell is autologous to the subject.

159. The method of any one of claims 49-157, wherein the T cells are allogeneic to the subject.

160. The method of any one of claims 157-179, wherein the subject has cancer, and optionally the cancer is leukemia or lymphoma.

161. The method of claim 160, wherein the subject is identified or known to have a high tumor burden at the time the cells are obtained.

162. The sum of two-way product (SPD) of the tumor in the subject is 30cm 2 Super, 40cm 2 Super, 50cm 2 Super, 60cm 2 Over or 70cm 2 Over or about 30cm 2 Super, about 40cm 2 Super, about 50cm 2 Super, about 60cm 2 Over or about 70cm 2 if greater than 100, the subject has a high tumor burden; and / or a subject has a high tumor burden if C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 milligrams / liter, greater than 10 milligrams / liter, greater than 15 milligrams / liter, greater than 20 milligrams / liter, greater than 25 milligrams / liter, greater than 30 milligrams / liter, greater than 40 milligrams / liter, or greater than 50 milligrams / liter, or greater than about 5 milligrams / liter, greater than about 10 milligrams / liter, greater than about 15 milligrams / liter, greater than about 20 milligrams / liter, greater than about 25 milligrams / liter, greater than about 30 milligrams / liter, greater than about 40 milligrams / liter, or greater than about 50 milligrams / liter; 162. The method of claim 161.

163. The SPD of the tumor in the subject is 50 cm 2 Over or about 50cm 2 163. The method of claim 162, wherein if ≥ 1, the subject has a high tumor burden.

164. 164. The method of claim 162 or 163, wherein the subject has high tumor burden if CRP is greater than or greater than about 20 milligrams / liter in the biological sample.

165. 165. An output composition produced by the method of any one of claims 106-164.

166. 165. A unit dose determined or produced by the method of any one of claims 82-164.

167. 167. A pharmaceutical composition comprising the unit dose of claim 166.

168. 168. The pharmaceutical composition of claim 167, further comprising a pharmaceutical carrier.

169. 168. A method of treatment comprising administering to a mammalian subject all or a portion of the output composition of claim 165, the unit dose of claim 166, or the pharmaceutical composition of claim 167 or 168.

170. 168. Use of all or a portion of the output composition of claim 165, the unit dose of claim 166, or the pharmaceutical composition of claim 167 or 168 to treat cancer.

171. 168. Use of all or a portion of the output composition of claim 165, the unit dose of claim 166, or the pharmaceutical composition of claim 167 or 168 in the manufacture of a medicament for treating cancer.

172. 169. The output composition of claim 165, the unit dose of claim 166, or the pharmaceutical composition of claim 167 or 168 for use in the treatment of cancer.

173. A therapeutic cell composition comprising T cells expressing a recombinant receptor that binds to an antigen, which, after stimulation with a stimulatory agent, At least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%, or at least about 70%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, or at least about at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 70%, 80%, 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha; and / or at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are multifunctional for producing two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha; The therapeutic cell composition.

174. At least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor.

174. The therapeutic cell composition of claim 173, wherein at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha.

175. 175. The therapeutic cell composition of claim 173 or 174, wherein at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, or more of the total number of CD4+ T cells expressing the recombinant receptor in said composition are multifunctional for producing two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

176. 176. The therapeutic cell composition of any one of claims 173-175, wherein at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in said composition are multifunctional for producing the cytokines interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

177. The therapeutic cell composition of any one of claims 173-176, wherein the stimulatory agent is a non-specific or non-antigen-dependent T cell stimulatory agent.

178. The therapeutic cell composition of claim 177, wherein the non-specific or non-antigen-dependent T cell stimulatory agent is a polyclonal stimulatory agent.

179. The therapeutic cell composition of any one of claims 173-178, wherein the non-specific or non-antigen-dependent stimulatory agent comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA).

180. 180. The therapeutic cell composition of any one of claims 177-179, wherein the non-specific or non-antigen-dependent T cell stimulator comprises PMA / ionomycin.

181. The therapeutic cell composition of any one of claims 173-176, wherein the stimulating agent specifically binds to the recombinant receptor, and optionally, the stimulating agent is an antigen-specific stimulating reagent and / or comprises an antigen or portion thereof that is specifically recognized by the recombinant receptor.

182. The therapeutic cell composition of any one of claims 173-181, wherein cytokine production is measured by an intracellular cytokine assay.

183. at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%, or at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, or at least or at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 40%, about 50%, about 60%, about 70%, about The therapeutic cell composition of any one of claims 173-182, comprising 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% of the cells that are surface positive for CCR7 and / or CD27, and optionally, are further negative for surface expression of CD45RA.

184. The therapeutic cell composition of any one of claims 173-183, wherein said T cells are primary cells obtained from a subject.

