Biomarkers and uses thereof for treatment of cancer with CD19-targeted adoptive cell therapies

Abstract

The presently disclosed subject matter provides methods for identifying subjects that are responsive or non-responsive to certain adoptive cell therapies or compositions comprising the same. In addition, methods for identifying subjects that have an increased likelihood or a reduced likelihood to response to certain adoptive cell therapies or compositions comprising the same are also provided. Moreover, the presently disclosed subject matter provides cells and compositions comprising a CD19-targeted chimeric receptor.

Description

[0001] BIOMARKERS AND USES THEREOF FOR TREATMENT OF CANCER WITH CD19- TARGETED ADOPTIVE CELL THERAPIES

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application claims priority to U.S. Provisional Application No. 63 / 595,170, filed November 1, 2023, the content of which is incorporated by reference in its entirety, and to which priority is claimed.

[0004] SEQUENCE LISTING

[0005] The instant application contains a Sequence Listing which has been submitted herewith and is hereby incorporated by reference in its entirety. Said .xml copy, created on October 24, 2024 is named 0871080164, and is 49,080 bytes in size.

[0006] TECHNICAL FIELD

[0007] The presently disclosed subject matter provides methods for identifying subjects that are responsive or non-responsive to certain adoptive cell therapies or compositions comprising the same. In addition, methods for identifying subjects that have an increased likelihood or a reduced likelihood to respond to certain adoptive cell therapies or compositions comprising the same are also provided. Moreover, the presently disclosed subject matter provides cells and compositions comprising a CD19-targeted chimeric receptor.

[0008] BACKGROUND

[0009] Cell -based immunotherapy is a therapy with curative potential for the treatment of cancer. T-cells and other immune cells may be modified to target tumor antigens through the introduction of genetic material coding for artificial or synthetic receptors for antigens, termed Chimeric Antigen Receptors (CARs), specific to selected antigens. Targeted T-cell therapy using CARs has shown recent clinical success in treating hematologic malignancies.

[0010] CD19-targeted chimeric antigen receptor (CAR) T cells have transformed the treatment of patients with relapsed or refractory CD19-positive hematologic malignancies (Park et al., TV Engl J Med 378, 449-459 (2018); Lee et al., Lancet 385, 517-528 (2015); Davila et al., Science translational medicine 6, 224ra225 (2014); and Maude et al., N Engl JMed3rT&, 439-448 (2018)). Four CAR T cell therapies have been approved by the Food and Drug Administration for the treatment of CD19-positive hematologic malignancies (Maude et al., N Engl J Med 378, 439-448 (2018); Neelapu et al., N Engl J Med 377 , 2531-2544 (2017); Schuster et al., N Engl J Med 380, 45-56 (2018); Wang et al., N Engl J Med 382, 1331-1342 (2020); Chong et al., N Engl J Med 384, 673-674 (2021); and Abramson et al., Lancet 396, 839-852 (2020)). Despite the promising results, most patients experience a lack of response or disease relapse, which can occur due to a lack of CAR T cell persistence, T cell exhaustion, or loss of the target antigen due to deletions spanning the CD19 locus, CAR masking or alternative splicing (Chong et al., N Engl J Med 384, 673-674 (2021); Sotillo et al., Cancer Discov 5, 1282-1295 (2015); Orlando et al., Nature medicine 24, 1504-1506 (2018); Ruella et al., Nature medicine 24, 1499-1503 (2018); Spiegel et al., Blood 137, 1832-1835 (2021)). Accordingly, there remains a need for identifying biomarkers to enable more targeted use of CD19-targeted chimeric receptors and cells comprising the same in cancer patients.

[0011] SUMMARY OF THE INVENTION

[0012] In certain non-limiting embodiments, the presently disclosed subject matter provides a method comprising a) measuring an expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; and c) identifying the subject as responsive or as having an increased likelihood to respond to an adoptive cell therapy. In certain embodiments, the subject is responsive or has an increased likelihood to respond to the adoptive cell therapy if a) the expression level of CD57 is increased compared to the reference sample; b) the expression level of TIM3 is increased compared to the reference sample; c) the expression level of KLRG1 is reduced compared to the reference sample; d) the expression level of CTLA4 is reduced compared to the reference sample; e) the expression level of LAG3 is reduced compared to the reference sample; and / or f) the expression level of PD1 is increased compared to the reference sample.

[0013] In certain non-limiting embodiments, the presently disclosed subject matter provides a method comprising a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; and c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy. In certain embodiments, the subject is non- responsive or has a reduced likelihood to respond the adoptive cell therapy if a) the expression level of CD57 is reduced compared to the reference sample; b) the expression level of TIM3 is reduced compared to the reference sample; c) the expression level of KLRG1 is increased compared to the reference sample; d) the expression level of CTLA4 is increased compared to the reference sample; e) the expression level of LAG3 is increased compared to the reference sample; and / or f) the expression level of PD1 is reduced compared to the reference sample.

[0014] In certain embodiments, the adoptive cell therapy comprises a cell comprising a CD 19- targeted chimeric receptor.

[0015] In certain embodiments, the methods disclosed herein further comprise administering to the subject a cell comprising a CD19-targeted chimeric receptor or a composition comprising thereof. In certain embodiments, the CD19-targeted chimeric receptor comprises an extracellular antigen-binding domain comprising a heavy chain variable region (VH) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12, and a light chain variable region (VL) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15.

[0016] In certain embodiments, the extracellular antigen-binding domain comprises a scFv. In certain embodiments, the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 16; and the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 17. In certain embodiments, the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 16; and the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 17.

[0017] In certain embodiments, the chimeric receptor comprises a transmembrane domain comprising a CD28 polypeptide. In certain embodiments, the chimeric receptor comprises intracellular signaling domain comprises a CD3(^ polypeptide comprising a native ITAM1, an ITAM2 variant consisting of two loss-of-function mutations, and an ITAM3 consisting of two loss-of-function mutations. In certain embodiments, the native ITAM1 consists of the amino acid sequence set forth in SEQ ID NO: 28, the ITAM2 variant consists of the amino acid sequence set forth in SEQ ID NO: 34, and the ITAM3 variant consists of the amino acid sequence set forth in SEQ ID NO: 38. In certain embodiments, the chimeric receptor comprises an intracellular signaling domain comprising a CD3(^ polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 40. In certain embodiments, the intracellular signaling domain further comprises a CD28 polypeptide. In certain embodiments, the chimeric receptor comprises the amino acid sequence set forth in SEQ ID NO: 45.

[0018] In certain embodiments, the chimeric receptor is expressed from a vector. In certain embodiments, the vector is a retroviral vector. In certain embodiments, the chimeric receptor is constitutively expressed on the surface of the cell.

[0019] In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is an immunoresponsive cell. In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is a cell of the lymphoid lineage or a cell of the myeloid lineage. In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a stem cell from which lymphoid cells may be differentiated, and a stem cell from which myeloid cells may be differentiated. In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is a T cell. In certain embodiments, the T cell is selected from the group consisting of helper T cells, cytotoxic T cells, memory T cells, regulatory T cells, tumor-infiltrating lymphocyte (TIL), Natural Killer T cells, mucosal associated invariant T cells, and y5 T cells. In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is a NK cell. In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is derived from a stem cell. In certain embodiments, the stem cell is a pluripotent stem cell. In certain embodiments, the pluripotent stem cell is an embryoid stem cell or an induced pluripotent stem cell.

[0020] In certain embodiments, the cell comprising the CD19-targeted chimeric receptor is selected from a CD57+T cell, a TIM3+T cell, a KLRGT T cell, a CTLA4' T cell, a LAG3' T cell, a PD1+T cell, a CD57+TIM3+T cell, a CD57+KLRGT T cell, a CD57+CTLA4' T cell, a CD57+LAG3’ T cell, a CD57+PD1+T cell, a TIM3+KLRGT T cell, a TIM3+CTLA4' T cell, a TIM3+LAG3’ T cell, a TIM3+PD1+T cell, a KLRGT CTLA4' T cell, a KLRGT LAG3' T cell, a KLRGT PD1+T cell, a CTLA4' LAG3' T cell, a CTLA4' PD1+T cell, a LAG3' PD1+T cell, a CD57+TIM3+KLRGT T cell, a CD57+TIM3+CTLA4' T cell, a CD57+TIM3+LAG3' T cell, a CD57+TIM3+PD1+T cell, a CD57+KLRGT CTLA4' T cell, a CD57+KLRGT LAG3' T cell, a CD57+KLRGT PD1+T cell, a CD57+CTLA4' LAG3' T cell, a CD57+CTLA4' PD1+T cell, a CD57+LAG3’ PD1+T cell, a TIM3+KLRGT CTLA4' T cell, a TIM3+KLRGT LAG3' T cell, a TIM3+CTLA4’ LAG3’ T cell, a TIM3+KLRGT PD1+T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+CTLA4’ PD1+T cell, a TIM3+LAG3' PD1+T cell, a KLRGT CTLA4' LAG3' T cell, a KLRGT CTLA4’ PD1+T cell, a KLRGT LAG3' PD1+T cell, a CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' T cell, a CD57+TIM3+KLRGT LAG3' T cell, a CD57+TIM3+KLRGT PD1+T cell, a CD57+TIM3+CTLA4' LAG3' T cell, a CD57+TIM3+CTLA4' PD1+T cell, a CD57+TIM3+LAG3’ PD1+T cell, a CD57+KLRGT CTLA4' LAG3' T cell, a CD57+KLRGT CTLA4’ PD1+T cell, a CD57+KLRGT LAG3' PD1+T cell, a CD57+CTLA4' LAG3' PD1+T cell, a TIM3+KLRGT CTLA4' LAG3' T cell, a TIM3+KLRGT CTLA4' LAG3' T cell, a TIM3+KLRGT CTLA4’ PD1+T cell, a TIM3+KLRGT LAG3' PD1+T cell, a TIM3+CTLA4' LAG3' PD1+T cell, a KLRGT CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' LAG3' T cell, a CD57+TIM3+KLRGT CTLA4' PD1+T cell, a CD57+TIM3+KLRGT LAG3' PD1+T cell, a CD57+TIM3+CTLA4' LAG3' PD1+T cell, a CD57+KLRGT CTLA4' LAG3' PD1+T cell, a TIM3+KLRGT CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' LAG3' PD1+T cell, or a combination thereof.

[0021] In certain embodiments, the composition comprises at least about 5% of a CD57+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a TIM3+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' PD1+ T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGr CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGL CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, or a combination thereof.

[0022] In certain embodiments, the subject has a tumor and / or a neoplasm associated with CD 19. In certain embodiments, the tumor and / or neoplasm is a blood cancer. In certain embodiments, the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas. In certain embodiments, the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed-phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia. In certain embodiments, the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma. In certain embodiments, the tumor and / or neoplasm is a B cell malignancy. In certain embodiments, the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma. In certain embodiments, the tumor and / or neoplasm is B cell lymphoma. In certain embodiments, the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma.

[0023] In certain embodiments, the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 is measured on the surface of the immunoresponsive cell. In certain embodiments, the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 is measured by cytometry or by flow-cytometry. In certain embodiments, the immunoresponsive cell is a T cell. In certain embodiments, the T cell is selected from the group consisting of a helper T cell, a cytotoxic T cell, a memory T cell, a central memory T cell, a stemlike memory T cell, an effector memory T cell, a regulatory T cell, a tumor-infiltrating lymphocyte (TIL), a Natural killer T cells, a y5 T cell, a CD4+T cells, and a CD8+T cell. In certain embodiments, the T cell is a CD4+T cell or a CD8+T cell. In certain embodiments, the immunoresponsive cell is obtained from a sample of the subject. In certain embodiments, the sample is selected from bone marrow, thymus, tumor biopsy, tumor fragments, enzymatically digested tumor tissue, dissociated / suspended cells, a lymph node sample, or a bodily fluid sample. In certain embodiments, the bodily fluid sample is a blood sample, an ascites sample, or a lymph sample. In certain embodiments, the subject is a human subject.

[0024] In certain non-limiting embodiments, the presently disclosed subject matter provides a composition comprising a cell comprising a CD19-targeted chimeric receptor, wherein the CD 19- targeted chimeric receptor comprises an extracellular antigen-binding domain, a transmembrane domain, and an intracellular domain, wherein the extracellular antigen-binding domain comprises a heavy chain variable region (VH) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12, and a light chain variable region (VL) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15.

[0025] In certain embodiments, the extracellular antigen-binding domain comprises a scFv. In certain embodiments, the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 16; and the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 17. In certain embodiments, the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 16; and the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 17.

[0026] In certain embodiments, the transmembrane domain comprises a CD28 polypeptide. In certain embodiments, the intracellular signaling domain comprises a CD3(^ polypeptide comprising a native IT AMI, an ITAM2 variant consisting of two loss-of-function mutations, and an ITAM3 consisting of two loss-of-function mutations. In certain embodiments, the native ITAM1 consists of the amino acid sequence set forth in SEQ ID NO: 28, the ITAM2 variant consists of the amino acid sequence set forth in SEQ ID NO: 34, and the ITAM3 variant consists of the amino acid sequence set forth in SEQ ID NO: 38. In certain embodiments, the intracellular signaling domain comprises a CD3(^ polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 40. In certain embodiments, the intracellular signaling domain further comprises a CD28 polypeptide. In certain embodiments, the chimeric receptor comprises the amino acid sequence set forth in SEQ ID NO: 45.

[0027] In certain embodiments, the chimeric receptor is expressed from a vector. In certain embodiments, the vector is a retroviral vector. In certain embodiments, the chimeric receptor is constitutively expressed on the surface of the cell. In certain embodiments, the cell is an immunoresponsive cell. In certain embodiments, the cell is a cell of the lymphoid lineage or a cell of the myeloid lineage. In certain embodiments, the cell is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a stem cell from which lymphoid cells may be differentiated, and a stem cell from which myeloid cells may be differentiated. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is selected from the group consisting of helper T cells, cytotoxic T cells, memory T cells, regulatory T cells, tumor-infiltrating lymphocyte (TIL), Natural Killer T cells, mucosal associated invariant T cells, and y5 T cells. In certain embodiments, the cell is an NK cell. In certain embodiments, the cell is derived from a stem cell. In certain embodiments, the stem cell is a pluripotent stem cell. In certain embodiments, the pluripotent stem cell is an embryoid stem cell or an induced pluripotent stem cell.

[0028] In certain embodiments, the the cell is selected from a CD57+T cell, a TIM3+T cell, a KLRG1' T cell, a CTLA4' T cell, a LAG3' T cell, a PD1+T cell, a CD57+TIM3+T cell, a CD57+ KLRGL T cell, a CD57+CTLA4' T cell, a CD57+LAG3' T cell, a CD57+PD1+T cell, a TIM3+KLRGL T cell, a TIM3+CTLA4' T cell, a TIM3+LAG3' T cell, a TIM3+PD1+T cell, a KLRGL CTLA4’ T cell, a KLRGL LAG3' T cell, a KLRGr PD1+T cell, a CTLA4' LAG3' T cell, a CTLA4’ PD1+T cell, a LAG3' PD1+T cell, a CD57+TIM3+KLRGL T cell, a CD57+TIM3+CTLA4’ T cell, a CD57+TIM3+LAG3' T cell, a CD57+TIM3+PD1+T cell, a CD57+KLRGL CTLA4’ T cell, a CD57+KLRGL LAG3' T cell, a CD57+KLRGL PD1+T cell, a CD57+CTLA4' LAG3’ T cell, a CD57+CTLA4' PD1+T cell, a CD57+LAG3' PD1+T cell, a TIM3+KLRGL CTLA4’ T cell, a TIM3+KLRGL LAG3' T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+KLRGL PD1+T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+CTLA4' PD1+T cell, a TIM3+LAG3' PD1+T cell, a KLRGL CTLA4' LAG3' T cell, a KLRGL CTLA4' PD1+T cell, a KLRGL LAG3' PD1+T cell, a CTLA4’ LAG3' PD1+T cell, a CD57+TIM3+KLRGL CTLA4' T cell, a CD57+TIM3+KLRGL LAG3’ T cell, a CD57+TIM3+KLRGL PD1+T cell, a CD57+TIM3+CTLA4' LAG3' T cell, a CD57+TIM3+CTLA4' PD1+T cell, a CD57+TIM3+LAG3' PD1+T cell, a CD57+KLRGL CTLA4’ LAG3’ T cell, a CD57+KLRGL CTLA4' PD1+T cell, a CD57+KLRGL LAG3' PD1+T cell, a CD57+CTLA4' LAG3' PD1+T cell, a TIM3+KLRGL CTLA4' LAG3' T cell, a TIM3+KLRGL CTLA4’ LAG3' T cell, a TIM3+KLRGL CTLA4' PD1+T cell, a TIM3+KLRGL LAG3' PD1+T cell, a TIM3+CTLA4' LAG3' PD1+T cell, a KLRGL CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGL CTLA4' LAG3' T cell, a CD57+TIM3+KLRGL CTLA4' PD1+T cell, a CD57+TIM3+KLRGL LAG3' PD1+T cell, a CD57+TIM3+CTLA4' LAG3' PD1+T cell, a CD57+KLRGL CTLA4' LAG3' PD1+T cell, a TIM3+KLRGL CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGL CTLA4' LAG3' PD1+T cell, or a combination thereof.