185. The therapeutic cell composition of claim 184, wherein the subject has cancer, and optionally, the cancer is leukemia or lymphoma.

186. The therapeutic cell composition of claim 185, wherein the subject is identified or known to have a high tumor burden at the time the cells are obtained.

187. The sum of two-way product (SPD) of the tumor in the subject is 30cm 2 Super, 40cm 2 Super, 50cm 2 Super, 60cm 2 Over or 70cm 2 Over or about 30cm 2 Super, about 40cm 2 Super, about 50cm 2 Super, about 60cm 2 Over or about 70cm 2 if greater than 100, the subject has a high tumor burden; and / or 187. The therapeutic cell composition of claim 186, wherein a subject has a high tumor burden when C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 milligrams / liter, greater than 10 milligrams / liter, greater than 15 milligrams / liter, greater than 20 milligrams / liter, greater than 25 milligrams / liter, greater than 30 milligrams / liter, greater than 40 milligrams / liter, or greater than 50 milligrams / liter, or greater than about 5 milligrams / liter, greater than about 10 milligrams / liter, greater than about 15 milligrams / liter, greater than about 20 milligrams / liter, greater than about 25 milligrams / liter, greater than about 30 milligrams / liter, greater than about 40 milligrams / liter, or greater than about 50 milligrams / liter.

188. The SPD of the tumor in the subject is 50 cm 2 Over or about 50cm 2 The therapeutic cell composition of claim 187, wherein if ≥ 100, the subject has a high tumor burden.

189. The therapeutic cell composition of claim 187 or 188, wherein the subject has a high tumor burden when CRP is greater than or greater than about 20 milligrams / liter in the biological sample.

190. The therapeutic cell composition may be enriched for CD4+ T cells and / or total recombinant receptors in the engineered composition. + At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least 97% of the cells, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD4 + 190. The therapeutic cell composition of any one of claims 173-189, wherein

191. The therapeutic cell composition is enriched for CD3+ T cells, enriched for CD4+ and CD8+ T cells, and / or total recombinant receptor in the engineered composition. + At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least 97%, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD3+ or CD4 + and CD8+.

192. The therapeutic cell composition of claim 191, comprising CD4+ and CD8+ T cells in a ratio of CD4+ T cells to CD8+ T cells that is about 1:3 to about 3:1, optionally about 1:2 to 2:

1.

193. 193. The therapeutic composition of any one of claims 173-192, wherein the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor.

194. The therapeutic composition of any one of claims 173-193, wherein the recombinant receptor is capable of binding to a target antigen associated with, specific for, and / or expressed in a cell or tissue of the disease, disorder, or condition.

195. 195. The therapeutic composition of claim 194, wherein the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer.

196. 196. The therapeutic composition of claim 194 or 195, wherein the target antigen is a tumor antigen.

197. Target antigens include αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, also known as CAIX or G250), cancer-testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, and CD12. 3, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutated (EGFRvIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OG D2), ganglioside GD3, glycoprotein 100 (gp100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPCR5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), quinine kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM),197. The therapeutic composition of any one of claims 194-196, wherein the antigen is selected from among oncofetal antigen, preferentially expressed antigen in melanoma (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), a pathogen-specific or pathogen-expressed antigen, or a universal tag-related antigen, and / or a biotinylated molecule, and / or a molecule expressed by HIV, HCV, HBV or other pathogens. ,

198. The recombinant receptor Functional non-TCR antigen receptors or TCRs or antigen-binding fragments thereof 198. The therapeutic composition of any one of claims 173-197, which is or comprises:

199. The therapeutic composition of any one of claims 173-198, wherein the recombinant receptor is a chimeric antigen receptor (CAR).

200. 200. The therapeutic composition of any one of claims 173-199, wherein the recombinant receptor comprises an extracellular domain comprising an antigen binding domain, and an intracellular signaling region comprising an intracellular signaling domain.

201. 201. The therapeutic composition of claim 200, wherein the antigen binding domain is or comprises an antibody or an antibody fragment thereof, optionally a single chain fragment.

202. 202. The therapeutic composition of claim 201, wherein said fragment comprises antibody variable regions linked by a flexible linker.

203. The therapeutic composition of claim 201 or 202, wherein the fragment comprises an scFv.

204. The intracellular signaling domain is a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain containing an immunoreceptor tyrosine-based activation motif (ITAM).

204. The therapeutic composition of any one of claims 200-203, which is or comprises:

205. The intracellular signaling domain is the intracellular signaling domain of the CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof 205. The therapeutic composition of any one of claims 200-204, which is or comprises:

206. The therapeutic composition of any one of claims 200-205, wherein the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region.

207. The therapeutic composition of any one of claims 200-206, wherein the intracellular signaling region further comprises a costimulatory signaling domain.