[0029] In certain embodiments, the composition comprises at least about 5% of a CD57+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGL T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRG1" CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRG1" LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRG1" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG I ' CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGI" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGI" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGI" CTLA4" T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a TIM3+KLRGI" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGI" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGI" CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGI" CTLA4" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP LAG3" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4" LAG3" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGI" CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGI" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGI" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+ LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG1" CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRG1" CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRG1" CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG1" CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRG1" CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, or a combination thereof.

[0030] In certain embodiments, the composition is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier. In certain embodiments, the composition comprises between about 1 x 106and about 5 * 108cells. In certain embodiments, the composition comprises between about 1 x 106and about 1 x 108cells. In certain embodiments, the composition comprises about 1 x io6and about 5 x io7cells. In certain embodiments, the composition comprises about 2.5 x io7cells.

[0031] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of reducing tumor burden in a subject, comprising administering to the subject a composition disclosed herein. In certain embodiments, the method reduces the number of tumor cells, reduces tumor size, and / or eradicates the tumor in the subject.

[0032] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of increasing or lengthening survival of a subject having a neoplasm, comprising administering to the subject a composition disclosed herein. In certain non-limiting embodiments, the presently disclosed subject matter provides a method of treating and / or preventing a neoplasm in a subject, comprising administering to the subject a composition of disclosed herein.

[0033] In certain embodiments, the tumor and / or neoplasm is associated with CD 19. In certain embodiments, the tumor and / or neoplasm is a blood cancer. In certain embodiments, the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas. In certain embodiments, the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed-phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia. In certain embodiments, the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma. In certain embodiments, the tumor and / or neoplasm is a B cell malignancy. In certain embodiments, the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma. In certain embodiments, the tumor and / or neoplasm is B cell lymphoma. In certain embodiments, the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma. In certain embodiments, the subject is a human subject.

[0034] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of reducing tumor burden in a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy if i) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample, nd / or vi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition disclosed herein.

[0035] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of increasing or lengthening survival of a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy if i) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample, and / or vi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition disclosed herein.

[0036] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of treating and / or preventing a neoplasm in a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy if i) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample, and / or vi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition disclosed herein.

[0037] In certain embodiments, the tumor and / or neoplasm is associated with CD 19. In certain embodiments, the tumor and / or neoplasm is a blood cancer. In certain embodiments, the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas. In certain embodiments, the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed-phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia. In certain embodiments, the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma. In certain embodiments, the tumor and / or neoplasm is a B cell malignancy. In certain embodiments, the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma. In certain embodiments, the tumor and / or neoplasm is B cell lymphoma. In certain embodiments, the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma. In certain embodiments, the subject is a human subject.

[0038] Finally, in certain non-limiting embodiments, the presently disclosed subject matter provides a kit for performing any of the methods disclosed herein.

[0039] BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figures 1A and IB illustrate positive association between response and CD57 and TIM3 expression independently on CAR+Cells and CAR' Cells as well as negative association between response and CTLA4, KLRG1, LAG3 expression independently in the administered compositions / final infusion product. CR: complete response; PR: partial response; NR: noresponse.

[0041] Figure 2 illustrates that untransduced cells (UTD) show a similar trend as the final infusion product for these markers. CR: complete response; PR: partial response; NR: no-response.

[0042] Figure 3 illustrates associations between response and CD57, CTLA4, and PD1 are not seen in starting / apheresis material (e.g., a sample from a subject), potential association with KLRG1, LAG3 and TIM3.

[0043] DETAILED DESCRIPTION

[0044] The presently disclosed subject matter is based, at least in part, on the discovery that expression levels of certain cell surface markers (e.g., CD57, KLRG1, CTLA4, LAG3, TIM3 and PD1) in CD8+and / or CD4+T cells can predict the responsiveness of patients treated with adoptive cell therapies including an anti-CD19 chimeric antigen receptor (e.g., CD19(T2)28zlxx).

[0045] In certain non-limiting embodiments, the presently disclosed subject matter provides a method of identifying a subject having cancer that is responsive to the adoptive cell therapy. In certain embodiments, the method comprises measuring the expression level of CD57, KLRG1, CTLA4, LAG3, TIM3, and / or PD1 in CD8+and / or CD4+T cells from the subject. In certain embodiments, the subject is responsive to the adoptive cell therapy if the expression level of CTLA4, LAG3, and / or KLRG1 in the CD8+and / or CD4+T cells from the subject is reduced as compared to a reference sample. In certain embodiments, the subject is responsive to the adoptive cell therapy if the expression level of CD57, TIM3, and / or PD1 in the CD8+and / or CD4+T cells from the subject is increased as compared to a reference sample. Additionally, in certain non-limiting embodiments, the presently disclosed subject matter provides an immunoresponsive cell (e.g., a T cell) that has increased therapeutic efficacy. In certain embodiments, the immunoresponsive cell comprises a reduced expression level of CTLA4, LAG3, and / or KLRG1 as compared to a reference sample. In certain embodiments, the immunoresponsive cell comprises an increased expression level of CD57, TIM3, and / or PD1 as compared to a reference sample. Without being bound by any theory, it is believed that CD8+and / or CD4+T cells that express reduced levels of CTLA4, LAG3, and / or KLRG1 and / or increased levels of CD57, TIM3, and / or PD1 have increased functional activity (e.g., cytotoxicity, proliferation, cytokine secretion) in the context of adoptive cell therapies.

[0046] Non-limiting embodiments of the present disclosure are described by the present specification and Examples.

[0047] For purposes of clarity of disclosure and not by way of limitation, the detailed description is divided into the following subsections:

[0048] 1. Definitions;

[0049] 2. Cells;

[0050] 3. Nucleic Acid Molecules and Vectors;

[0051] 4. Formulations and Administration;

[0052] 5. Biomarkers;

[0053] 6. Methods of Treatment;

[0054] 7. Kits; and

[0055] 8. Exemplary Embodiments.

[0056] / . Definitions

[0057] Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by a person skilled in the art to which the presently disclosed subject matter belongs. The following references provide one of skill with a general definition of many of the terms used in the presently disclosed subject matter: Singleton et al., Dictionary of Microbiology and Molecular Biology (2nded. 1994); The Cambridge Dictionary of Science and Technology (Walker ed., 1988); The Glossary of Genetics, 5thEd., R. Rieger et al. (eds.), Springer Verlag (1991); and Hale & Marham, The Harper Collins Dictionary of Biology (1991). As used herein, the following terms have the meanings ascribed to them below, unless specified otherwise.

[0058] As used herein, the term “about” or “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, z.e., the limitations of the measurement system. For example, “about” can mean within 3 or more than 3 standard deviations, per the practice in the art. Alternatively, “about” can mean a range of up to 20%, preferably up to 10%, more preferably up to 5%, and more preferably still up to 1% of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold, of a value.

[0059] By “immunoresponsive cell” is meant a cell that functions in an immune response or a progenitor, or progeny thereof. In certain embodiments, the immunoresponsive cell is a cell of lymphoid lineage. Non-limiting examples of cells of lymphoid lineage include T cells, Natural Killer (NK) cells, B cells, and stem cells from which lymphoid cells may be differentiated. In certain embodiments, the immunoresponsive cell is a cell of myeloid lineage.

[0060] By “activates an immunoresponsive cell” is meant induction of signal transduction or changes in protein expression in the cell resulting in initiation of an immune response. For example, when CD3 Chains cluster in response to ligand binding and immunoreceptor tyrosinebased inhibition motifs (ITAMs) a signal transduction cascade is produced. In certain embodiments, when an endogenous TCR or an exogenous CAR binds to an antigen, a formation of an immunological synapse occurs that includes clustering of many molecules near the bound receptor (e.g. CD4 or CD8, CD3v / 6 / s / (^, etc.). This clustering of membrane bound signaling molecules allows for ITAM motifs contained within the CD3 chains to become phosphorylated. This phosphorylation in turn initiates a T cell activation pathway ultimately activating transcription factors, such as NF-KB and AP-1. These transcription factors induce global gene expression of the T cell to increase IL-2 production for proliferation and expression of master regulator T cell proteins in order to initiate a T cell mediated immune response.

[0061] By “stimulates an immunoresponsive cell” is meant a signal that results in a robust and sustained immune response. In various embodiments, this occurs after immune cell (e.g., T-cell) activation or concomitantly mediated through receptors including, but not limited to, CD28, CD137 (4-1BB), 0X40, CD40 and ICOS. Receiving multiple stimulatory signals can be important to mount a robust and long-term T cell mediated immune response. T cells can quickly become inhibited and unresponsive to antigen. While the effects of these co-stimulatory signals may vary, they generally result in increased gene expression in order to generate long lived, proliferative, and anti-apoptotic T cells that robustly respond to antigen for complete and sustained eradication.

[0062] As used herein, the term “antibody” means not only intact antibody molecules, but also fragments of antibody molecules that retain immunogen-binding ability. Such fragments are also well known in the art and are regularly employed both in vitro and in vivo. Accordingly, as used herein, the term “antibody” means not only intact immunoglobulin molecules but also the well- known active fragments F(ab')2, and Fab. F(ab')2, and Fab fragments that lack the Fe fragment of intact antibody, clear more rapidly from the circulation, and may have less non-specific tissue binding of an intact antibody (Wahl et al., NuclMed (1983);24:316-325). As used herein, include whole native antibodies, bispecific antibodies; chimeric antibodies; Fab, Fab’, single chain V region fragments (scFv), fusion polypeptides, and unconventional antibodies. In certain embodiments, an antibody is a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant (CH) region. The heavy chain constant region is comprised of three domains, CHI, CH2 and CH3. Each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant CL region. The light chain constant region is comprised of one domain, CL. The VH and VL regions can be further sub-divided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Clq) of the classical complement system.

[0063] As used herein, “CDRs” are defined as the complementarity determining region amino acid sequences of an antibody which are the hypervariable regions of immunoglobulin heavy and light chains. See, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 4th U. S. Department of Health and Human Services, National Institutes of Health (1987), or IMGT numbering system (Lefranc, The Immunologist (1999);7: 132-136; Lefranc et al., Dev. Comp. Immunol. (2003);27:55-77). Generally, antibodies comprise three heavy chain and three light chain CDRs or CDR regions in the variable region. CDRs provide the majority of contact residues for the binding of the antibody to the antigen or epitope. In certain embodiments, the CDRs regions are delineated using the IMGT numbering system. In certain embodiments, the CDR regions are delineated using the IMGT numbering system accessible at http : / / www.imgt. org / IMGT_vquest / input.

[0064] As used herein, the term “single-chain variable fragment” or “scFv” is a fusion protein of the variable regions of the heavy (VH) and light chains (VL) of an immunoglobulin (e.g., mouse or human) covalently linked to form a VH: :VL heterodimer. The heavy (VH) and light chains (VL) are either joined directly or joined by a peptide-encoding linker (e.g., 10, 15, 20, 25 amino acids), which connects the N-terminus of the VH with the Cterminus of the VL, or the C-terminus of the VH with the N-terminus of the VL. The linker is usually rich in glycine for flexibility, as well as serine or threonine for solubility. The linker can link the heavy chain variable region and the light chain variable region of the extracellular antigen-binding domain. Non-limiting examples of linkers are disclosed in Shen et al., Anal. Chem. 80(6): 1910-1917 (2008) and WO 2014 / 087010, the contents of which are hereby incorporated by reference in their entireties. In certain embodiments, the linker is a G4S linker.

[0065] In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 1, which is provided below:

[0066] GGGGSGGGGSGGGSGGGGS [ SEQ ID NO : 1 ]

[0067] In certain embodiments, the linker comprise or consists of the amino acid sequence set forth in SEQ ID NO: 2, which is provided below:

[0068] GGGGSGGGGSGGGGS [ SEQ ID NO : 2 ]

[0069] In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 3, which is provided below:

[0070] GGGGSGGGGSGGGGSGGGSGGGGS [ SEQ ID NO : 3 ]

[0071] In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 4, which is provided below:

[0072] GGGGSGGGGSGGGGSGGGGSGGGSGGGGS [ SEQ ID NO : 4 ]

[0073] In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 5, which is provided below:

[0074] GGGGS [ SEQ ID NO : 5 ]

[0075] In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 6, which is provided below:

[0076] GGGGSGGGGS [ SEQ ID NO : 6 ]

[0077] In certain embodiments, the linker comprises the first three amino acids of the heavy chain constant region. In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 7, which is provided below:

[0078] ASTGGGGSGGGGSGGGGS [ SEQ ID NO : 7 ]

[0079] Additionally or alternately, the linker can be a Whitlow / 218 linker disclosed in Whitlow, M. et al. (1993) Protein Eng 6, 989-95, the contents of which are hereby incorporated by reference in their entireties.

[0080] Despite removal of the constant regions and the introduction of a linker, scFv proteins retain the specificity of the original immunoglobulin. Single chain Fv polypeptide antibodies can be expressed from a nucleic acid comprising VH - and VL encoding sequences as described by Huston, et al. Proc. Nat. Acad. Sci. USA, (1988);85:5879-5883; U.S. Patent Nos. 5,091,513, 5,132,405 and 4,956,778; and U.S. Patent Publication Nos. 20050196754 and 20050196754. Antagonistic scFvs having inhibitory activity have been described (see, e.g., Zhao et al., Hybridoma (Larchmt) (2008);27(6):445-51; Peter et al., J Cachexia Sarcopenia Muscle (2012);August 12; Shieh et al., J Imunol (2009);183(4):2277-85; Giomarelli et al., Thromb Haemost (2007);97(6):955-63; Fife eta., J Clin Invst (2006);l 16(8):2252-61; Brocks et al., Immunotechnology 1997 3(3): 173-84; Moosmayer et al., Ther Immunol 1995 2(10:31-40). Agonistic scFvs having stimulatory activity have been described (Peter et al., J Biol Chem (2003);25278(38):36740-7; Xie et al., Nat Biotech 1997 15(8):768-71; Ledbetter et al., Crit Rev Immunol (1997);17(5-6):427-55; Ho et al., BioChim Biophys Acta (2003);1638(3):257-66).

[0081] The term “chimeric antigen receptor” or “CAR” as used herein refers to a molecule comprising an extracellular antigen-binding domain that is fused to an intracellular signaling domain that is capable of activating or stimulating an immunoresponsive cell. In certain embodiments, the CAR also comprises a transmembrane domain. In certain embodiments, the extracellular antigen-binding domain of a CAR comprises an scFv. The scFv can be derived from fusing the variable heavy and light regions of an antibody. Alternatively or additionally, the scFv may be derived from Fab’s (instead of from an antibody, e.g., obtained from Fab libraries). In certain embodiments, the scFv is fused to the transmembrane domain and then to the intracellular signaling domain.

[0082] The term “chimeric co-stimulating receptor” or “CCR” refers to a chimeric receptor that binds to an antigen and provides co-stimulatory signals, but does not alone provide an activation signal. CCR is described in Krause, et al., J. Exp. Med. (1998);188(4):619-626, and US20020018783, the contents of which are incorporated by reference in their entirety. CCRs mimic co-stimulatory signals, but unlike, CARs, do not alone provide an activation signal, e.g., CCRs lack a CD3^ polypeptide.

[0083] By “substantially identical” or “substantially homologous” is meant a polypeptide or nucleic acid molecule exhibiting at least about 50% homologous or identical to a reference amino acid sequence (for example, any of the amino acid sequences described herein) or a reference nucleic acid sequence (for example, any of the nucleic acid sequences described herein). In certain embodiments, such a sequence is at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, or at least about 100% homologous or identical to the sequence of the amino acid or nucleic acid used for comparison.

[0084] Sequence identity can be measured by using sequence analysis software (for example, Sequence Analysis Software Package of the Genetics Computer Group, University of Wisconsin Biotechnology Center, 1710 University Avenue, Madison, Wis. 53705, BLAST, BESTFIT, GAP, or PILEUP / PRETTYBOX programs). Such software matches identical or similar sequences by assigning degrees of homology to various substitutions, deletions, and / or other modifications. Conservative substitutions typically include substitutions within the following groups: glycine, alanine; valine, isoleucine, leucine; aspartic acid, glutamic acid, asparagine, glutamine; serine, threonine; lysine, arginine; and phenylalanine, tyrosine. In an exemplary approach to determining the degree of identity, a BLAST program may be used, with a probability score between e-3 and e-100 indicating a closely related sequence.

[0085] As used herein, the percent homology between two amino acid sequences is equivalent to the percent identity between the two sequences. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences (z.e., % homology = # of identical positions / total # of positions x 100), taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm.

[0086] The percent homology between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. AppL Biosci.. 4: 11-17 (1988)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In addition, the percent homology between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol. 48:444-453 (1970)) algorithm which has been incorporated into the GAP program in the GCG software package (available at www.gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6.

[0087] Additionally or alternatively, the amino acids sequences of the presently disclosed subject matter can further be used as a “query sequence” to perform a search against public databases to, for example, identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10. BLAST protein searches can be performed with the XBLAST program, score = 50, wordlength = 3 to obtain amino acid sequences homologous to the specified sequences (e.g., heavy and light chain variable region sequences of scFv m903, m904, m905, m906, and m900) disclosed herein. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. An “effective amount” is an amount sufficient to affect a beneficial or desired clinical result upon treatment. An effective amount can be administered to a subject in one or more doses. In certain embodiments, an effective amount can be an amount that is sufficient to palliate, ameliorate, stabilize, reverse or slow the progression of the disease, or otherwise reduce the pathological consequences of the disease. The effective amount can be determined by a physician on a case-by-case basis and is within the skill of one in the art. Several factors are typically taken into account when determining an appropriate dosage to achieve an effective amount. These factors include age, sex and weight of the subject, the condition being treated, the severity of the condition and the form and effective concentration of the cells administered.

[0088] As used herein, the term “endogenous” refers to a nucleic acid molecule or polypeptide that is normally expressed in a cell or tissue.

[0089] As used herein, the term “exogenous” refers to a nucleic acid molecule or polypeptide that is not endogenously present in a cell. The term “exogenous” would therefore encompass any recombinant nucleic acid molecule or polypeptide expressed in a cell, such as foreign, heterologous, and over-expressed nucleic acid molecules and polypeptides. By “exogenous” nucleic acid is meant a nucleic acid not present in a native wild-type cell; for example, an exogenous nucleic acid may vary from an endogenous counterpart by sequence, by position / location, or both. For clarity, an exogenous nucleic acid may have the same or different sequence relative to its native endogenous counterpart; it may be introduced by genetic engineering into the cell itself or a progenitor thereof, and may optionally be linked to alternative control sequences, such as a non-native promoter or secretory sequence.

[0090] By a “heterologous nucleic acid molecule or polypeptide” is meant a nucleic acid molecule (e.g., a cDNA, DNA or RNA molecule) or polypeptide that is not normally present in a cell or sample obtained from a cell. This nucleic acid may be from another organism, or it may be, for example, an mRNA molecule that is not normally expressed in a cell or sample.

[0091] By “modulate” is meant positively or negatively alter. Exemplary modulations include a about 1%, about 2%, about 5%, about 10%, about 25%, about 50%, about 75%, or about 100% change.

[0092] By “increase” is meant to alter positively by at least about 5%. An alteration may be by about 5%, about 10%, about 25%, about 30%, about 50%, about 75%, about 100% or more.

[0093] By “reduce” is meant to alter negatively by at least about 5%. An alteration may be by about 5%, about 10%, about 25%, about 30%, about 50%, about 75%, or even by about 100%.

[0094] The terms “isolated,” “purified,” or “biologically pure” refer to material that is free to varying degrees from components which normally accompany it as found in its native state. “Isolate” denotes a degree of separation from original source or surroundings. “Purify” denotes a degree of separation that is higher than isolation. A “purified” or “biologically pure” protein is sufficiently free of other materials such that any impurities do not materially affect the biological properties of the protein or cause other adverse consequences. That is, a nucleic acid or peptide is purified if it is substantially free of cellular material, viral material, or culture medium when produced by recombinant DNA techniques, or chemical precursors or other chemicals when chemically synthesized. Purity and homogeneity are typically determined using analytical chemistry techniques, for example, polyacrylamide gel electrophoresis or high-performance liquid chromatography. The term “purified” can denote that a nucleic acid or protein gives rise to essentially one band in an electrophoretic gel. For a protein that can be subjected to modifications, for example, phosphorylation or glycosylation, different modifications may give rise to different isolated proteins, which can be separately purified.

[0095] By “isolated cell” is meant a cell that is separated from the molecular and / or cellular components that naturally accompany the cell.

[0096] The term “antigen-binding domain” as used herein refers to a domain capable of specifically binding a particular antigenic determinant or set of antigenic determinants present on a cell.

[0097] By “receptor” is meant a polypeptide, or portion thereof, present on a cell membrane that selectively binds one or more ligand.

[0098] By “signal sequence” or “leader sequence” is meant a peptide sequence (e.g., 5, 10, 15, 20, 25 or 30 amino acids) present at the N-terminus of newly synthesized proteins that directs their entry to the secretory pathway

[0099] The terms “comprises”, “comprising”, and are intended to have the broad meaning ascribed to them in U.S. Patent Law and can mean “includes”, “including” and the like.

[0100] As used herein, “treatment” refers to clinical intervention in an attempt to alter the disease course of the individual or cell being treated, and can be performed either for prophylaxis or during the course of clinical pathology. Therapeutic effects of treatment include, without limitation, preventing occurrence or recurrence of disease, alleviation of symptoms, diminishment of any direct or indirect pathological consequences of the disease, preventing metastases, decreasing the rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. By preventing progression of a disease or disorder, a treatment can prevent deterioration due to a disorder in an affected or diagnosed subject or a subject suspected of having the disorder, but also a treatment may prevent the onset of the disorder or a symptom of the disorder in a subject at risk for the disorder or suspected of having the disorder.

[0101] An “individual” or “subject” herein is a vertebrate, such as a human or non-human animal, for example, a mammal. Mammals include, but are not limited to, humans, primates, farm animals, sport animals, rodents and pets. Non-limiting examples of non-human animal subjects include rodents such as mice, rats, hamsters, and guinea pigs; rabbits; dogs; cats; sheep; pigs; goats; cattle; horses; and non-human primates such as apes and monkeys. As used herein, the term “a conservative sequence modification” refers to an amino acid modification that does not significantly affect or alter the binding characteristics of the presently disclosed chimeric receptors comprising the amino acid sequence. Conservative modifications can include amino acid substitutions, additions and deletions. Modifications can be introduced into the extracellular antigen-binding domain of the presently disclosed chimeric receptors by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Amino acids can be classified into groups according to their physicochemical properties such as charge and polarity. Conservative amino acid substitutions are ones in which the amino acid residue is replaced with an amino acid within the same group. For example, amino acids can be classified by charge: positively-charged amino acids include lysine, arginine, histidine, negatively-charged amino acids include aspartic acid, glutamic acid, neutral charge amino acids include alanine, asparagine, cysteine, glutamine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. In addition, amino acids can be classified by polarity: polar amino acids include arginine (basic polar), asparagine, aspartic acid (acidic polar), glutamic acid (acidic polar), glutamine, histidine (basic polar), lysine (basic polar), serine, threonine, and tyrosine; non-polar amino acids include alanine, cysteine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, and valine. Thus, one or more amino acid residues within a CDR region can be replaced with other amino acid residues from the same group and the altered antibody can be tested for retained function (z.e., the functions set forth in (c) through (1) above) using the functional assays described herein. In certain embodiments, no more than one, no more than two, no more than three, no more than four, no more than five residues within a specified sequence or a CDR region are altered.

[0102] As used herein, the term “reference sample” refers to a representative sample of normal measurement of a biomarker (e.g., CD57, TIM3, KLRG1, CTLA4, LAG3, PD1) obtained from a cohort of healthy subject or a cohort of heathy and diseased subjects. In certain embodiments, the reference sample refers to a baseline amount of a biomarker. In certain embodiments, the reference sample refers to isotype controls. In certain embodiments, the reference sample refers to untransduced (UTD) cells or patient apheresis / starting material (SM). In certain embodiments, the reference sample refers to a measurement of a biomarker (e.g., CD57, TIM3, PD1, KLRG1, CTLA4, LAG3) obtained from NR (no response) patients.

[0103] Other aspects of the presently disclosed subject matter are described in the following disclosure and are within the ambit of the presently disclosed subject matter.

[0104] 2. Cells The presently disclosed subject matter provides cells comprising a chimeric receptor comprising an extracellular antigen-binding domain that binds to CD 19.

[0105] In certain embodiments, the cell is selected from the group consisting of cells of lymphoid lineage and cells of myeloid lineage. In certain embodiments, the cell is an immunoresponsive cell. In certain embodiments, the immunoresponsive cell is a cell of lymphoid lineage.

[0106] In certain embodiments, the cell is a cell of the lymphoid lineage. Cells of the lymphoid lineage can provide the production of antibodies, regulation of the cellular immune system, detection of foreign agents in the blood, detection of cells foreign to the host, and the like. Nonlimiting examples of cells of the lymphoid lineage include T cells, Natural Killer (NK) cells, B cells, dendritic cells, and stem cells from which lymphoid cells may be differentiated. In certain embodiments, the stem cell is a pluripotent stem cell. In certain embodiments, the pluripotent stem cell is an embryonic stem cell (ESC) or an induced pluripotent stem cell (iPSC).

[0107] In certain embodiments, the cell is a T cell. T cells can be lymphocytes that mature in the thymus and are chiefly responsible for cell-mediated immunity. T cells are involved in the adaptive immune system. The T cells of the presently disclosed subject matter can be any type of T cells, including, but not limited to, helper T cells, cytotoxic T cells, memory T cells (including central memory T cells, stem-cell-like memory T cells (or stem-like memory T cells), and two types of effector memory T cells: e.g., TEM cells and TEMRA cells, Regulatory T cells (also known as suppressor T cells), tumor-infiltrating lymphocyte (TIL), Natural killer T cells, Mucosal associated invariant T cells, and y5 T cells. Cytotoxic T cells (CTL or killer T cells) are a subset of T lymphocytes capable of inducing the death of infected somatic or tumor cells. A patient’s own T cells may be genetically modified to target specific antigens through the introduction of a chimeric receptor, e.g., a CAR or a HIT. In certain embodiments, the immunoresponsive cell is a T cell. The T cell can be a CD4+T cell or a CD8+T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is a CD8+T cell.

[0108] In certain embodiments, the cell is an NK cell. Natural killer (NK) cells can be lymphocytes that are part of cell-mediated immunity and act during the innate immune response. NK cells do not require prior activation in order to perform their cytotoxic effect on target cells. In certain embodiments, the cell is a genetically modified NK cell. In certain embodiments, the cell is an edited NK cell. In certain embodiments, the cell is an NK cell derived from a stem cell. In certain embodiments, the cell is an NK cell derived from a pluripotent stem cell. In certain embodiments, the cell is an induced pluripotent stem cell (iPSC)-derived NK cell.

[0109] Types of human lymphocytes of the presently disclosed subject matter include, without limitation, peripheral donor lymphocytes, e.g., those disclosed in Sadelain et al., Nat Rev Cancer (2003); 3 :35-45 (disclosing peripheral donor lymphocytes genetically modified to express CARs), in Morgan, R.A., et al. 2006 Science 314:126-129 (disclosing peripheral donor lymphocytes genetically modified to express a full-length tumor antigen-recognizing T cell receptor complex comprising the a and P heterodimer), in Panelli et al., J Immunol (2000); 164:495-504; Panelli et al., J Immunol (2000); 164:4382-4392 (disclosing lymphocyte cultures derived from tumor infiltrating lymphocytes (TILs) in tumor biopsies), and in Dupont et al., Cancer Res (2005);65:5417-5427; Papanicolaou et al., Blood (2003); 102:2498-2505 (disclosing selectively in vztro-expanded antigen-specific peripheral blood leukocytes employing artificial antigen- presenting cells (AAPCs) or pulsed dendritic cells).

[0110] The cells (e.g., T cells or NK cells) can be autologous, non-autologous (e.g., allogeneic), or derived in vitro from engineered progenitor or stem cells.

[0111] The cells of the presently disclosed subject matter can be cells of the myeloid lineage. Non-limiting examples of cells of the myeloid lineage include monocytes, macrophages, neutrophils, dendritic cells, basophils, neutrophils, eosinophils, megakaryocytes, mast cells, erythrocytes, thrombocytes, and stem cells from which myeloid cells may be differentiated. In certain embodiments, the stem cell is a pluripotent stem cell (e.g., an embryonic stem cell or an induced pluripotent stem cell).

[0112] In certain embodiments, the presently disclosed cells are capable of modulating the tumor microenvironment. Tumors have a microenvironment that is hostile to the host immune response involving a series of mechanisms by malignant cells to protect themselves from immune recognition and elimination. This “hostile tumor microenvironment” comprises a variety of immune suppressive factors including infiltrating regulatory CD4+T cells (Tregs), myeloid- derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), immune suppressive cytokines including TGF-P, and expression of ligands targeted to immune suppressive receptors expressed by activated T cells (CTLA-4 and PD-1). These mechanisms of immune suppression play a role in the maintenance of tolerance and suppressing inappropriate immune responses, however, within the tumor microenvironment, these mechanisms prevent an effective anti-tumor immune response. Collectively these immune suppressive factors can induce either marked anergy or apoptosis of adoptively transferred modified T cells upon encounter with targeted tumor cells.

[0113] In certain embodiments, the cells can be transduced with a CD19-targeted chimeric receptor disclosed herein such that the cells express the chimeric receptor(s).

[0114] 2.1. CD 19

[0115] CD 19 is a cell-surface 95 kDa glycoprotein present on normal B cells from early in their development until differentiation into plasma cells. CD19 is not present in other normal tissues, including pluripotent blood stem cells. It is expressed in B cell lymphomas, acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), hairy cell leukemias, and a subset of acute myelogenous leukemias making it an attractive target for immunotherapy with minimal risk of autoimmune disease (other than B cell aplasia) or irreversible bone marrow toxicity.

[0116] CD 19 is a member of the immunoglobulin superfamily and a component of a cell surface signal transduction complex that includes Leul3, CD81, and CD21, which positively regulates signal transduction through the B cell receptor.

[0117] In certain embodiments, the presently disclosed chimeric receptor (e.g., CAR) binds to human CD 19. In certain embodiments, the human CD 19 comprises or consists of the amino acid sequence with an NCBI Reference No: NP 001171569.1 (SEQ ID NO: 8), or a fragment thereof.

[0118] SEQ ID NO: 8 is provided below:

[0119] MPPPRLLFFLLFLTPMEVRPEEPLWKVEEGDNAVLQCLKGTSDGPTQQLTWSRESPLKPFLKL SLGLPGLGIHMRPLAIWLFI FNVSQQMGGFYLCQPGPPSEKAWQPGWTVNVEGSGELFRWNVSD LGGLGCGLKNRSSEGPSSPSGKLMSPKLYVWAKDRPEIWEGEPPCLPPRDSLNQSLSQDLTMAP GSTLWLSCGVPPDSVSRGPLSWTHVHPKGPKSLLSLELKDDRPARDMWVMETGLLLPRATAQDA GKYYCHRGNLTMSFHLEITARPVLWHWLLRTGGWKVSAVTLAYLI FCLCSLVGILHLQRALVLR RKRKRMTDPTRRFFKVTPPPGSGPQNQYGNVLSLPTPTSGLGRAQRWAAGLGGTAPSYGNPSSD VQADGALGSRSPPGVGPEEEEGEGYEEPDSEEDSEFYENDSNLGQDQLSQDGSGYENPEDEPLG PEDEDSFSNAESYENEDEELTQPVARTMDFLSPHGSAWDPSREATSLAGSQSYEDMRGILYAAP QLRS IRGQPGPNHEEDADSYENMDNPDGPDPAWGGGGRMGTWSTR [ SEQ ID NO : 8 ]

[0120] In certain embodiments, the human CD 19 comprises or consists of the amino acid sequence with an NCBI Reference No: NP_001761.3 (SEQ ID NO: 9), or a fragment thereof. SEQ ID NO: 9 is provided below:

[0121] MPPPRLLFFLLFLTPMEVRPEEPLWKVEEGDNAVLQCLKGTSDGPTQQLTWSRESPLKPFLKL SLGLPGLGIHMRPLAIWLFI FNVSQQMGGFYLCQPGPPSEKAWQPGWTVNVEGSGELFRWNVSD LGGLGCGLKNRSSEGPSSPSGKLMSPKLYVWAKDRPEIWEGEPPCLPPRDSLNQSLSQDLTMAP GSTLWLSCGVPPDSVSRGPLSWTHVHPKGPKSLLSLELKDDRPARDMWVMETGLLLPRATAQDA GKYYCHRGNLTMSFHLEITARPVLWHWLLRTGGWKVSAVTLAYLI FCLCSLVGILHLQRALVLR RKRKRMTDPTRRFFKVTPPPGSGPQNQYGNVLSLPTPTSGLGRAQRWAAGLGGTAPSYGNPSSD VQADGALGSRSPPGVGPEEEEGEGYEEPDSEEDSEFYENDSNLGQDQLSQDGSGYENPEDEPLG PEDEDSFSNAESYENEDEELTQPVARTMDFLSPHGSAWDPSREATSLGSQSYEDMRGILYAAPQ LRS IRGQPGPNHEEDADSYENMDNPDGPDPAWGGGGRMGTWSTR [ SEQ ID NO : 9 ]

[0122] In certain embodiments, the chimeric receptor binds to the extracellular domain of CD 19. In certain embodiments, the chimeric receptor binds to the extracellular domain of human CD 19. In certain embodiments, the extracellular domain of human CD 19 comprises or consists of amino acids 20 to 291 of SEQ ID NO: 8. In certain embodiments, the extracellular domain of human CD19 comprises or consists of amino acids 20 to 291 of SEQ ID NO: 9.

[0123] In certain embodiments, the CD 19 comprises or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, 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% identical to the amino acid sequence set forth in SEQ ID NO: 8 or a fragment thereof.

[0124] In certain embodiments, the CD 19 comprises or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, 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% identical to the amino acid sequence set forth in SEQ ID NO: 9 or a fragment thereof.

[0125] 2.2. Chimeric Receptors

[0126] In certain embodiments, the presently disclosed chimeric receptor comprises an extracellular antigen-binding domain that binds to CD 19. The extracellular antigen-binding domain can be an antigen-binding fragment of an antibody, an antigen-binding fragment of a heavy chain variable region (VH) of an antibody, an antigen-binding fragment of a light chain variable region (VL) of an antibody, a single chain variable fragment (scFv), a Fab, or F(ab)2. In certain embodiments, the extracellular antigen-binding fragment is a single chain variable fragment (scFv). In certain embodiments, the scFv is a human scFv. In certain embodiments, the scFv is a humanized scFv. In certain embodiments, the scFv is a murine scFv. In certain embodiments, the Fab is crosslinked.