208. The therapeutic composition of claim 207, wherein said costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof.

209. The therapeutic composition of claim 207 or 208, wherein the costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof.

210. The therapeutic composition of any one of claims 207-209, wherein the costimulatory signaling domain is between the transmembrane domain and the intracellular signaling domain.

211. The therapeutic composition of any one of claims 173-210, wherein the T cells are primary T cells obtained from the subject.

212. The therapeutic composition of any one of claims 173-211, wherein the T cell is autologous to the subject.

213. 213. An article of manufacture comprising: a container containing the therapeutic cell composition of any one of claims 173-212; and instructions for administering the therapeutic cell composition, and optionally one or more unit doses thereof and / or an amount corresponding to such one or more unit doses, to a subject having a disease or condition.

214. 214. The article of manufacture of claim 213, wherein the instructions specify administering a plurality of unit doses contained in a plurality of separate compositions.

215. Multiple distinct compositions are identified as CD8 + T cells and CD4 + a first composition comprising a therapeutic cell composition comprising one of CD8 T cells; + T cells and CD4 + and a second composition comprising another of the T cells, wherein the second composition comprises a recombinant receptor or a cell expressing a recombinant receptor specific for an antigen or a cell expressed by a disease or condition, and optionally the second composition is the composition of any one of claims 1-40.

216. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, and CD4 + 216. The article of manufacture of claim 215, wherein the instructions specify that the T cells or one or more unit doses thereof and / or compositions containing an amount corresponding to such one or more unit doses are administered no more than 48 hours apart, no more than 36 hours apart, no more than 24 hours apart, no more than 12 hours apart, no more than 6 hours apart, no more than 2 hours apart, no more than 1 hour apart, or simultaneously.

217. CD8 + T cells or one or more unit doses thereof and / or a composition containing an amount corresponding to such one or more unit doses, + 217. The manufacture or article of manufacture of claim 215 or 216, wherein the instructions specify administration prior to administration of a composition containing the cells or one or more unit doses thereof and / or an amount corresponding to such one or more unit doses.

218. A method of treatment comprising administering to a subject having a disease or condition one or more unit doses of the therapeutic cell composition of any one of claims 173-212.

219. The method of claim 218, wherein said therapeutic composition is a first therapeutic cell composition comprising one of CD8+ T cells and CD4+ T cells, and wherein one or more unit doses further comprise a second therapeutic cell composition comprising the other of CD8+ T cells and CD4+ cells, said second composition comprising cells expressing a recombinant receptor or a recombinant receptor specific for an antigen or cells expressed by the disease or condition, and optionally wherein said second composition is the composition of any one of claims 173-212.

220. at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100%, or at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93% of the total number of T cells in the second therapeutic cell composition or of the total number of T cells expressing the recombinant receptor in said composition 48. The method of claim 47, wherein at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or at least about 100%, or 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the cells are surface positive for CCR7 and / or CD27, and optionally further negative for surface expression of CD45RA.

221. The second therapeutic cell composition may be enriched for CD8+ T cells and / or may contain total recombinant receptor agonists in the engineered composition. + At least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, or at least 97% of the cells, or about 60% to 99%, about 75% to 99%, about 85% to 99%, or about 90% to 99%, inclusive, of the total cells in the composition are CD8 + 221. The method of claim 219 or 220, wherein

222. 222. The method of claim 221, wherein the first therapeutic composition and the second therapeutic composition are administered within at or about 48 hours, within at or about 36 hours, within at or about 24 hours, within at or about 12 hours, within at or about 6 hours, within at or about 2 hours, or within at or about 1 hour of each other.

223. The method of claim 221 or 222, wherein the second therapeutic cell composition comprising CD8+ T cells is administered before the first composition comprising CD4+ T cells.

224. The method of any one of claims 218-223, wherein one or more unit doses of cells comprise a defined ratio of CD4+ T cells to CD8+ T cells, wherein the ratio is at or about 1:3 and 3:1 or about 3:1, optionally 1:2 or about 1:2 and 2:1 or about 2:1, optionally 1:1 or about 1:

1.