[0127] In certain embodiments, the presently disclosed chimeric receptor is a chimeric antigen receptor (CAR). In certain embodiments, the chimeric receptor is a TCR like fusion molecule. 2.2.1. Chimeric Antigen Receptor ( CAR)

[0128] In certain embodiments, the chimeric receptor is a CAR. CARs are engineered receptors, which graft or confer a specificity of interest onto an immune effector cell. CARs can be used to graft the specificity of a monoclonal antibody onto a T cell; with transfer of their coding sequence facilitated by retroviral vectors.

[0129] There are three generations of CARs. “First generation” CARs are typically composed of an extracellular antigen-binding domain (e.g., an scFv), which is fused to a transmembrane domain, which is fused to the cytoplasmic / intracellular signaling domain. “First generation” CARs can provide de novo antigen recognition and cause activation of both CD4+and CD8+T cells through their CD3(^ chain signaling domain in a single fusion molecule, independent of HLA- mediated antigen presentation. “Second generation” CARs add intracellular signaling domains from various co-stimulatory molecules (e.g., CD28, 4- IBB, ICOS, 0X40) to the cytoplasmic tail of the CAR to provide additional signals to the T cell. “Second generation” CARs comprise those that provide both co-stimulation (e.g., CD28 or 4-1BB) and activation (CD3Q. “Third generation” CARs comprise those that provide multiple co-stimulation (e.g., CD28 and 4- IBB) and activation (CD3Q. In certain embodiments, the chimeric receptor is a second generation CAR. In certain embodiments, the chimeric receptor is a CAR that comprises an intracellular domain of a costimulatory molecule or a fragment thereof.

[0130] 2, 2, 1, 1, Extracellular Antigen-Binding Domain of a CAR

[0131] In certain embodiments, the extracellular antigen-binding domain is a single chain variable fragment (scFv). In certain embodiments, the scFv is a human scFv. In certain embodiments, the scFv is a humanized scFv. In certain embodiments, the scFv is a murine scFv. In certain embodiments, the scFv is identified by screening scFv phage library with an antigen-Fc fusion protein.

[0132] In certain embodiments, the extracellular antigen-binding domain is a Fab. In certain embodiments, the Fab is crosslinked. In certain embodiments, the extracellular antigen-binding domain is a F(ab)2. Any of the foregoing molecules may be comprised in a fusion protein with a heterologous sequence to form the extracellular antigen-binding domain.

[0133] Binding of the extracellular antigen-binding domain of a chimeric receptor, e.g., a CAR, can be confirmed by, for example, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), FACS analysis, bioassay (e.g., growth inhibition), or Western Blot assay. Each of these assays generally detect the presence of protein-antibody complexes of particular interest by employing a labeled reagent (e.g., an antibody, or an scFv) specific for the complex of interest. For example, the scFv can be radioactively labeled and used in a radioimmunoassay (RIA) (see, for example, Weintraub, B., Principles of Radioimmunoassay, Seventh Training Course on Radioligand Assay Techniques, The Endocrine Society, March, 1986, which is incorporated by reference herein). The radioactive isotope can be detected by such means as the use of a y counter, a scintillation counter, or by autoradiography. In certain embodiments, the extracellular antigen-binding domain of the CAR is labeled with a fluorescent marker. Nonlimiting examples of fluorescent markers include green fluorescent protein (GFP), blue fluorescent protein (e.g., EBFP, EBFP2, Azurite, and mKalamal), cyan fluorescent protein (e.g., ECFP, Cerulean, and CyPet), and yellow fluorescent protein (e.g., YFP, Citrine, Venus, and YPet).

[0134] In certain embodiments, the extracellular antigen-binding domain of the CAR binds to CD19 (e.g., human CD19) with a dissociation constant (Ka) of at least about 1 x 10'6M, at least about 1 x 10'7M, at least about 1 x 10'8M, at least about 1 x 10'9M, or at least about 1 x 10'10M. In certain embodiments, the extracellular antigen -binding domain of the CAR binds to CD 19 (e.g., human CD 19) with a dissociation constant (Ka) of at least about 2 x 10’8M. In certain embodiments, the extracellular antigen-binding domain of the CAR binds to CD 19 (e.g., human CD 19) with a dissociation constant (Ka) of between about 2 x 10'8M and about 8 x 10'9M.

[0135] In certain embodiments, the extracellular antigen-binding domain of the CAR binds to CD19 (e.g., human CD19) with a dissociation constant (Ka) of between about 1 nM and 50 nM, between about 5 nM and 30 nM, between about 5 nM and 25 nM, or between about 8 nM and 20 nM. In certain embodiments, the extracellular antigen-binding domain of the CAR binds to CD 19 (e.g., human CD 19) with a dissociation constant (Ka) of at least about 50 nM, at least about 40 nM, at least about 35 nM, at least about 30 nM, at least about 25 nM, at least about 20 nM, at least about 19 nM, at least about 18 nM, at least about 17 nM, at least about 16 nM, at least about 15 nM, at least about 14 nM, at least about 13 nM, at least about 12 nM, at least about 11 nM, at least about 10 nM, at least about 9 nM, at least about 8 nM, at least about 7 nM, at least about 6 nM, or at least about 5 nM.

[0136] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10 or a conservative modification thereof, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11 or a conservative modification thereof, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12 or a conservative modification thereof. SEQ ID NOs: 10-12 are provided in Table 1.

[0137] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VL comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13 or a conservative modification thereof, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14 or a conservative modification thereof, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15 or a conservative modification thereof. SEQ ID NOs: 13-15 are provided in Table 1.

[0138] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10 or a conservative modification thereof, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11 or a conservative modification thereof, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12 or a conservative modification thereof; and a VL comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13 or a conservative modification thereof, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14 or a conservative modification, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15 or a conservative modification thereof.

[0139] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12; and a VL comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a VL CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a VL CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15.

[0140] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising an amino acid sequence that is at least about 80% (e.g., at least about 85%, at least about 90%, or at least about 95%) homologous or identical to the amino acid sequence set forth in SEQ ID NO: 16. For example, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising an amino acid sequence that is about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% homologous or identical to SEQ ID NO: 16.

[0141] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 16. SEQ ID NO: 16 is provided in Table 1 below.

[0142] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises a VL comprising an amino acid sequence that is at least about 80% (e.g., at least about 85%, at least about 90%, or at least about 95%) homologous or identical to the amino acid sequence set forth in SEQ ID NO: 17. For example, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g., a CD 19- targeted scFv) comprises a VL comprising an amino acid sequence that is about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% homologous or identical to SEQ ID NO: 17. In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g, a CD19-targeted scFv) comprises a VL comprising the amino acid sequence set forth in SEQ ID NO: 17. SEQ ID NO: 17 is provided in Table 1 below.

[0143] In certain embodiments, the extracellular antigen-binding domain of the CD19-targeted chimeric receptor (e.g, a CD19-targeted scFv) comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 16, and a VL comprising the amino acid sequence set forth in SEQ ID NO: 17. In certain embodiments, the VH and VL are linked via a linker. In certain embodiments, the linker comprises the amino acid sequence set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, or SEQ ID NO: 7.

[0144] In certain embodiments, the variable regions within the extracellular antigen-binding domain of the CD19-targeted chimeric receptor have to be linked one after another such that at the N-terminus of the extracellular antigen-binding domain, a heavy chain variable region (VH) is positioned. In certain embodiments, if the extracellular antigen-binding domain of the CD 19- targeted chimeric receptor is an scFv, the variable regions are positioned from the N- to the C- terminus: VH-VL. In certain embodiments, the CD19-targeted scFv comprises or consists of the amino acid sequence set forth in SEQ ID NO: 18. An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 18 is set forth in SEQ ID NO: 19. SEQ ID NOs: 18 and 19 are provided in Table 1. The CDRs provided in Table 1 are identified according to the Kabat numbering system.

[0145] Table 1

[0146] The VH and / or VL amino acid sequences having at least about 80%, at least about 80%, at least about 85%, at least about 90%, or at least about 95% (e.g., about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99%) homology or identity to a specific sequence (e.g., SEQ ID NO: 16 or SEQ ID NO: 17) may contain substitutions (e.g., conservative substitutions), insertions, or deletions relative to the specified sequence(s), but retain the ability to bind to a target antigen (e.g., CD 19). In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in a specific sequence (e.g., SEQ ID NO: 16 or SEQ ID NO: 17). In certain embodiments, substitutions, insertions, or deletions occur in regions outside the CDRs (e.g., in the FRs) of the extracellular antigen-binding domain. In certain embodiments, the extracellular antigen-binding domain comprises VH and / or VL sequence selected from SEQ ID NO: 16 or SEQ ID NO: 17, including post-translational modifications of that sequence (SEQ ID NO: 16 or SEQ ID NO: 17).

[0147] In addition, the extracellular antigen-binding domain can comprise a leader or a signal peptide that directs the nascent protein into the endoplasmic reticulum. Signal peptide or leader can be essential if the CAR is to be glycosylated and anchored in the cell membrane. The signal sequence or leader can be a peptide sequence (about 5, about 10, about 15, about 20, about 25, or about 30 amino acids long) present at the N-terminus of newly synthesized proteins that directs their entry to the secretory pathway. In certain embodiments, the signal peptide is covalently joined to the 5’ terminus (N-terminus) of the extracellular antigen-binding domain. In certain embodiments, the signal peptide comprises a CD8 polypeptide, e.g., the CAR comprises a truncated CD8 signal peptide. In certain embodiments, the signal peptide is generated from the antibody from it is derived. In certain embodiments, the signal peptide is set forth in SEQ ID NO: 19, which is provided below:

[0148] MDMRVPAQLLGLLLLWLPDTRC [ SEQ ID NO : 19 ]

[0149] Additionally or alternatively, the extracellular antigen-binding domain of the CD 19- targeted chimeric receptor (e.g., a CD19-targeted scFv) comprises an extracellular antigenbinding domain disclosed in International Patent Application No. PCT / US2021 / 029138, the content of which is incorporated by reference in its entirety.

[0150] 2, 2, 1,2, Transmembrane Domain of a CAR

[0151] In certain embodiments, the transmembrane domain of the CAR comprises a hydrophobic alpha helix that spans at least a portion of the membrane. Different transmembrane domains result in different receptor stability. After antigen recognition, receptors cluster and a signal are transmitted to the cell. In accordance with the presently disclosed subject matter, the transmembrane domain of the CAR can comprise a native or modified transmembrane domain of CD8 or a fragment thereof, a native or modified transmembrane domain of CD28 or a fragment thereof, a native or modified transmembrane domain of CD3(^ or a fragment thereof, a native or modified transmembrane domain of CD4 or a fragment thereof, a native or modified transmembrane domain of 4-1BB or a fragment thereof, a native or modified transmembrane domain of 0X40 or a fragment thereof, a native or modified transmembrane domain of ICOS or a fragment thereof, a native or modified transmembrane domain of CD84 or a fragment thereof, a native or modified transmembrane domain of CD 166 or a fragment thereof, a native or modified transmembrane domain of CD8a or a fragment thereof, a native or modified transmembrane domain of CD8b or a fragment thereof, a native or modified transmembrane domain of ICAM-1 or a fragment thereof, a native or modified transmembrane domain of CTLA-4 or a fragment thereof, a native or modified transmembrane domain of CD27 or a fragment thereof, a native or modified transmembrane domain of CD40 or a fragment thereof, NKGD2 or a fragment thereof, or a combination thereof.

[0152] In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide (e.g., a transmembrane domain of CD8 or a fragment thereof).

[0153] In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide (e.g., a transmembrane domain of CD8 or a fragment thereof). In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide (e.g., a transmembrane domain of human CD8 or a fragment thereof). In certain embodiments, the CD8 polypeptide comprises or consists of an amino acid sequence that is at least about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% homologous or identical to the amino acid sequence having a NCBI Reference No: NP_001139345.1 (SEQ ID NO: 20) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD8 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 20, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, at least about 60, at least about 70, and up to about 235 amino acids in length. In certain embodiments, the CD8 polypeptide comprises or consists of amino acids 1 to 235, 1 to 50, 50 to 100, 100 to 150, 150 to 200, 137 to 207, or 200 to 235 of SEQ ID NO: 20. In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide comprising or consisting of amino acids 137 to 207 of SEQ ID NO: 20. SEQ ID NO: 20 is provided below.

[0154] MALPVTALLLPLALLLHAARPSQFRVSPLDRTWNLGETVELKCQVLLSNPTSGCSWLFQPRGAA ASPTFLLYLSQNKPKAAEGLDTQRFSGKRLGDTFVLTLSDFRRENEGYYFCSALSNS IMYFSHF VPVFLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGT CGVLLLSLVITLYCNHRNRRRVCKCPRPWKSGDKPSLSARYV [ SEQ ID NO : 20 ]

[0155] An exemplary nucleotide sequence encoding amino acids 137 to 207 of SEQ ID NO: 20 is set forth in SEQ ID NO: 21, which is provided below.

[0156] CCCACCACGACGCCAGCGCCGCGACCACCAACCCCGGCGCCCACGATCGCGTCGCAGCCCCTGT CCCTGCGCCCAGAGGCGTGCCGGCCAGCGGCGGGGGGCGCAGTGCACACGAGGGGGCTGGACTT CGCCTGTGATATCTACATCTGGGCGCCCCTGGCCGGGACTTGTGGGGTCCTTCTCCTGTCACTG GTTATCACCCTTTACTGCAAC [ SEQ ID NO : 21 ]

[0157] In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide (e.g., a transmembrane domain of mouse CD8 or a fragment thereof). In certain embodiments, the CD8 polypeptide comprises or consists of an amino acid sequence that is at least about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% homologous or identical to the amino acid sequence having a NCBI Reference No: AAA92533.1 (SEQ ID NO: 22) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD8 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 22, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, or at least about 60, or at least about 70, or at least about 100, or at least about 200, and up to about 247 amino acids in length. In certain embodiments, the CD8 polypeptide comprises or consists of amino acids 1 to 247, 1 to 50, 50 to 100, 100 to 150, 150 to 200, 151 to 219, or 200 to 247 of SEQ ID NO: 22. In certain embodiments, the transmembrane domain of the CAR comprises a CD8 polypeptide comprising or consisting of amino acids 151 to 219 of SEQ ID NO: 22. SEQ ID NO: 22 is provided below.

[0158] MASPLTRFLSLNLLLMGES I ILGSGEAKPQAPELRI FPKKMDAELGQKVDLVCEVLGSVSQGCS WLFQNSSSKLPQPTFWYMASSHNKITWDEKLNSSKLFSAVRDTNNKYVLTLNKFSKENEGYYF CSVISNSVMYFSSWPVLQKVNSTTTKPVLRTPSPVHPTGTSQPQRPEDCRPRGSVKGTGLDFA GDI YIWAPLAGICVAPLLSLI ITLICYHRSRKRVCKCPRPLVRQEGKPRPSEKIV [ SEQ ID NO : 22 ]

[0159] In certain embodiments, the transmembrane domain of a presently disclosed CAR comprises a CD28 polypeptide (e.g., a transmembrane domain of CD28 or a fragment thereof).

[0160] In certain embodiments, the transmembrane domain of the CAR comprises a CD28 polypeptide e.g., a transmembrane domain of human CD28 or a fragment thereof). In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is at least about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99% or 100% homologous or identical to the amino acid sequence having an NCBI Reference No: NP 006130 (SEQ ID NO: 23) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 10, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, and up to about 220 amino acids in length. In certain embodiments, the CD28 polypeptide comprises or consists of amino acids 1 to 220, 1 to 50, 50 to 100, 100 to 150, 150 to 200, 153 to 179, or 200 to 220 of SEQ ID NO: 23. In certain embodiments, the transmembrane domain of the CAR comprises a CD28 polypeptide comprising or consisting of amino acids 153 to 179 of SEQ ID NO: 23. SEQ ID NO: 23 is provided below: MLRLLLALNLFPS IQVTGNKILVKQSPMLVAYDNAVNLSCKYSYNLFSREFRASLHKGLDSAVE VCWYGNYSQQLQVYSKTGFNCDGKLGNESVTFYLQNLYVNQTDI YFCKIEVMYPPPYLDNEKS NGTI IHVKGKHLCPSPLFPGPSKPFWVLVWGGVLACYSLLVTVAFI I FWVRSKRSRLLHSDYM NMTPRRPGPTRKHYQPYAPPRDFAAYRS [ SEQ ID NO : 23 ]

[0161] An exemplary nucleotide sequence encoding amino acids 153 to 179 of SEQ ID NO: 23 is set forth in SEQ ID NO: 24, which is provided below.

[0162] TTTTGGGTGCTGGTGGTGGTTGGTGGAGTCCTGGCTTGCTATAGCTTGCTAGTAACAGTGGCCT TTATTATTTTCTGGGTG [ SEQ ID NO : 24 ]

[0163] In certain embodiments, the transmembrane domain of the CAR comprises a CD28 polypeptide (e.g., a transmembrane domain of mouse CD28 or a fragment thereof). In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is at least about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or 100% homologous or identical to the amino acid sequence having an NCBI Reference No: NP 031668.3 (SEQ ID NO: 25) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 25, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, and up to about 218 amino acids in length. In certain embodiments, the CD28 polypeptide comprises or consists of amino acids 1 to 220, 1 to 50, 50 to 100, 100 to 150, 150 to 200, 151 to 177, or 200 to 218 of SEQ ID NO: 25. In certain embodiments, the transmembrane domain of the CAR comprises a CD28 polypeptide comprising or consisting of amino acids 151 to 177 of SEQ ID NO: 25. SEQ ID NO: 25 is provided below:

[0164] MTLRLLFLALNFFSVQVTENKILVKQSPLLWDSNEVSLSCRYSYNLLAKEFRASLYKGVNSDV EVCVGNGNFTYQPQFRSNAEFNCDGDFDNETVTFRLWNLHVNHTDI YFCKIEFMYPPPYLDNER SNGTI IHIKEKHLCHTQSSPKLFWALVWAGVLFCYGLLVTVALCVIWTNSRRNRLLQSDYMNM TPRRPGLTRKPYQPYAPARDFAAYRP [ SEQ ID NO : 25 ]

[0165] In certain embodiments, the CAR further comprises a spacer region that links the extracellular antigen-binding domain to the transmembrane domain. The spacer region can be flexible enough to allow the antigen binding domain to orient in different directions to facilitate antigen recognition while preserving the activating activity of the CAR.

[0166] In certain embodiments, the hinge / spacer region of the CAR comprises a native or modified hinge region of CD8 or a fragment thereof, a native or modified hinge region of CD28 or a fragment thereof, a native or modified hinge region of CD3^ or a fragment thereof, a native or modified hinge region of CD40 or a fragment thereof, a native or modified hinge region of 4- IBB or a fragment thereof, a native or modified hinge region of 0X40 or a fragment thereof, a native or modified hinge region of CD84 or a fragment thereof, a native or modified hinge region of CD 166 or a fragment thereof, a native or modified hinge region of CD8a or a fragment thereof, a native or modified hinge region of CD8b or a fragment thereof, a native or modified hinge region of ICOS or a fragment thereof, a native or modified hinge region of ICAM-1 or a fragment thereof, a native or modified hinge region of CTLA-4 or a fragment thereof, a native or modified hinge region of CD27 or a fragment thereof, a native or modified hinge region of CD40 or a fragment thereof, a native or modified hinge region of NKGD2 or a fragment thereof, a synthetic polypeptide (not based on a protein associated with the immune response), or a combination thereof. The hinge / spacer region can be the hinge region from IgGl, the CH2CH3 region of immunoglobulin and portions of CD3, a portion of a CD28 polypeptide (e.g. , a portion of SEQ ID NO: 23 or SEQ ID NO: 25), a portion of a CD8 polypeptide (e.g, a portion of SEQ ID NO: 20 or SEQ ID NO: 22), a variation of any of the foregoing which is at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% homologous or identical thereto, or a synthetic spacer sequence.

[0167] In certain embodiments, the hinge / spacer region of the CAR comprises a CD28 polypeptide. In certain embodiments, the hinge / spacer region of the CAR comprises a CD28 polypeptide comprising or consisting of amino acids 114 to 152 of SEQ ID NO: 23.

[0168] An exemplary nucleotide sequence encoding amino acids 114 to 152 of SEQ ID NO: 23 is set forth in SEQ ID NO: 26, which is provided below.

[0169] ATT GAAG TTATGTATCCTCCTCCTTACC T AGAC AAT GAGAAGAG C AAT G GAAC CATTATCCATG TGAAAGGGAAACACCTTTGTCCAAGTCCCCTATTTCCCGGACCTTCTAAGCCC [ SEQ ID NO : 26 ]

[0170] 2, 2, 1,3, Intracellular Signaling Domain of a CAR

[0171] In certain embodiments, the CAR comprises an intracellular signaling domain. In certain embodiments, the intracellular signaling domain of the CAR comprises a CD3(^ polypeptide. CD3(^ can activate or stimulate a cell (e.g, a cell of the lymphoid lineage, e.g., a T cell). Wild type (“native”) CD3(^ comprises three functional immunoreceptor tyrosine-based activation motifs (ITAMs), and three functional basic-rich stretch (BRS) regions (BRS1, BRS2, and BRS3). CD3(^ transmits an activation signal to the cell (e.g., a cell of the lymphoid lineage, e.g., a T cell) after the antigen is bound. The intracellular signaling domain of the CD3(^-chain is the primary transmitter of signals from endogenous TCRs.

[0172] In certain embodiments, the intracellular signaling domain of the CAR comprises a native CD3(^. In certain embodiments, the native CD3(^ polypeptide comprises or consists of an amino acid sequence that is at least about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% homologous or identical to the amino acid sequence having an NCBI Reference No: NP_932170 (SEQ ID NO: 27) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the native CD3(^ polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 27, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, at least about 100, at least about 110, and up to about 164 amino acids in length. In certain embodiments, the native CD3(^ polypeptide comprises or consists of amino acids 1 to 164, 1 to 50, 50 to 100, 52 to 164, 100 to 150, or 150 to 164 of SEQ ID NO: 27. In certain embodiments, the intracellular signaling domain of the CAR comprises a CD3(^ polypeptide comprising or consisting of amino acids 52 to 164 of SEQ ID NO: 27. SEQ ID NO: 27 is provided below:

[0173] MKWKALFTAAILQAQLPITEAQSFGLLDPKLCYLLDGILFIYGVILTALFLRVKFSRSADAPAY QQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPQRRKNPQEGLYNELQKDKMAEAYSEIGM KGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR [ SEQ ID NO : 27 ]

[0174] In certain embodiments, the intracellular signaling domain of the CAR comprises a modified CD3(^ polypeptide. In certain embodiments, the modified CD3(^ polypeptide comprises one, two, or three ITAMs. In certain embodiments, the modified CD3(^ polypeptide comprises a native IT AMI. In certain embodiments, the native IT AMI comprises or consists of the amino acid sequence set forth in SEQ ID NO: 28.

[0175] QNQLYNELNLGRREEYDVLDKR [ SEQ ID NO : 28 ]

[0176] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 28 is set forth in SEQ ID NO: 29, which is provided below.

[0177] CAGAACCAGCTCTATAACGAGCTCAATCTAGGACGAAGAGAGGAGTACGATGTTTTGGACAAGA GA [ SEQ ID NO : 29 ]

[0178] In certain embodiments, the modified CD3(^ polypeptide comprises an ITAM1 variant comprising one or more loss-of-function mutations. In certain embodiments, the ITAM1 variant comprises or consists of two loss-of-function mutations. In certain embodiments, each of the one or more (e.g. , two) loss of function mutations comprises a mutation of a tyrosine residue in IT AMI . In certain embodiments, the IT AMI variant consists of two loss-of-function mutations. In certain embodiments, the ITAM1 variant comprises or consists of the amino acid sequence set forth in SEQ ID NO: 30, which is provided below. QNQLFNELNLGRREEFDVLDKR [ SEQ ID NO : 30 ]

[0179] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 30 is set forth in SEQ ID NO: 31, which is provided below.

[0180] CAGAACCAGCTCTTTAACGAGCTCAATCTAGGACGAAGAGAGGAGTTCGATGTTTTGGACAAGA GA [ SEQ ID NO : 31 ]

[0181] In certain embodiments, the modified CD3(^ polypeptide comprises a native ITAM2. In certain embodiments, the native ITAM2 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 32, which is provided below.

[0182] QEGLYNELQKDKMAEAYSEIGMK [ SEQ ID NO : 32 ]

[0183] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 32 is set forth in SEQ ID NO: 33, which is provided below.

[0184] CAGGAAGGCCTGTACAATGAACTGCAGAAAGATAAGATGGCGGAGGCCTACAGTGAGATTGGGA TGAAA [ SEQ ID NO : 33 ]

[0185] In certain embodiments, the modified CD3(^ polypeptide comprises an ITAM2 variant. In certain embodiments, the ITAM2 variant comprises or consists of one or more loss-of-function mutations. In certain embodiments, the ITAM2 variant comprises or consists of two loss-of- function mutations. In certain embodiments, each of the one or more (e.g., two) loss of function mutations comprises a mutation of a tyrosine residue in ITAM2. In certain embodiments, the ITAM2 variant consists of two loss-of-function mutations. In certain embodiments, the ITAM2 variant comprises or consists of the amino acid sequence set forth in SEQ ID NO: 21, which is provided below.

[0186] QEGLFNELQKDKMAEAFSEIGMK [ SEQ ID NO : 34 ]

[0187] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 34 is set forth in SEQ ID NO: 35, which is provided below.

[0188] CAGGAAGGCCTGTTCAATGAACTGCAGAAAGATAAGATGGCGGAGGCCTTCAGTGAGATTGGGA TGAAA [ SEQ ID NO : 35 ]

[0189] In certain embodiments, the modified CD3(^ polypeptide comprises a native ITAM3. In certain embodiments, the native ITAM3 comprises or consists of the amino acid sequence set forth in SEQ ID NO: 36, which is provided below. HDGLYQGLSTATKDTYDALHMQ [ SEQ ID NO : 36 ]

[0190] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 36 is set forth in SEQ ID NO: 37, which is provided below.

[0191] CACGATGGCCTTTACCAGGGTCTCAGTACAGCCACCAAGGACACCTACGACGCCCTTCACATGC AG [ SEQ ID NO : 37 ]

[0192] In certain embodiments, the modified CD3(^ polypeptide comprises an ITAM3 variant. In certain embodiments, the IT AMS variant comprises or consists of two loss-of-function mutations. In certain embodiments, each of the one or more (e.g., two) loss of function mutations comprises a mutation of a tyrosine residue in ITAM3. In certain embodiments, the ITAM3 variant comprises or consists of two loss-of-function mutations. In certain embodiments, the ITAM3 variant comprises or consists of the amino acid sequence set forth in SEQ ID NO: 38, which is provided below.

[0193] HDGLFQGLSTATKDTFDALHMQ [ SEQ ID NO : 38 ]

[0194] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 38 is set forth in SEQ ID NO: 39, which is provided below.

[0195] CACGATGGCCTTTTCCAGGGTCTCAGTACAGCCACCAAGGACACCTTCGACGCCCTTCACATGC AG [ SEQ ID NO : 39 ]

[0196] Various modified CD3(^ polypeptides and CARs comprising modified CD3(^ polypeptides are disclosed in International Patent Application Publication No. WO2019 / 133969, which is incorporated by reference hereby in its entirety.

[0197] In certain embodiments, the intracellular signaling domain of the CAR comprises a modified CD3(^ polypeptide comprising a native ITAM1, an ITAM2 variant comprising or consisting of one or more (e.g., two) loss-of-function mutations, and an ITAM3 variant comprising or consisting of one or more (e.g., two) loss-of-function mutations. In certain embodiments, the intracellular signaling domain of the CAR comprises a modified CD3(^ polypeptide comprising a native IT AMI, an ITAM2 variant consisting of two loss-of-function mutations, and an ITAM3 variant consisting of two loss-of-function mutations. In certain embodiments, the intracellular signaling domain of the CAR comprises a modified CD3(^ polypeptide comprising a native IT AMI consisting of the amino acid sequence set forth in SEQ ID NO: 28, an ITAM2 variant consisting of the amino acid sequence set forth in SEQ ID NO: 34, and an ITAM3 variant consisting of the amino acid sequence set forth in SEQ ID NO: 38. In certain embodiments, the modified CD3(^ polypeptide is designated as “1XX”. In certain embodiments, the modified CD3(^ polypeptide comprises or consists of the amino acid sequence set forth in SEQ ID NO: 40. SEQ ID NO: 40 is provided below.

[0198] RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLFNELQK DKMAEAFSEIGMKGERRRGKGHDGLFQGLSTATKDTFDALHMQALPPR [ SEQ ID NO : 40 ]

[0199] In certain embodiments, the intracellular signaling domain of the CAR comprises a modified CD3(^ polypeptide comprising or consisting of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, at least about 100% identical to SEQ ID NO: 40 or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions.

[0200] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 40 is set forth in SEQ ID NO: 41, which is provided below.

[0201] AGAGTGAAGTTCAGCAGGAGCGCAGACGCCCCCGCGTACCAGCAGGGCCAGAACCAGCTCTATA ACGAGCTCAATCTAGGACGAAGAGAGGAGTACGATGTTTTGGACAAGAGACGTGGCCGGGACCC TGAGATGGGGGGAAAGCCGAGAAGGAAGAACCCTCAGGAAGGCCTGTTCAATGAACTGCAGAAA GATAAGATGGCGGAGGCCTTCAGTGAGATTGGGATGAAAGGCGAGCGCCGGAGGGGCAAGGGGC ACGATGGCCTTTTCCAGGGTCTCAGTACAGCCACCAAGGACACCTTCGACGCCCTTCACATGCA GGCCCTGCCCCCTCGC [ SEQ ID NO : 41 ]

[0202] In certain embodiments, the CAR is a second-generation CAR. In certain embodiments, the intracellular signaling domain of the CAR further comprises at least a co-stimulatory signaling region. In certain embodiments, the co-stimulatory signaling region comprises an intracellular domain of at least one co-stimulatory molecule or a fragment thereof.

[0203] As used herein, a “co-stimulatory molecule” refers to a cell surface molecule other than antigen receptor or its ligand that can provide an efficient response of lymphocytes to an antigen. In certain embodiments, a co-stimulatory molecule can provide optimal lymphocyte activation. Non-limiting examples of co-stimulatory molecules include CD28, 4-1BB, 0X40, ICOS, DAP- 10, CD27, CD40, and NKGD2. The co-stimulatory molecule can bind to a co-stimulatory ligand, which is a protein expressed on cell surface that upon binding to its receptor produces a co- stimulatory response, i.e., an intracellular response that effects the stimulation provided when an chimeric receptor (e.g., a chimeric antigen receptor (CAR)) binds to its target antigen. As one example, a 4-1BB ligand (i.e., 4-1BBL) may bind to 4-1BB for providing an intracellular signal that in combination with a CAR signal induces an effector cell function of the CAR+ T cell. In certain embodiments, the intracellular signaling domain of the CAR comprises a costimulatory signaling region that comprises a CD28 polypeptide, e.g., an intracellular domain of CD28 or a fragment thereof. In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO: 23 or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 23, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, and up to about 220 amino acids in length. In certain embodiments, the CD28 polypeptide comprises or consists of amino acids 1 to 220, 1 to 50, 50 to 100, 100 to 150, 114 to 220, 150 to 200, 180 to 220, or 200 to 220 of SEQ ID NO: 25. In certain embodiments, the intracellular signaling domain of the CAR comprises a co-stimulatory signaling region that comprises a CD28 polypeptide comprising or consisting of amino acids 180 to 220 of SEQ ID NO: 23.

[0204] An exemplary nucleic acid sequence encoding amino acids 180 to 220 of SEQ ID NO: 23 is set forth in SEQ ID NO: 42, which is provided below.

[0205] AGGAGTAAGAGGAGCAGGCTCCTGCACAGTGACTACATGAACATGACTCCCCGCCGCCCCGGGC CCACCCGCAAGCATTACCAGCCCTATGCCCCACCACGCGACTTCGCAGCCTATCGCTCC [ SEQ ID NO : 42 ]

[0206] In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, at least about 100% homologous or identical to the amino acid sequence set forth in SEQ ID NO: 25 or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the CD28 polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 25, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, and up to 218 amino acids in length. In certain embodiments, the CD28 polypeptide comprises or consists of amino acids 1 to 218, 1 to 50, 50 to 100, 100 to 150, 150 to 218, 178 to 218, or 200 to 218 of SEQ ID NO: 5. In certain embodiments, the co-stimulatory signaling region of a presently disclosed CAR comprises a CD28 polypeptide that comprises or consists of the amino acids 178 to 218 of SEQ ID NO: 25.

[0207] In certain embodiments, the intracellular signaling domain of the CAR comprises a co- stimulatory signaling region that comprises a 4-1BB polypeptide, e.g., an intracellular domain of 4-1BB or a fragment thereof (e.g., an intracellular domain of human 4-1BB or a fragment thereof). The 4-1BB polypeptide can comprise or consists of an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99%, at least about 100% homologous or identical to the amino acid sequence having a NCBI Ref. No.: NP_001552 (SEQ ID NO: 43) or a fragment thereof, and / or may optionally comprise up to one or up to two or up to three conservative amino acid substitutions. In certain embodiments, the 4-1BB polypeptide comprises or consists of an amino acid sequence that is a consecutive portion of SEQ ID NO: 43, which is at least about 20, or at least about 30, or at least about 40, or at least about 50, or at least about 100, or at least about 150, or at least about 150, and up to about 255 amino acids in length. In certain embodiments, the 4-1BB polypeptide comprises or consists of amino acids 1 to 255, 1 to 50, 50 to 100, 100 to 150, 150 to 200, 200 to 255, or 214 to 255 of SEQ ID NO: 43. In certain embodiments, the intracellular signaling domain of the CAR comprises a co-stimulatory signaling region that comprises a 4- IBB polypeptide comprising or consisting of amino acids 214 to 255 of SEQ ID NO: 43. SEQ ID NO: 43 is provided below.