225. (A) assaying an engineered cell composition comprising T cells expressing a recombinant receptor for a factor indicative of the function or phenotype of the engineered cells in the composition, wherein the factor is selected from (i) the percentage of T cells expressing the recombinant receptor that are surface positive for CCR7 and / or CD27, and optionally surface negative for CD45RA; and / or (ii) the percentage of CD4+ T cells expressing the recombinant receptor that are capable of producing IL-2, TNF-alpha, and / or IFN-gamma after stimulation with a stimulatory agent; (B) administering a therapy to a subject having a disease or condition, (1) administering to the subject one or more unit doses of cells of an engineered cell composition comprising T cells expressing a recombinant receptor when an element indicative of the function or phenotype of cells of the engineered T cell composition is at or above a threshold value; or (2) if an element indicative of cellular function or phenotype of the engineered T cell composition is below a threshold value for that element; (a) one or more unit doses of cells of an engineered cell composition, and an agent capable of increasing the expansion, proliferation, or efficacy of T cells of the engineered cell composition in the subject; (b) optionally, an increased dose of cells of the engineered cell composition compared to one or more unit doses administered to a similarly situated subject having the same disease or condition but exhibiting at or above the threshold value of said factor; or (c) another therapeutic treatment for treating the disease or condition other than one or more unit doses of the engineered cell composition. administering a therapy selected from the administering step selected from the group consisting of Including, The threshold of the element is (i) at or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of T cells in the composition, or of the total number of T cells expressing a recombinant receptor in the composition, are surface positive for CCR7 and / or CD27; and / or (ii) at or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of CD4+ T cells expressing the recombinant receptor in the composition are capable of producing interleukin-2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or (iii) at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are polyfunctional to produce two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and TNF-alpha after stimulation with a stimulatory agent. A method for treating a subject selected from the group consisting of:

226. administering a therapy to a subject having a disease or condition, (1) administering to the subject one or more unit doses of cells of an engineered cell composition comprising T cells expressing a recombinant receptor when an element indicative of the function or phenotype of cells of the engineered cell composition is at or above a threshold value; or (2) if an element indicative of cellular function or phenotype of the engineered T cell composition is below a threshold value for that element; (a) one or more unit doses of cells of an engineered cell composition, and an agent capable of increasing the expansion, proliferation, or efficacy of T cells of the engineered cell composition in the subject; (b) optionally, an increased dose of cells of the engineered cell composition compared to one or more unit doses administered to a similarly situated subject having the same disease or condition but exhibiting at or above the threshold value of said factor; or (c) another therapeutic treatment for treating the disease or condition other than the administration of the engineered cell composition. administering a therapy selected from the administering step selected from Including, The threshold of the element is (i) at or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of T cells in the composition, or of the total number of T cells expressing a recombinant receptor in the composition, are surface positive for CCR7 and / or CD27; and / or (ii) at or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of CD4+ T cells expressing the recombinant receptor in the composition are capable of producing interleukin-2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or (iii) at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are polyfunctional to produce two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and TNF-alpha after stimulation with a stimulatory agent. A method for treating a subject selected from the group consisting of:

227. The method of any one of claims 218-226, wherein the cells of said cell composition are autologous to the subject.

228. The method of any one of claims 218-227, wherein at least 35%, at least 40%, at least 50%, at least 60%, or at least 70%, or at least 75% of subjects treated according to said method achieve progression free survival of at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 9 months, or at least 12 months after administration of the one or more unit doses.

229. The method of any of claims 218-228, wherein at least 35%, at least 40%, at least 50%, at least 60%, or at least 70%, or at least 75% of subjects treated according to said method are disease free for at least or at least about 3 months, at least 4 months, at least or at least about 4 months, at least 5 months, at least or at least about 5 months, at least 6 months, at least or at least about 6 months, at least 9 months, or at least about 9 months, or at least 12 months, after administration of the one or more unit doses.

230. A subject treated according to the method comprises: pharmacokinetic properties of the cells of the cell composition in a biological sample, optionally a blood or serum sample, that are improved on average compared to a group of similarly situated subjects having the same disease or condition and receiving a similar dose of an autologous cell composition comprising T cells expressing the recombinant receptor but with a factor indicative of cellular function or phenotype below a threshold. The method of any one of claims 218 to 229, wherein the method achieves the above.

231. 231. The method of claim 230, wherein said pharmacokinetic property is total exposure over time (AUC) or peak cell number in a biological sample.

232. (a) assaying an engineered cell composition comprising T cells expressing a recombinant receptor for a factor indicative of the function or phenotype of the engineered cells in the composition, wherein the factor is selected from (i) the percentage of T cells expressing the recombinant receptor that are surface positive for CCR7 and / or CD27, and optionally surface negative for CD45RA; and / or (ii) the percentage of CD4+ T cells expressing the recombinant receptor that are capable of producing IL-2, TNF-alpha, and / or IFN-gamma after stimulation with a stimulatory agent; (b) determining the likelihood of a response after administering a cell therapy comprising a dose of engineered cells, wherein if the factor is at or above a threshold, the subject is identified as likely to achieve a durable response or progression-free survival, optionally for at least three months, after administering the therapy; or if the factor is below a threshold, the subject is identified as unlikely to exhibit a durable response or progression-free survival to the therapy; Including, The threshold of the element is (i) at or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of T cells in the composition, or of the total number of T cells expressing a recombinant receptor in the composition, are surface positive for CCR7 and / or CD27; and / or (ii) at or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of CD4+ T cells expressing the recombinant receptor in the composition are capable of producing interleukin-2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or (iii) at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are polyfunctional to produce two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and TNF-alpha after stimulation with a stimulatory agent.