[0208] MGNSCYNIVATLLLVLNFERTRSLQDPCSNCPAGTFCDNNRNQICSPCPPNSFSSAGGQRTCDI CRQCKGVFRTRKECSSTSNAECDCTPGFHCLGAGCSMCEQDCKQGQELTKKGCKDCCFGTFNDQ KRGICRPWTNCSLDGKSVLVNGTKERDWCGPSPADLSPGASSVTPPAPAREPGHSPQI I SEEL ALTSTALLFLLFFLTLRFSWKRGRKKLLYI FKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL [ SEQ ID NO : 43 ]

[0209] An exemplary nucleic acid sequence encoding amino acids 214 to 255 of SEQ ID NO: 43 is set forth in SEQ ID NO: 44, which is provided below.

[0210] AAACGGGGCAGAAAGAAGCTCCTGTATATATTCAAACAACCATTTATGAGACCAGTACAAACTA CTCAAGAGGAAGATGGCTGTAGCTGCCGATTTCCAGAAGAAGAAGAAGGAGGATGTGAACTG [ SEQ ID NO : 44 ]

[0211] In certain embodiments, the intracellular signaling domain of the CAR comprises a costimulatory signaling region that comprises intracellular domains of two or more co-stimulatory molecules or portions thereof, e.g., an intracellular domain of CD28 or a fragment thereof and an intracellular domain of 4-1BB or a fragment thereof, or an intracellular domain of CD28 or a fragment thereof and an intracellular domain of 0X40 or a fragment thereof.

[0212] 2, 2, 1,4, Exemplified CAR

[0213] In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CD19-targeted CAR comprises (a) an extracellular antigen-binding domain comprising (i) a VH comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12, and (ii) a VL comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15, (b) a hinge domain comprising a CD28 polypeptide (e.g., a hinge domain of human CD28 or a portion thereof, e.g., a CD28 polypeptide consisting of the amino acid sequence of amino acids 114 to 153 of SEQ ID NO: 23); (c) a transmembrane domain comprising a CD28 polypeptide (e.g., a transmembrane domain of human CD28 or a portion thereof, e.g., a CD28 polypeptide consisting of the amino acid sequence of amino acids 154 to 179 of SEQ ID NO: 23), and (d) an intracellular signaling domain comprising (i) a CD3(^ polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 40, and (ii) a costimulatory signaling region comprising a CD28 polypeptide (e.g., an intracellular domain of human CD28 or a portion thereof, e.g., a CD28 polypeptide consisting of the amino acid sequence of amino acids 180 to 220 of SEQ ID NO: 23). In certain embodiments, the VH and VL are linked via a linker consisting of the amino acid sequence set forth in SEQ ID NO: 1. In certain embodiments, the VH and VL are positioned from the N- to the C-terminus: VL-VH. In certain embodiments, the CAR is designed as “19(T2)28zlXX”. In certain embodiments, the CAR comprises the amino acid sequence set forth in SEQ ID NO: 45, which is provided below.

[0214] MDMRVPAQLLGLLLLWLPDTRCEIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPG QAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQAGAVPITFGGGTKVE IKGGGGSGGGGSGGGGSQVQLVESGGGWQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEW VALIWYDGSNKYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKPVEGLLRGFDYWG QGTLVTVSSRAAAIEVMYPPPYLDNEKSNGTI IHVKGKHLCPSPLFPGPSKPFWVLVWGGVLA CYSLLVTVAFI I FWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLFNELQKDKMAEAFS EIGMKGERRRGKGHDGLFQGLSTATKDTFDALHMQALPPR [ SEQ ID NO : 45 ]

[0215] An exemplary nucleic acid sequence encoding the amino acid sequence of SEQ ID NO: 45 is set forth in SEQ ID NO: 46, which is provided below.

[0216] ATGGATATGAGAGTACCAGCTCAGCTGCTGGGCCTGCTGCTTTTGTGGTTGCCGGACACACGCT GTGAGATTGTCCTGACTCAGTCTCCCGGGACTCTGTCCCTCAGCCCCGGTGAACGCGCTACCCT TTCATGCAGAGCCTCTCAGTCTGTGTCCAGCAGCTACCTCGCATGGTATCAGCAGAAGCCCGGA CAGGCTCCCAGGCTGTTGATCTATGGAGCTAGTAGTCGAGCAACAGGCATCCCAGATCGCTTCT CAGGGAGCGGTTCAGGTACAGACTTCACGCTGACGATTTCAAGGCTGGAACCCGAAGATTTTGC CGTCTATTATTGTCAACAGGCAGGGGCTGTGCCAATCACTTTCGGGGGCGGGACCAAGGTGGAA ATCAAAGGAGGCGGAGGAAGTGGAGGAGGAGGGAGCGGTGGAGGAGGGTCACAGGTGCAGCTGG TAGAATCTGGCGGAGGGGTCGTTCAACCAGGGAGGTCATTGCGGTTGAGCTGCGCAGCGAGTGG TTTTACCTTCAGCAGTTATGGAATGCATTGGGTGAGACAAGCACCAGGAAAAGGTCTGGAGTGG GTGGCTTTGATTTGGTACGACGGCAGTAATAAATACTACGCCGATTCTGTTAAGGGCAGATTTA CTATTTCTCGCGACAACAGCAAGAACACGCTGTACCTGCAGATGAACTCTCTGAGAGCCGAAGA TACAGCAGTGTACTATTGTGCTAAGCCCGTAGAAGGGCTCCTGAGGGGATTCGATTATTGGGGC CAGGGTACGCTTGTGACAGTGTCTAGTCGGGCGGCCGCAATTGAAGTTATGTATCCTCCTCCTT AC C T AGAC AAT GAGAAGAG C AAT G GAAC CAT TAT C CAT G T GAAAG G GAAAC AC C T T T G T C C AAG TCCCCTATTTCCCGGACCTTCTAAGCCCTTTTGGGTGCTGGTGGTGGTTGGTGGAGTCCTGGCT TGCTATAGCTTGCTAGTAACAGTGGCCTTTATTATTTTCTGGGTGAGGAGTAAGAGGAGCAGGC TCCTGCACAGTGACTACATGAACATGACTCCCCGCCGCCCCGGGCCCACCCGCAAGCATTACCA GCCCTATGCCCCACCACGCGACTTCGCAGCCTATCGCTCCAGAGTGAAGTTCAGCAGGAGCGCA GACGCCCCCGCGTACCAGCAGGGCCAGAACCAGCTCTATAACGAGCTCAATCTAGGACGAAGAG AGGAGTACGATGTTTTGGACAAGAGACGTGGCCGGGACCCTGAGATGGGGGGAAAGCCGAGAAG GAAGAACCCTCAGGAAGGCCTGTTCAATGAACTGCAGAAAGATAAGATGGCGGAGGCCTTCAGT GAGATTGGGATGAAAGGCGAGCGCCGGAGGGGCAAGGGGCACGATGGCCTTTTCCAGGGTCTCA GTACAGCCACCAAGGACACCTTCGACGCCCTTCACATGCAGGCCCTGCCCCCTCGC [ SEQ ID NO : 4 6 ]

[0217] 2.3. TCR like Fusion Molecules

[0218] In certain embodiments, the chimeric receptor is a TCR like fusion molecule. Nonlimiting examples of TCR fusion molecules include HLA-Independent TCR-based Chimeric Antigen Receptor (also known as “HIT”, e.g, those disclosed in International Patent Application No. PCT / US19 / 017525, which is incorporated by reference in its entirety), T cell receptor fusion constructs (TRuCs) (e.g, those disclosed in Baeuerle et al., “Synthetic TRuC receptors engaging the complete T cell receptor for potent anti -turn or response,” Nature Communications volume 10, Article number: 2087 (2019), which is incorporated by reference in its entirety), synthetic T cell receptor and antigen receptor (STAR) (e.g., those disclosed in Liu et al. Science Translational Medicine (2021); 13(586):eabb5191, which is incorporated by reference in its entirety), antibody- T-cell receptor (AbTCR) (e.g., those disclosed in Xu et al. Cell Discovery (2018) 4:62, which is incorporated by reference in its entirety), and T cell antigen coupler (TAC) (e.g., those disclosed in Helsen et al. Nature Communications (2018);9:3049, which is incorporated by reference in its entirety).

[0219] In certain embodiments, the TCR like fusion molecule comprises an antigen binding chain that comprises an extracellular antigen-binding domain and a constant domain, wherein the TCR like fusion molecule binds to an antigen in an HLA-independent manner. In certain embodiments, the constant domain comprises a T cell receptor constant region selected from the group consisting of a native or modified TRAC polypeptide, a native or modified TRBC polypeptide, a native or modified TRDC polypeptide, a native or modified TRGC polypeptide and any variants or functional fragments thereof. In certain embodiments, the constant domain comprises a native or modified TRAC polypeptide. In certain embodiments, the constant domain comprises a native or modified TRBC polypeptide. In certain embodiments, the constant domain is capable of forming a homodimer or a heterodimer with another constant domain. In certain embodiments, the antigen binding chain is capable of associating with a CD3(^ polypeptide. In certain embodiments, the antigen binding chain, upon binding to an antigen (e.g., CD 19), is capable of activating the CD3^ polypeptide associated to the antigen binding chain. In certain embodiments, the activation of the CD3(^ polypeptide is capable of activating an immunoresponsive cell. In certain embodiments, the TCR like fusion molecule is capable of integrating with a CD3 complex and providing HLA-independent antigen recognition. In certain embodiments, the TCR like fusion molecule replaces an endogenous TCR in a CD3 / TCR complex. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule is capable of dimerizing with another extracellular antigen-binding domain. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises a ligand for a cell-surface receptor, a receptor for a cell surface ligand, an antigen binding portion of an antibody or a fragment thereof or an antigen binding portion of a TCR. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises one or two immunoglobulin variable region(s). In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises a heavy chain variable region (VH) of an antibody. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises a light chain variable region (VL) of an antibody. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule is capable of dimerizing with another extracellular antigen-binding domain. In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises a VH of an antibody, wherein the VH is capable of dimerizing with another extracellular antigen-binding domain comprising a VL of the antibody and form a fragment variable (Fv). In certain embodiments, the extracellular antigen-binding domain of the TCR like fusion molecule comprises a VL of an antibody, wherein the VL is capable of dimerizing with another extracellular antigen-binding domain comprising a VH of the antibody and form a fragment variable (Fv).

[0220] 2.4. Exemplified Cells

[0221] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell.

[0222] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+.

[0223] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+.

[0224] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1".

[0225] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CTLA4'.

[0226] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is LAG3'.

[0227] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is PD1+.

[0228] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+.

[0229] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1".

[0230] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+CTLA4'.

[0231] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+LAG3'. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+PD1+.

[0232] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1".

[0233] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+CTLA4'.

[0234] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+LAG3'.

[0235] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+PD1+.

[0236] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" CTLA4'.

[0237] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" LAG3'.

[0238] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" PD1+.

[0239] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CTLA4" LAG3".

[0240] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CTLA4" PD1+.

[0241] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is LAG3" PD1+.

[0242] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRG1".

[0243] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4".

[0244] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+LAG3-.

[0245] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+PD1+.

[0246] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRGl" CTLA4". In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1" LAGS’.

[0247] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1" PD1+.

[0248] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+CTLA4' LAGS’.

[0249] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+CTLA4' PD1+.

[0250] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+LAG3' PD1+.

[0251] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’.

[0252] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" LAG3-. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" PD1+.

[0253] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+CTLA4' LAG3;

[0254] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+CTLA4' PD1+.

[0255] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+LAG3' PD1+.

[0256] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" CTLA4' LAG3;

[0257] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" CTLA4' PD1+.

[0258] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRG1" LAG3' PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CTLA4' LAG3' PD1+.

[0259] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGL CTLA4’.

[0260] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGL LAG3’.

[0261] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGC PD1+.

[0262] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’ LAG3’.

[0263] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’ PD1+.

[0264] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+LAG3 PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1" CTLA4’ LAG3’.

[0265] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1" CTLA4’ PD1+.

[0266] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRG1" LAG3 PD1+.

[0267] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+CTLA4' LAG3 PD1+.

[0268] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’ LAG3’.

[0269] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’ PD1+.

[0270] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" LAG3 PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+CTLA4" LAG3 PD1+.

[0271] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is KLRGl" CTLA4" LAG3 PD1+.

[0272] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" CTLA4" LAG3".

[0273] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" CTLA4" PD1+.

[0274] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRG l" LAG3" PD I .

[0275] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4" LAG3 PD1+.

[0276] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+KLRGl" CTLA4" LAG3 PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’ LAG3 PD1+.

[0277] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD8+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRG1" CTLA4’ LAG3' PD1+.

[0278] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell.

[0279] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+.

[0280] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+.

[0281] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRG1".

[0282] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CTLA4'.

[0283] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is LAG3'. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is PD1+.

[0284] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+.

[0285] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1".

[0286] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+CTLA4'.

[0287] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+LAG3'.

[0288] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+PD1+.

[0289] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1".

[0290] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+CTLA4'.

[0291] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+LAG3'.

[0292] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+PD1+.

[0293] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRG1" CTLA4'.

[0294] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRG1" LAG3'.

[0295] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRG1" PD1+.

[0296] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CTLA4' LAG3'.

[0297] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CTLA4' PD1+.

[0298] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is LAG3' PD1+.

[0299] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl". In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’.

[0300] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+LAG3-.

[0301] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+PD1+.

[0302] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" CTLA4’.

[0303] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" LAG3-.

[0304] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" PD1+.

[0305] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+CTLA4' LAG3-. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+CTLA4' PD1+.

[0306] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+LAG3' PD1+.

[0307] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’.

[0308] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" LAG3-.

[0309] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" PD1+.

[0310] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+CTLA4' LAG3-.

[0311] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+CTLA4' PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+LAG3' PD1+.

[0312] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRGl" CTLA4" LAG3;

[0313] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRGl" CTLA4" PD1+.

[0314] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRGl" LAG3" PD1+.

[0315] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CTLA4" LAG3" PD1+.

[0316] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" CTLA4".

[0317] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" LAG3". In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRG1 PD1+.

[0318] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’ LAG3’.

[0319] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’ PD1+.

[0320] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+LAG3 PD1+.

[0321] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" CTLA4’ LAG3’.

[0322] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" CTLA4’ PD1+.

[0323] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRG1" LAG3 PD1+. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+CTLA4' LAG3 PD1+.

[0324] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’ LAG3’.

[0325] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" CTLA4’ PD1+.

[0326] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRG1" LAG3 PD1+.

[0327] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+CTLA4' LAG3 PD1+.

[0328] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is KLRG1" CTLA4' LAG3 PD1+.

[0329] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" CTLA4’ LAG3'. In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGP CTLA4’ PD1+.

[0330] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRG1 LAG3 PD1+.

[0331] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+CTLA4’ LAG3 PD1+.

[0332] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+KLRGl" CTLA4’ LAG3 PD1+.

[0333] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is TIM3+KLRGl" CTLA4" LAG3 PD1+.

[0334] In certain embodiments, the cell comprises a CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the CAR is 19(T2)28zlXX. In certain embodiments, the cell is a T cell. In certain embodiments, the T cell is a CD4+T cell. In certain embodiments, the T cell is CD57+TIM3+KLRGl" CTLA4" LAG3" PD1+.

[0335] 3. Nucleic Acid Molecules, Vector and Genetic Modifications

[0336] The presently disclosed subject matter provides nucleic acid molecules encoding the presently disclosed CD19-targeted chimeric receptors (e.g., those disclosed in Section 2.2). In certain embodiments, the nucleic acid molecule further comprises a promoter that is operably linked to the presently disclosed CD19-targeted chimeric receptor. Also provided are cells comprising such nucleic acid molecules.

[0337] In certain embodiments, the promoter is endogenous or exogenous. In certain embodiments, the exogenous promoter is selected from the group consisting of an elongation factor (EF)-l promoter, a cytomegalovirus immediate-early promoter (CMV) promoter, a simian virus 40 early promoter (SV40) promoter, a phosphoglycerate kinase (PGK) promoter, a metallothionein promoter, and Ubiquitin C promoter. In certain embodiments, the endogenous promoter is selected from a TCR alpha promoter, a TCR beta promoter, and a beta 2-microglobulin promoter. In certain embodiments, the promoter is an inducible promoter. In certain embodiments, the inducible promoter is selected from the group consisting of a NF AT transcriptional response element (TRE) promoter, a CD69 promoter, a CD25 promoter, an IL-2 promoter, a 4- IBB promoter, a PDl promoter, and a LAG3 promoter.

[0338] The presently disclosed subject matter also provides vectors comprising the presently disclosed nucleic acid molecules. In certain embodiments, the vector is a viral vector. In certain embodiments, the viral vector is a retroviral vector. In certain embodiments, the retroviral vector is a gamma retroviral vector or lentiviral vector.

[0339] In certain embodiments, the vector comprises a nucleic acid molecule encoding a presently disclosed CD19-targeted chimeric receptor. In certain embodiments, the CD19-targeted chimeric receptor is a CD19-targeted CAR. In certain embodiments, the nucleic acid molecule encodes a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 45. In certain embodiments, the nucleic acid molecule comprises the nucleotide sequence set forth in SEQ ID NO: 46.

[0340] The nucleic acid molecules can be delivered into cells by art-known methods or as described herein. Genetic modification of a cell can be accomplished by transducing a substantially homogeneous cell composition with a recombinant DNA construct. In certain embodiments, a retroviral vector (e.g., gammaretroviral vector or lentiviral vector) is employed for the introduction of the DNA construct into the cell. For example, a polynucleotide encoding a presently disclosed chimeric receptor can be cloned into a retroviral vector and expression can be driven from its endogenous promoter, from the retroviral long terminal repeat, or from a promoter specific for a target cell type of interest. Non-viral vectors may be used as well.

[0341] For initial genetic modification of a cell to include the presently disclosed chimeric receptor, a retroviral vector can be employed for transduction, however, any other suitable viral vector or non-viral delivery system can be used. The chimeric receptor can be constructed in a single, multi ci str onic expression cassette, in multiple expression cassettes of a single vector, or in multiple vectors. Examples of elements that create polycistronic expression cassette include, but is not limited to, various viral and non-viral Internal Ribosome Entry Sites (IRES, e.g., FGF-1 IRES, FGF-2 IRES, VEGF IRES, IGF-II IRES, NF-KB IRES, RUNX1 IRES, p53 IRES, hepatitis A IRES, hepatitis C IRES, pestivirus IRES, aphthovirus IRES, picornavirus IRES, poliovirus IRES and encephalomyocarditis virus IRES) and cleavable linkers (e.g., 2A peptides, e.g., P2A, T2A, E2A and F2A peptides). An exemplary P2A peptide comprises or consists of the amino acid sequence set forth in SEQ ID NO: 47, which is provided below:

[0342] GSGATNFSLLKQAGDVEENPGP [ SEQ ID NO : 47 ]

[0343] An exemplary nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 47 is set forth in 48, which is provided below:

[0344] GGAAGCGGAGCTACTAACTTCAGCCTGCTGAAGCAGGCTGGAGACGTGGAGGAGAACCCTGGAC CC [ SEQ ID NO : 48 ]

[0345] Combinations of retroviral vectors and an appropriate packaging lines are also suitable, where the capsid proteins will be functional for infecting human cells. Various amphotropic virusproducing cell lines are known, including, but not limited to, PA12 (Miller etal., (V9 5)Mol Cell Biol (1985);5:431-437); PA317 (Miller., et al., Mol Cell Biol (1986); 6:2895-2902); and CRIP (Danos et al., Proc Natl Acad Sci USA (1988);85:6460-6464). Non-amphotropic particles are suitable too, e.g., particles pseudotyped with VSVG, RD114, or GALV envelope, and any other known in the art.

[0346] Possible methods of transduction also include direct co-culture of the cells with producer cells (Bregni et al., Blood (1992);80: 1418-1422), or culturing with viral supernatant alone or concentrated vector stocks with or without appropriate growth factors and polycations (Xu et al., Exp Hemat (1994); 22:223-230; and Hughes et al. J Clin Invest (1992); 89: 1817).

[0347] Other transducing viral vectors can be used to modify a cell. In certain embodiments, the chosen vector exhibits high efficiency of infection and stable integration and expression (see, e.g., Cayouette et al., Human Gene Therapy 8:423-430, 1997; Kido et al., Current Eye Research 15:833-844, 1996; Bloomer et al., Journal of Virology 71 :6641-6649, 1997; Naldini et al., Science 272:263-267, 1996; and Miyoshi et al., Proc. Natl. Acad. Sci. U.S.A. 94: 10319, 1997). Other viral vectors that can be used include, for example, adenoviral, lentiviral, and adena-associated viral vectors, vaccinia virus, a bovine papilloma virus, or a herpes virus, such as Epstein-Barr Virus (also see, for example, the vectors of Miller, Human Gene Thera (1990); 15- 14; Friedman, Science 244: 1275-1281, 1989; Eglitis et al., BioTechniques (1988);6:608-614; Tolstoshev et al., Cur Opin Biotechnol (1990); 1 :55-61; Sharp, The Lancet (1991);337: 1277-78; Cornetta et al., Nucleic Acid Research and Molecular Biology 36:311-22, 1987; Anderson, Science (1984);226:401-409; Moen, Blood Cells 17:407-16, 1991; Miller et al., Biotechnol (1989);7:980-90; LeGal La Salle et al., Science (1993);259:988-90; and Johnson, Chest (1995)107:77S- 83S). Retroviral vectors are particularly well developed and have been used in clinical settings (Rosenberg et al., N Engl J Afe7 (1990);323:370, 1990; Anderson et al., U.S. Patent. No. 5,399,346).

[0348] Non-viral approaches can also be employed for genetic modification of a cell. For example, a nucleic acid molecule can be introduced into a cell by administering the nucleic acid in the presence of lipofection (Feigner et al., Proc Natl Acad Sci U.S.A. (1987);84:7413; Ono et al., Neurosci Lett (I990);I7:259; Brigham et al., Am J Med Sci (1989);298:278; Staubinger et al., Methods in Enzymol (1983); 101 :512, Wu et al., J Biol Chem (1988);263: 14621; Wu et al., J Biol Chem (1989);264: 16985), or by micro-injection under surgical conditions (Wolff et al., Science (1990);247: 1465). Other non-viral means for gene transfer include transfection in vitro using calcium phosphate, DEAE dextran, electroporation, and protoplast fusion. Liposomes can also be potentially beneficial for delivery of DNA into a cell. Transplantation of normal genes into the affected tissues of a subject can also be accomplished by transferring a normal nucleic acid into a cultivatable cell type ex vivo (e.g., an autologous or heterologous primary cell or progeny thereof), after which the cell (or its descendants) are injected into a targeted tissue or are injected systemically. Recombinant receptors can also be derived or obtained using transposases or targeted nucleases (e.g. Zinc finger nucleases, meganucleases, or TAKEN, CRISPR). Transient expression may be obtained by RNA electroporation.

[0349] Any targeted genome editing methods can also be used to deliver the presently disclosed chimeric receptor (e.g., a presently disclosed CD19-targeted chimeric receptor) to a cell. In certain embodiments, a CRISPR system is used to deliver the presently disclosed chimeric receptor (e.g., a presently disclosed CD19-targeted chimeric receptor ). In certain embodiments, zinc-finger nucleases are used to deliver the presently disclosed chimeric receptor (e.g., a presently disclosed CD19-targeted chimeric receptor). In certain embodiments, a TAKEN system is used to deliver the presently disclosed chimeric receptor (e.g., a presently disclosed CD19-targeted chimeric receptor).

[0350] Clustered regularly-interspaced short palindromic repeats (CRISPR) system is a genome editing tool discovered in prokaryotic cells. When utilized for genome editing, the system includes Cas9 (a protein able to modify DNA utilizing crRNA as its guide), CRISPR RNA (crRNA, contains the RNA used by Cas9 to guide it to the correct section of host DNA along with a region that binds to tracrRNA (generally in a hairpin loop form) forming an active complex with Cas9), trans-activating crRNA (tracrRNA, binds to crRNA and forms an active complex with Cas9), and an optional section of DNA repair template (DNA that guides the cellular repair process allowing insertion of a specific DNA sequence). CRISPR / Cas9 often employs a plasmid to transfect the target cells. The crRNA needs to be designed for each application as this is the sequence that Cas9 uses to identify and directly bind to the target DNA in a cell. The repair template carrying a chimeric receptor expression cassette (e.g., a CAR expression cassette) needs also be designed for each application, as it must overlap with the sequences on either side of the cut and code for the insertion sequence. Multiple crRNA's and the tracrRNA can be packaged together to form a single-guide RNA (sgRNA). This sgRNA can be joined together with the Cas9 gene and made into a plasmid in order to be transfected into cells.

[0351] A zinc-finger nuclease (ZFN) is an artificial restriction enzyme, which is generated by combining a zinc finger DNA-binding domain with a DNA-cleavage domain. A zinc finger domain can be engineered to target specific DNA sequences which allows a zinc-finger nuclease to target desired sequences within genomes. The DNA-binding domains of individual ZFNs typically contain a plurality of individual zinc finger repeats and can each recognize a plurality of base pairs. The most common method to generate new zinc-finger domain is to combine smaller zinc-finger "modules" of known specificity. The most common cleavage domain in ZFNs is the non-specific cleavage domain from the type Ils restriction endonuclease Fokl. Using the endogenous homologous recombination (HR) machinery and a homologous DNA template carrying a chimeric receptor expression cassette (e.g., a CAR expression cassette), ZFNs can be used to insert the CAR expression cassette into the genome. When the targeted sequence is cleaved by ZFNs, the HR machinery searches for homology between the damaged chromosome and the homologous DNA template, and then copies the sequence of the template between the two broken ends of the chromosome, whereby the homologous DNA template is integrated into the genome.

[0352] Transcription activator-like effector nucleases (TALEN) are restriction enzymes that can be engineered to cut specific sequences of DNA. TALEN system operates on almost the same principle as ZFNs. They are generated by combining a transcription activator-like effectors DNA- binding domain with a DNA cleavage domain. Transcription activator-like effectors (TALEs) are composed of 33-34 amino acid repeating motifs with two variable positions that have a strong recognition for specific nucleotides. By assembling arrays of these TALEs, the TALE DNA- binding domain can be engineered to bind desired DNA sequence, and thereby guide the nuclease to cut at specific locations in genome. cDNA expression for use in polynucleotide therapy methods can be directed from any suitable promoter (e.g., the human cytomegalovirus (CMV), simian virus 40 (SV40), metallothionein promoters, or Ubiquitin C promoter), and regulated by any appropriate mammalian regulatory element or intron (e.g. the elongation factor la enhancer / promoter / intron structure). For example, if desired, enhancers known to preferentially direct gene expression in specific cell types can be used to direct the expression of a nucleic acid. The enhancers used can include, without limitation, those that are characterized as tissue- or cell-specific enhancers. Alternatively, if a genomic clone is used as a therapeutic construct, regulation can be mediated by the cognate regulatory sequences or, if desired, by regulatory sequences derived from a heterologous source, including any of the promoters or regulatory elements described above.

[0353] Methods for delivering the genome editing agents / sy stems can vary depending on the need. In certain embodiments, the components of a selected genome editing method are delivered as DNA constructs in one or more plasmids. In certain embodiments, the components are delivered via viral vectors. Common delivery methods include but is not limited to, electroporation, microinjection, gene gun, impalefection, hydrostatic pressure, continuous infusion, sonication, magnetofection, adeno-associated viruses, envelope protein pseudotyping of viral vectors, replication-competent vectors cis and trans-acting elements, herpes simplex virus, and chemical vehicles (e.g., oligonucleotides, lipoplexes, polymersomes, polyplexes, dendrimers, inorganic Nanoparticles, and cell-penetrating peptides).

[0354] In certain embodiments, the delivery methods include use of colloids. As used herein, the term “colloid” refers to systems in which there are two or more phases, with one phase (e.g., the dispersed phase) distributed in the other phase (e.g., the continuous phase). Moreover, at least one of the phases has small dimensions (in the range of about 109to about 106m). Non-limiting examples of colloids encompassed by the presently disclosed subject matter include macromolecule complexes, nanocapsules, microspheres, beads, and lipid-based systems (e.g., micelles, liposomes, and lipid nanoparticles).

[0355] In certain embodiments, the delivery methods include use of liposomes. The term “liposome,” as used herein, refers to single- or multi-layered spherical lipid bilayer structures produced from lipids dissolved in organic solvents and then dispersed in aqueous media. Experimentally and therapeutically used for delivering an active pharmaceutical ingredient (e.g., nucleic acid compositions disclosed herein) to cells, liposomes fuse with cell membranes so the contents are transferred into the cytoplasm.

[0356] In certain embodiments, the delivery methods include use of lipid nanoparticles. As used herein, the term “lipid nanoparticle” refers to a particle having at least one dimension in the order of nanometers (e.g., from about 1 nm to about 1,000 nm) and including at least one lipid. In certain embodiments, the lipid nanoparticles can include an active pharmaceutical ingredient (e.g., nucleic acid compositions disclosed herein) for delivering to cells. The morphology of the lipid nanoparticles can be different from liposomes. While liposomes are characterized by a lipid bilayer surrounding a hydrophilic core, lipid nanoparticles have an electron-dense core where cationic lipids and / or ionizable lipids are organized into inverted micelles around an active pharmaceutical ingredient (e.g., nucleic acid compositions disclosed herein). Additional information on the morphology and properties of lipid nanoparticles and liposomes can be found in Wilczewska, et al., Pharmacological reports 64, no. 5 (2012): 1020-1037; Eygeris et al., Accounts of Chemical Research 55, no. 1 (2021): 2-12; Zhang et al., Chemical Reviews 121, no. 20 (2021): 12181-12277; and Fan et al., Journal of pharmaceutical and biomedical analysis 192 (2021): 113642.

[0357] In certain embodiments, the lipid nanoparticles have a mean diameter of from about 30 nm to about 150 nm, from about 40 nm to about 150 nm, from about 50 nm to about 150 nm, from about 60 nm to about 130 nm, from about 70 nm to about 110 nm, from about 70 nm to about 100 nm, from about 80 nm to about 100 nm, from about 90 nm to about 100 nm, from about 70 to about 90 nm, from about 80 nm to about 90 nm, from about 70 nm to about 80 nm, or about 30 nm, 35 nm, 40 nm, 45 nm, 50 nm, 55 nm, 60 nm, 65 nm, 70 nm, 75 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, 120 nm, 125 nm, 130 nm, 135 nm, 140 nm, 145 nm, or 150 nm.

[0358] In certain embodiments, the lipid nanoparticles can include a cationic lipid or an ionizable lipid. The term “cationic lipid” refers to lipids including a head group with permanent positive charges. Non-limiting examples of cationic lipids encompassed by the presently disclosed subject matter include l,2-di-O-octadecenyl-3 -trimethylammonium -propane (DOTMA), l,2-dioleoyl-3- trimethyl ammonium -propane (DOTAP), 2, 3-di oleyloxy -N-[2-(sperminecarboxamido)ethyl]- N,N-dimethyl-l-propanaminium trifluoroacetate (DOSPA), and ethylphosphatidylcholine (ePC).

[0359] As used herein, the term “ionizable lipid” refers to lipids that are protonated at low pH and are neutral at physiological pH. The pH-sensitivity of ionizable lipids is particularly beneficial for delivery in vivo (e.g., delivery of nucleic acid compositions disclosed herein), because neutral lipids have less interactions with the anionic membranes of blood cells and, thus, improve the biocompatibility of the lipid nanoparticles. Once trapped in endosomes, ionizable lipids are protonated and promote membrane destabilization to allow the endosomal escape of the nanoparticles. Non-limiting example of ionizable lipids encompassed by the presently disclosed subject matter include tetrakis(8-methylnonyl) 3,3',3",3"'-(((methylazanediyl) bis(propane-3,l diyl))bis (azanetriyl))tetrapropionate; decyl (2-(dioctylammonio)ethyl) phosphate; ((4- hydroxybutyl)azanediyl)bis(hexane-6,l-diyl)bis(2 -hexyldecanoate); bis(2-

[0360] (dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6- diazahexacosyl)azanediyl)dipropionate; l,l'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl) (2-hydroxydodecyl)amino)ethyl) piperazin- l-yl)ethyl)azanediyl) bis(dodecan-2-ol); cKK-E12, 3, 6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2, 5-dione; (6Z,9Z,28Z,31Z)- heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino) butanoate; hexa(octan-3-yl) 9, 9', 9", 9"', 9'"', 9"'"- ((((benzene-l,3,5-tricarbonyl)yris(azanediyl)) tris (propane-3,1 -diyl)) tris(azanetriyl))hexanonanoate; heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6- (undecyloxy)hexyl)amino) octanoate; and (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4, 1- diyl))bis(azanetriyl))tetrakis(ethane-2, l-diyl) (9Z,9'Z,9"Z,9"'Z, 12Z, 12'Z,12"Z,12"'Z)-tetrakis

[0361] (octadeca-9, 12-dienoate).

[0362] Additionally, in certain embodiments, the lipid nanoparticles can include other lipids. For example, but without any limitation, the lipid nanoparticles of the presently disclosed subject matter can include phospholipids, cholesterol, polyethylene glycol (PEG)-functionalized lipids

[0363] (PEG-lipids). These lipids can improve certain properties of the lipid nanoparticles (e.g., stability, biodistribution, etc.). For example, cholesterol enhances the stability of the lipid nanoparticles by modulating the integrity and rigidity. Non-limiting examples of other lipids present in lipid nanoparticles include cholesterol, DC-cholesterol, P-sitosterol, BHEM-cholesterol, ALC-0159, di stearoylphosphatidylcholine (DSPC), dioleoylphosphatidylcholine (DOPC), dipalmitoylphosphatidylcholine (DPPC), dioleoylphosphatidylglycerol (DOPG), dipalmitoylphosphatidylglycerol (DPPG), di ol eoy Iphosphati dy 1 ethanol amine (DOPE), palmitoyloleoylphosphatidylcholine (POPC), palmitoyloleoyl-phosphatidylethanolamine (POPE) and dioleoyl-phosphatidylethanolamine 4-(N- maleimidom ethyl) -cyclohexane -1 -carboxylate (DOPE-mal), dipalmitoyl phosphatidyl ethanolamine (DPPE), dimyristoylphosphoethanolamine (DMPE), distearoylphosphatidylethanolamine (DSPE), 16-0-monom ethyl PE, 16-O-dimethyl PE, 18-1 -trans PE, 1- stearioyl-2-oleoyl-phosphatidy ethanol amine (SOPE), and 1,2-dielaidoyl-sn- glycero-3- phophoethanolamine (transDOPE).