20. A method for predicting the likelihood of response to a therapeutic T cell composition selected from:

233. The method of claim 232, wherein the cells of said cell composition are autologous to the subject.

234. 234. The method of Claim 232 or 233, wherein if the subject is identified as likely to achieve a durable response or progression-free survival to the therapy, one or more unit doses of cells of the engineered T cell composition are administered to the subject.

235. One or more unit doses are 1 x 10 inclusive. 5 Or about 1 x 10 5 and 5×10 8 Or about 5 x 10 8 Between 1×10 5 Or about 1 x 10 5 and 1×10 8 Or about 1 x 10 8 Between 5x10 5 Or about 5 x 10 5 and 1×10 7 Or about 1 x 10 7 Between or 1×10 6 Or about 1 x 10 6 and 1×10 7 Or about 1 x 10 7 Between the total T cells expressing the recombinant receptor (receptor + 235. The method of any one of claims 218-231, 233, and 234, comprising a total number of T cells (i.e., a subset of T cells) or total T cells.

236. One or more unit doses may contain about 5 x 10 8 Below, approximately 1×10 8 Below, about 5×10 7 Below, approximately 1×10 7 Below, about 5×10 6 Below, approximately 1×10 6 or less, or about 5 x 10 5 Total receptors below + The method of any one of claims 218-231 and 233-235, comprising total T cells or total T cells.

237. 237. The method of any one of claims 225-236, wherein the element threshold is that at or about 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of T cells in said composition, or the total number of T cells expressing a recombinant receptor in said composition, are surface positive for CCR7 and / or CD27.

238. 238. The method of any one of claims 225-237, wherein the element threshold is that at or about 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the total number of CD4+ T cells expressing the recombinant receptor in the composition are capable of producing a cytokine selected from interleukin 2 (IL-2) and / or TNF-alpha.

239. The method of any one of claims 225-238, wherein the element threshold is that at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are multifunctional for producing two or more cytokines selected from among interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

240. The method of any one of claims 225-239, wherein the element threshold is that at or about 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or more of the total number of CD4+ T cells expressing the recombinant receptor in the composition are polyfunctional for producing the cytokines interferon-gamma (IFN-gamma), interleukin 2 (IL-2), and TNF-alpha.

241. If a subject is identified as unlikely to achieve a durable response or progression-free survival, (a) a dose of cells of an engineered cell composition, and an agent capable of increasing T cell expansion, proliferation, or efficacy of the engineered cell composition in the subject; (b) optionally, an increased dose of cells of the engineered cell composition compared to the dose administered to a similarly situated subject with the same disease or condition but who exhibits a greater likelihood of achieving progression-free survival or a durable response to cell therapy; or (c) another therapeutic treatment for treating the disease or condition other than the administration of the engineered cell composition. The method of any one of claims 232-240, wherein the subject is administered a therapy selected from the group consisting of:

242. The method of any one of claims 225-231 and 241, comprising administering a therapy comprising a dose of cells of the engineered cell composition and an agent capable of increasing the expansion, proliferation, or efficacy of T cells of said engineered cell composition in a subject, wherein the agent is an anti-idiotypic antibody or antigen-binding fragment thereof specific for a CAR, an immune checkpoint inhibitor, a modulator of a metabolic pathway, an adenosine receptor antagonist, a kinase inhibitor, an anti-TGFβ antibody, or an anti-TGFβR antibody, or a cytokine.

243. The method of claim 242, wherein said agent is administered before, simultaneously with, or after administration of a therapeutic T cell composition.

244. The method of any one of claims 218-242, wherein the subject is identified or known to have a high tumor burden at the time of the assay and / or prior to administration.

245. The sum of two-way product (SPD) of the tumor in the subject is 30cm 2 Super, 40cm 2 Super, 50cm 2 Super, 60cm 2 Over or 70cm 2 Over or about 30cm 2 Super, about 40cm 2 Super, about 50cm 2 Super, about 60cm 2 Over or about 70cm 2 if greater than 100, the subject has a high tumor burden; and / or a subject has a high tumor burden if C-reactive protein (CRP) in a biological sample, optionally a serum sample, from the subject is greater than 5 milligrams / liter, greater than 10 milligrams / liter, greater than 15 milligrams / liter, greater than 20 milligrams / liter, greater than 25 milligrams / liter, greater than 30 milligrams / liter, greater than 40 milligrams / liter, or greater than 50 milligrams / liter, or greater than about 5 milligrams / liter, greater than about 10 milligrams / liter, greater than about 15 milligrams / liter, greater than about 20 milligrams / liter, greater than about 25 milligrams / liter, greater than about 30 milligrams / liter, greater than about 40 milligrams / liter, or greater than about 50 milligrams / liter; The method of claim 244.

246. The SPD of the tumor in the subject is 50 cm 2 Over or about 50cm 2 The method of claim 245, wherein if β is greater than 1, then the subject has a high tumor burden.

247. The method of claim 245 or 246, wherein the subject has high tumor burden if CRP is greater than or greater than about 20 milligrams / liter in the biological sample.

248. The method of any one of claims 225-247, wherein the stimulating agent is a non-specific or non-antigen-dependent T cell stimulating agent.

249. The method of claim 248, wherein the non-specific or non-antigen-dependent T cell stimulatory agent is a polyclonal stimulatory agent.

250. The method of claim 248 or 249, wherein the non-specific or non-antigen-dependent stimulating agent comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA).

251. The method of any one of claims 248-250, wherein the non-specific or non-antigen-dependent T cell stimulatory agent comprises PMA / ionomycin.

252. The method of any one of claims 225-247, wherein the stimulating agent specifically binds to the recombinant receptor, and optionally the stimulating agent is an antigen-specific stimulating reagent and / or comprises an antigen or portion thereof that is specifically recognized by the recombinant receptor.

253. The method of any one of claims 225-252, wherein cytokine production is measured by an intracellular cytokine assay.

254. The method of any one of claims 218-253, wherein the recombinant receptor is or comprises a chimeric receptor and / or a recombinant antigen receptor.

255. The method of any one of claims 218-254, wherein the recombinant receptor is capable of binding to a target antigen associated with, specific for, and / or expressed in a cell or tissue of the disease, disorder, or condition.

256. The method of claim 255, wherein the disease, disorder, or condition is an infectious disease or disorder, an autoimmune disease, an inflammatory disease, or a tumor or cancer.

257. The method of claim 255 or 256, wherein the target antigen is a tumor antigen.

258. Target antigens include αvβ6 integrin (avb6 integrin), B-cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, also known as CAIX or G250), cancer-testis antigen, cancer / testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24, CD30, CD33, CD38, CD44, CD44v6, CD44v7 / 8, and CD12. 3, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), epidermal growth factor receptor type III mutated (EGFRvIII), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), folate-binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OG D2), ganglioside GD3, glycoprotein 100 (gp100), glypican-3 (GPC3), G protein-coupled receptor 5D (GPCR5D), Her2 / neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22Ra), IL-13 receptor alpha 2 (IL-13Rα2), quinine kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, leucine-rich repeat-containing 8 family member A (LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met, murine cytomegalovirus (CMV), mucin 1 (MUC1), MUC16, natural killer group 2 member D (NKG2D) ligand, melan-A (MART-1), neural cell adhesion molecule (NCAM),258. The method of any one of claims 255-257, wherein the antigen is selected from among oncofetal antigen, preferentially expressed antigen in melanoma (PRAME), progesterone receptor, prostate-specific antigen, prostate stem cell antigen (PSCA), prostate-specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor 2 (VEGFR2), Wilms' tumor 1 (WT-1), a pathogen-specific or pathogen-expressed antigen, or a universal tag-related antigen, and / or a biotinylated molecule, and / or a molecule expressed by HIV, HCV, HBV or other pathogens. ,

259. The recombinant receptor Functional non-TCR antigen receptors or TCRs or antigen-binding fragments thereof 259. The method of any one of claims 218-258, comprising or comprising:

260. The method of any one of claims 218-259, wherein the recombinant receptor is a chimeric antigen receptor (CAR).

261. The method of any one of claims 218-260, wherein the recombinant receptor comprises an extracellular domain comprising an antigen binding domain, and an intracellular signaling region comprising an intracellular signaling domain.

262. 262. The method of claim 261, wherein the antigen-binding domain is or comprises an antibody or an antibody fragment thereof, optionally a single chain fragment.

263. The method of claim 262, wherein said fragment comprises antibody variable regions linked by a flexible linker.

264. The method of claim 262 or 263, wherein the fragment comprises an scFv.

265. The intracellular signaling domain is a primary signaling domain, a signaling domain capable of inducing a primary activation signal in a T cell, a signaling domain of a T cell receptor (TCR) component, and / or a signaling domain containing an immunoreceptor tyrosine-based activation motif (ITAM).

265. The method of any one of claims 261-264, comprising or comprising:

266. The intracellular signaling domain is the intracellular signaling domain of the CD3 chain, optionally the CD3-zeta (CD3ζ) chain, or a signaling portion thereof 266. The method of claim 265, which is or comprises:

267. The method of any one of claims 261-266, wherein the recombinant receptor further comprises a transmembrane domain disposed between the extracellular domain and the intracellular signaling region.

268. The method of any one of claims 261-267, wherein said intracellular signaling region further comprises a costimulatory signaling domain.

269. The method of claim 268, wherein said costimulatory signaling domain comprises the intracellular signaling domain of a T cell costimulatory molecule, or a signaling portion thereof.

270. The method of claim 268 or 269, wherein said costimulatory signaling domain comprises the intracellular signaling domain of CD28, 4-1BB, or ICOS, or a signaling portion thereof.

271. The method of claim 269 or 270, wherein said costimulatory signaling domain is between said transmembrane domain and said intracellular signaling domain.

272. The method of any one of claims 218-271, wherein the T cell is a primary T cell obtained from the subject.

273. The method of any one of claims 218-272, wherein the T cell is autologous to the subject.

274. (a) incubating an input composition under stimulatory conditions, thereby producing a stimulated composition, wherein the input composition is capable of inducing a defined number of T cells that do not express apoptotic markers (apoptotic marker-negative ( - )) (b) introducing a recombinant receptor into cells from the stimulated composition, thereby producing an engineered cell composition, wherein the introducing comprises contacting cells of the stimulated composition with an agent comprising a polynucleotide encoding the recombinant receptor; A method for producing a therapeutic T cell composition, comprising:

275. Apoptosis marker-negative T cells Apoptosis marker-negative CD3 + Cells, apoptosis marker negative CD8 + cells, apoptosis marker-negative CD4+ cells, or apoptosis marker-negative CD8+ cells and apoptosis marker-negative CD4+ cells 275. The method of claim 274, which is or comprises:

276. The method of claim 274 or 275, wherein the apoptosis marker is annexin V.

277. The method of claim 274 or 275, wherein the apoptosis marker is activated caspase 3.

278. Defined numbers of T cells are at least 50 x 10 6 The method of any one of claims 274-276, comprising apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells.

279. Defined numbers of T cells are at least 100 x 10 6 The method of any one of claims 274-278, comprising apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells.

280. The prescribed number of T cells is 100 x 10 6 or about 100 x 10 6 and 500 x 10 6 or approximately 500 x 10 6 The method of any one of claims 274-279, comprising apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells, between

281. The prescribed number of T cells is 300 × 10 6 or approximately 300 x 10 6 The method of any one of claims 274-280, comprising apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells.

282. The method of any one of claims 274-281, wherein the percentage of apoptotic marker negative T cells as a percentage of total T cells in the input composition is greater than or about 70%, greater than or about 80%, greater than or about 90%, or greater than or about 95%.

283. The method of any one of claims 274-282, wherein prior to incubating, an input composition is obtained comprising a defined number of apoptotic marker-negative T cells.

284. The method of any one of claims 274-283, wherein prior to incubating, apoptosis marker-negative T cells, optionally apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells, are isolated, selected or enriched from the biological sample.

285. (a) T cells that do not express apoptosis markers (apoptosis marker negative ( - )), optionally isolating, selecting, or enriching a population of apoptosis marker-negative CD4+ and / or apoptosis marker-negative CD8+ T cells from the biological sample, thereby obtaining an input composition; (b) incubating the input composition under stimulated conditions, thereby producing a stimulated composition; (c) introducing a recombinant receptor into cells from the stimulated composition, thereby producing an engineered cell composition, wherein the introducing comprises contacting cells of the stimulated composition with an agent comprising a polynucleotide encoding the recombinant receptor; A method for producing a therapeutic T cell composition, comprising:

286. The method of claim 284 or 285, wherein the biological sample is primary T cells obtained from the subject.

287. The method of claim 286, wherein the subject is a human subject.

288. The biological sample is Whole blood sample, buffy coat sample, peripheral blood mononuclear cell (PBMC) sample, unfractionated T cell sample, lymphocyte sample, white blood cell sample, apheresis product, or leukapheresis product 288. The method of any one of claims 284-287, comprising or comprising:

289. The method of any one of claims 284-288, wherein the biological sample comprises an apheresis product or a leukapheresis product.

290. The method of any one of claims 284-289, wherein said T cells are isolated, selected or enriched from said biological sample within 36 hours of obtaining said biological sample from the subject.

291. The method of any one of claims 284-289, wherein the biological sample is cryogenically frozen in the presence of a cryoprotectant within 36 hours of being obtained from the subject and thawed prior to cell isolation, selection, or enrichment, optionally wherein the cryogenically frozen sample is thawed immediately prior to, or within 6 hours, 4 hours, 2 hours, or 1 hour prior to cell isolation, selection, or enrichment.

292. The method of any one of claims 274-291, wherein said incubation is carried out in the presence of one or more cytokines, optionally in serum-free medium.

293. 293. The method of claim 292, wherein the one or more cytokines are selected from recombinant IL-2, recombinant IL-7, and / or recombinant IL-15.

294. 294. The method of claim 293, wherein the one or more cytokines comprise 10 to 200 IU / mL recombinant IL-2; 100 IU / mL to 1,000 IU / mL recombinant IL-7; and / or 10 to 200 IU / mL recombinant IL-15.

295. The method of any one of claims 274-294, wherein the stimulatory reagent specifically binds to a member of the TCR complex, and optionally comprises a primary agent that specifically binds to CD3, and optionally the primary agent is an antibody or antigen-binding fragment thereof.

296. 296. The method of claim 295, wherein the stimulatory reagent further comprises a secondary agent that specifically binds to a T cell costimulatory molecule, optionally wherein the costimulatory molecule is selected from CD28, CD137 (4-1-BB), OX40, or ICOS, and optionally wherein the secondary agent is an antibody or antigen-binding fragment thereof.

297. The method of any one of claims 274-296, wherein the stimulating reagent comprises incubating with an anti-CD3 antibody and an anti-CD28 antibody, or antigen-binding fragment thereof.

298. The method of any one of claims 295-297, wherein the primary agent and / or the secondary agent are present on the surface of a solid support.

299. 299. The method of claim 298, wherein the solid support is or comprises a bead.

300. 300. The method of any one of claims 295-299, wherein the primary agent and the secondary agent are immobilized on or reversibly bound to the surface of an oligomeric particle reagent comprising a plurality of molecules of streptavidin or streptavidin mutein.

301. The method of any one of claims 274-300, wherein the input composition is incubated under stimulation conditions for 12 hours to 36 hours, inclusive, optionally 24 hours or about 24 hours.

302. The method of any one of claims 274-301, wherein said contacting is performed by viral transduction, optionally with a retroviral vector, optionally with a lentiviral vector.

303. The method of any one of claims 274-302, further comprising culturing the engineered composition under conditions that promote the growth and / or expansion of the engineered cells, thereby producing an output composition comprising the engineered T cells.

304. The method of claim 303, wherein said culturing is performed in the presence of one or more cytokines, optionally in serum-free medium.

305. 305. The method of claim 304, wherein the one or more cytokines are selected from recombinant IL-2, recombinant IL-7, and / or recombinant IL-15.

306. The method of claim 304 or 305, wherein the one or more cytokines comprise 50 to 400 IU / mL of recombinant IL-2; 100 IU / mL to 2,000 IU / mL of recombinant IL-7; and / or 50 to 400 IU / mL of recombinant IL-15.

307. the input composition comprises CD4+ T cells and CD8+ T cells in a ratio of CD4+ T cells to CD8+ T cells of 2:1 to 1:2; or the input composition comprises apoptosis marker-negative CD4+ T cells and apoptosis marker-negative CD8+ T cells at a ratio of CD4+ T cells to CD8+ T cells of 2:1 to 1:2; The method of any one of claims 274-306.

308. The method of claim 307, wherein the CD4+ T cells and CD8+ T cells are separately selected, isolated or enriched from the same biological sample or a sample derived therefrom and are combined prior to incubating.

309. The method of any one of claims 274-308, wherein the recombinant receptor is a chimeric antigen receptor (CAR).

310. The output composition is at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% of the total number of T cells in the composition or of the total number of T cells expressing a recombinant receptor in the composition are surface positive for CCR7 and / or CD27; and / or at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or at least 100% of the total number of CD4+ T cells in the composition expressing the recombinant receptor are capable of producing interleukin-2 (IL-2) or TNF-alpha after stimulation with a stimulatory agent; and / or At least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60% or more of the total number of CD4+ T cells in the composition expressing the recombinant receptor are multifunctional to produce two or more cytokines selected from interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and TNF-alpha after stimulation with a stimulatory agent. The method of any one of claims 303-309, comprising:

311. The method of claim 310, wherein the stimulating agent is a non-specific or non-antigen-dependent T cell stimulating agent.

312. The method of claim 311, wherein the non-specific or non-antigen-dependent T cell stimulatory agent is a polyclonal stimulatory agent.

313. The method of claim 311 or 312, wherein the non-specific or non-antigen-dependent stimulating agent comprises PMA / ionomycin, anti-CD3 / anti-CD28, phytohemagglutinin (PHA), or concanavalin A (ConA).

314. The method of any one of claims 311-313, wherein the non-specific or non-antigen-dependent T cell stimulatory agent comprises PMA / ionomycin.

315. 311. The method of claim 310, wherein the stimulating agent specifically binds to the recombinant receptor, and optionally, the stimulating agent is an antigen-specific stimulating reagent and / or comprises an antigen or portion thereof that is specifically recognized by the recombinant receptor.

316. The method of any one of claims 310-315, wherein cytokine production is measured by an intracellular cytokine assay.

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