[0364] In certain embodiments, the lipid nanoparticles can include a targeting moiety that binds to a ligand. The use of the targeting moieties allows selective delivery of an active pharmaceutical ingredient (e.g., nucleic acid compositions disclosed herein) to target cells expressing the ligand (e.g., T cells). In certain embodiments, the targeting moiety can be an antibody or antigen-binding fragment thereof that binds to a cell surface receptor. For example, but without any limitation, the targeting domain is an antibody or antigen-binding fragment thereof that binds to a receptor expressed on the surface of a T cell (e.g., CD3, CD4, CD8, CD16, CD40L, CD95, FasL, CTLA- 4, 0X40, GITR, LAG3, ICOS and PD-1).

[0365] In certain embodiments, the delivery methods are in vivo delivery methods. In certain embodiments, the delivery methods are ex vivo delivery methods.

[0366] 4. Formulations and Administration

[0367] The presently disclosed subject matter also provides compositions comprising the presently disclosed cells (e.g., those disclosed in Section 2). For example, but without any limitation, the cells of the presently disclosed compositions comprise a chimeric receptor (e.g., those disclosed in Section 2.2). In certain embodiments, the composition is a pharmaceutical composition that further comprises a pharmaceutically acceptable carrier.

[0368] In certain embodiments, the presently disclosed compositions comprise cells (e.g., those disclosed in Section 2) that have been selected for the presence or absence of certain biomarkers. In certain embodiments, the biomarkers are selected from CD57, TIM3, KLRG1, CTLA4, LAG3, PD1, or any combination thereof.

[0369] In certain embodiments, the presently disclosed compositions comprise cells (e.g., those disclosed in Section 2) that have been selected for the expression level of certain biomarkers compared to the expression level of a reference sample. In certain embodiments, the compositions comprise cells having an expression level of CD57, TIM3, and / or PD1 that is increased compared to the expression level of a reference sample. In certain embodiments, the compositions comprise cells having an expression level of KLRG1, CTLA4, and / or LAG3 that is reduced compared to the expression level of a reference sample.

[0370] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0371] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+cells (e.g., T cells).

[0372] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0373] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% TIM3+cells (e.g.,

[0374] T cells). In certain embodiments, the composition comprises at least about 70% TIM3+cells (e.g.,

[0375] T cells). In certain embodiments, the composition comprises at least about 75% TIM3+cells (e.g.,

[0376] T cells). In certain embodiments, the composition comprises at least about 80% TIM3+cells (e.g.,

[0377] T cells). In certain embodiments, the composition comprises at least about 85% TIM3+cells (e.g.,

[0378] T cells). In certain embodiments, the composition comprises at least about 90% TIM3+cells (e.g.,

[0379] T cells). In certain embodiments, the composition comprises at least about 95% TIM3+cells (e.g.,

[0380] T cells). In certain embodiments, the composition comprises about 100% TIM3+cells (e.g., T cells).

[0381] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises about 100% KLRGr cells (e.g., T cells).

[0382] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CTLA4' cells (e.g., T cells).

[0383] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0384] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% LAG3' cells (e.g., T cells).

[0385] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0386] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% PD1+cells (e.g.,

[0387] T cells). In certain embodiments, the composition comprises at least about 15% PD1+cells (e.g.,

[0388] T cells). In certain embodiments, the composition comprises at least about 20% PD1+cells (e.g.,

[0389] T cells). In certain embodiments, the composition comprises at least about 25% PD1+cells (e.g.,

[0390] T cells). In certain embodiments, the composition comprises at least about 30% PD1+cells (e.g.,

[0391] T cells). In certain embodiments, the composition comprises at least about 35% PD1+cells (e.g.,

[0392] T cells). In certain embodiments, the composition comprises at least about 40% PD1+cells (e.g.,

[0393] T cells). In certain embodiments, the composition comprises at least about 45% PD1+cells (e.g.,

[0394] T cells). In certain embodiments, the composition comprises at least about 50% PD1+cells (e.g.,

[0395] T cells). In certain embodiments, the composition comprises at least about 55% PD1+cells (e.g.,

[0396] T cells). In certain embodiments, the composition comprises at least about 60% PD1+cells (e.g.,

[0397] T cells). In certain embodiments, the composition comprises at least about 65% PD1+cells (e.g.,

[0398] T cells). In certain embodiments, the composition comprises at least about 70% PD1+cells (e.g.,

[0399] T cells). In certain embodiments, the composition comprises at least about 75% PD1+cells (e.g.,

[0400] T cells). In certain embodiments, the composition comprises at least about 80% PD1+cells (e.g.,

[0401] T cells). In certain embodiments, the composition comprises at least about 85% PD1+cells (e.g.,

[0402] T cells). In certain embodiments, the composition comprises at least about 90% PD1+cells (e.g.,

[0403] T cells). In certain embodiments, the composition comprises at least about 95% PD1+cells (e.g.,

[0404] T cells). In certain embodiments, the composition comprises about 100% PD1+cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+TIM3+cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0405] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+KLRGr KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+KLRGl" cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+CTLA4' CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0406] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+LAG3' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0407] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0408] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+KLRGL KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises about 100% TIM3+KLRGL cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0409] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+CTLA4' CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0410] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+LAG3' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0411] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% KLRG1" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% KLRG1" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% KLRGL CTLA4' CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% KLRGL CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises about 100% KLRGl" CTLA4" cells (e.g., T cells).

[0412] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0413] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% KLRGr LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% KLRGr LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% KLRGP LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% KLRGP LAG3' cells (e.g., T cells).

[0414] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% KLRG1" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% KLRG1" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% KLRGP PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% KLRG1" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% KLRGP PD1+cells (e.g., T cells).

[0415] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0416] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CTLA4' LAG3' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CTLA4' LAG3' cells (e.g., T cells).

[0417] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CTLA4' PD1+cells (e.g., T cells).

[0418] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0419] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% LAG3- PD1+cells (e.g., T cells).

[0420] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+TIM3+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+TIM3+KLRG1" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+TIM3+KLRGr cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+TIM3+KLRGr cells (e.g., T cells).

[0421] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+ TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+TIM3+CTLA4‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+TIM3+CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+TIM3+CTLA4- cells (e.g., T cells).

[0422] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+TIM3+LAG3‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+TIM3+LAG3‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+TIM3+LAG3‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+TIM3+LAG3‘ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+TIM3+LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+TIM3+LAG3- cells (e g., T cells).

[0423] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises atleast about 55% CD57+TIM3+PD1+ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+TIM3+PD1+' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+TIM3+PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+TIM3+PD1+- cells (e.g., T cells).

[0424] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+KLRG1' CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+KLRG1' CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+KLRG1" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+KLRGl" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+KLRGr CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+KLRGr CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+KLRGr CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+KLRGl" CTLA4" cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+KLRGl" CTLA4" cells (e.g., T cells).

[0425] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+KLRGl" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+KLRG1" LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+KLRG1" LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+KLRGI’ LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+KLRG1" LAG3’ cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+KLRGI’ LAG3’ cells (e.g., T cells).

[0426] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+KLRG1" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+KLRG1" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+KLRG r PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+KLRGI’ PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+KLRGI’ PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+KLRGr PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+KLRGl" PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+KLRGl" PD1+cells (e.g., T cells).

[0427] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0428] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+CTLA4" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+CTLA4" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+CTLA4" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+CTLA4" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+CTLA4" LAG3" cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+ CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+CTLA4' LAG3' cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+CTLA4’ LAG3' cells (e.g., T cells).

[0429] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+CTLA4' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+CTLA4- PD1+cells (e.g., T cells).

[0430] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about

[0431] 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 50% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 55% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 60% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 65% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 70% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 75% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 80% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 85% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 90% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises at least about 95% CD57+LAG3' PD1+cells (e.g., T cells). In certain embodiments, the composition comprises about 100% CD57+LAG3' PD1+cells (e.g., T cells).

[0432] In certain embodiments, the composition comprises from about 5% to about 100%, from about 10% to about 100%, from about 20% to about 100%, from about 30% to about 100%, from about 40% to about 100%, from about 50% to about 100%, from about 60% to about 100%, from about 70% to about 100%, from about 80% to about 100%, from about 90% to about 100%, from about 5% to about 10%, from about 5% to about 15%, from about 5% to about 25%, from about 5% to about 35%, from about 5% to about 45%, from about 5% to about 55%, from about 5% to about 65%, from about 5% to about 75%, from about 5% to about 85%, from about 5% to about 95% TIM3+KLRG1" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% TIM3+KLRG1" CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 5% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 10% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 15% TIM3+KLRGL CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 20% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 25% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 30% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 35% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 40% TIM3+KLRGr CTLA4' cells (e.g., T cells). In certain embodiments, the composition comprises at least about 45% TIM3+KLRGl" CTLA4' cells (e.g., T cells). In certain embodiments, the composition ...

Claims

WHAT IS CLAIMED IS:

1. A method comprising: a) measuring an expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; and c) identifying the subject as responsive or as having an increased likelihood to respond to an adoptive cell therapy.

2. The method of claim 1, wherein the subject is responsive or has an increased likelihood to respond to the adoptive cell therapy if a) the expression level of CD57 is increased compared to the reference sample; b) the expression level of TIM3 is increased compared to the reference sample; c) the expression level of KLRG1 is reduced compared to the reference sample; d) the expression level of CTLA4 is reduced compared to the reference sample; e) the expression level of LAG3 is reduced compared to the reference sample; and / or f) the expression level of PD1 is increased compared to the reference sample.

3. A method compri sing : a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; and c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy.

4. The method of claim 3, wherein the subject is non-responsive or has a reduced likelihood to respond the adoptive cell therapy if a) the expression level of CD57 is reduced compared to the reference sample; b) the expression level of TIM3 is reduced compared to the reference sample; c) the expression level of KLRG1 is increased compared to the reference sample; d) the expression level of CTLA4 is increased compared to the reference sample; e) the expression level of LAG3 is increased compared to the reference sample; and / or f) the expression level of PD1 is reduced compared to the reference sample.

5. The method of any one of claims 1-4, wherein the adoptive cell therapy comprises a cell comprising a CD19-targeted chimeric receptor.

6. The method of claim 5 further comprising administering to the subject a cell comprising a CD19-targeted chimeric receptor or a composition comprising thereof, wherein the CD 19- targeted chimeric receptor comprises an extracellular antigen-binding domain comprising a heavy chain variable region (VH) comprising a CDR1 comprising the amino acid sequence setforth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12, and a light chain variable region (VL) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15.

7. The method of claim 6, wherein the extracellular antigen-binding domain comprises a scFv.

8. The method of claim 6 , wherein a) the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 16; and b) the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 17.

9. The method of claim 8, wherein a) the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 16; and b) the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 17.

10. The method of claim 6, wherein the chimeric receptor comprises a transmembrane domain comprising a CD28 polypeptide.

11. The method of claim 6, wherein the chimeric receptor comprises an intracellular signaling domain comprising a CD3(^ polypeptide comprising a native ITAM1, an ITAM2 variant consisting of two loss-of-function mutations, and an ITAM3 consisting of two loss-of-function mutations.

12. The method of claim 11, wherein the native ITAM1 consists of the amino acid sequence set forth in SEQ ID NO: 28, the ITAM2 variant consists of the amino acid sequence set forth in SEQ ID NO: 34, and the ITAM3 variant consists of the amino acid sequence set forth in SEQ ID NO: 38.

13. The method of claim 11 , wherein the chimeric receptor comprises an intracellular signaling domain comprising a CD3(^ polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 40.

14. The method of claim 11, wherein the intracellular signaling domain further comprises a CD28 polypeptide.

15. The method of claim 6, wherein the chimeric receptor comprises the amino acid sequence set forth in SEQ ID NO: 45.

16. The method of claim 6, wherein the chimeric receptor is expressed from a vector.

17. The method of claim 16, wherein the vector is a retroviral vector.

18. The method of claim 6, wherein the chimeric receptor is constitutively expressed on the surface of the cell.

19. The method of claim 6, wherein the cell comprising the CD19-targeted chimeric receptor is an immunoresponsive cell.

20. The method of claim 19, wherein the cell comprising the CD19-targeted chimeric receptor is a cell of the lymphoid lineage or a cell of the myeloid lineage.

21. The method of claim 19, wherein the cell comprising the CD19-targeted chimeric receptor is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a stem cell from which lymphoid cells may be differentiated, and a stem cell from which myeloid cells may be differentiated.

22. The method of claim 21, wherein the cell comprising the CD19-targeted chimeric receptor is a T cell.

23. The method of claim 22, wherein the T cell is selected from the group consisting of helper T cells, cytotoxic T cells, memory T cells, regulatory T cells, tumor-infiltrating lymphocyte (TIL), Natural Killer T cells, mucosal associated invariant T cells, and y5 T cells.

24. The method of claim 21, wherein the cell comprising the CD19-targeted chimeric receptor is a NK cell.

25. The method of claim 21, wherein the cell comprising the CD19-targeted chimeric receptor is derived from a stem cell.

26. The method of claim 25, wherein the stem cell is a pluripotent stem cell.

27. The method of claim 26, wherein the pluripotent stem cell is an embryoid stem cell or an induced pluripotent stem cell.

28. The method of claim 6, wherein the cell comprising the CD19-targeted chimeric receptor is selected from a CD57+T cell, a TIM3+T cell, a KLRGT T cell, a CTLA4' T cell, a LAG3' T cell, a PD1+T cell, a CD57+TIM3+T cell, a CD57+KLRGT T cell, a CD57+CTLA4' T cell, a CD57+LAG3’ T cell, a CD57+PD1+T cell, a TIM3+KLRGT T cell, a TIM3+CTLA4' T cell, a TIM3+LAG3' T cell, a TIM3+PD1+T cell, a KLRGT CTLA4' T cell, a KLRGT LAG3' T cell, a KLRGT PD1+T cell, a CTLA4' LAG3' T cell, a CTLA4' PD1+T cell, a LAG3' PD1+T cell, a CD57+TIM3+KLRGT T cell, a CD57+TIM3+CTLA4' T cell, a CD57+TIM3+LAG3’ T cell, a CD57+TIM3+PD1+T cell, a CD57+KLRGT CTLA4' T cell, a CD57+KLRGT LAG3’ T cell, a CD57+KLRGT PD1+T cell, a CD57+CTLA4' LAG3' T cell, a CD57+CTLA4’ PD1+T cell, a CD57+LAG3' PD1+T cell, a TIM3+KLRGT CTLA4' T cell, a TIM3+KLRGT LAG3' T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+KLRGT PD1+T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+CTLA4' PD1+T cell, a TIM3+LAG3' PD1+T cell, a KLRGT CTLA4' LAG3' T cell, a KLRGT CTLA4' PD1+T cell, a KLRGT LAG3' PD1+T cell, a CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' T cell, a CD57+TIM3+KLRGT LAG3' T cell, a CD57+TIM3+KLRGT PD1+T cell, a CD57+TIM3+CTLA4'LAG3" T cell, a CD57+TIM3+CTLA4" PD1+T cell, a CD57+TIM3+LAG3" PD1+T cell, a CD57+KLRGl" CTLA4" LAG3" T cell, a CD57+KLRGl" CTLA4" PD1+T cell, a CD57+KLRGl" LAG3" PD1+T cell, a CD57+CTLA4" LAG3" PD1+T cell, a TIM3+KLRGl" CTLA4" LAG3" T cell, a TIM3+KLRGl" CTLA4" LAG3" T cell, a TIM3+KLRGl" CTLA4" PD1+T cell, a TIM3+KLRGl" LAG3" PD1+T cell, a TIM3+CTLA4" LAG3" PD1+T cell, a KLRGl" CTLA4" LAG3" PD1+T cell, a CD57+TIM3+KLRGl" CTLA4" LAG3" T cell, a CD57+TIM3+KLRGl" CTLA4" PD1+T cell, a CD57+TIM3+KLRGl" LAG3" PD1+T cell, a CD57+TIM3+CTLA4" LAG3" PD1+T cell, a CD57+KLRGl" CTLA4" LAG3" PD1+T cell, a TIM3+KLRGl" CTLA4" LAG3" PD1+T cell, a CD57+TIM3+KLRGl" CTLA4" LAG3" PD1+T cell, or a combination thereof.

29. The method of claim 6, wherein the composition comprises at least about 5% of a CD57+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGl" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGl" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGl" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGl" CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGl" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGl" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4" LAG3" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGl" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4" T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3" T cell comprising the CD19-targeted chimeric receptor, at least aboutabout 5% of a CD57+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP PD1+T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4 LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRG1 ’ CTLA4' LAG3' T cell comprising the CD 19- targeted chimeric receptor, at least about 5% of a KLRGP CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP LAG3' PD1+T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' PD1+T cell comprising the CD19-targetedchimeric receptor, at least about 5% of a TIM3+KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGL CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, or a combination thereof.

30. The method of claim 6, wherein the subject has a tumor and / or a neoplasm associated with CD 19.

31. The method of claim 30, wherein the tumor and / or neoplasm is a blood cancer.

32. The method of claim 31, wherein the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas.

33. The method of claim 32, wherein the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed- phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia.

34. The method of claim 33, wherein the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma.

35. The method of claim 31, wherein the tumor and / or neoplasm is a B cell malignancy.

36. The method of claim 35, wherein the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma.

37. The method of claim 31, wherein the tumor and / or neoplasm is B cell lymphoma.

38. The method of claim 37, wherein the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma.

39. The method of claim 5, wherein the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 is measured on the surface of the immunoresponsive cell.

40. The method of claim 5, wherein the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 is measured by cytometry or by flow-cytometry.

41. The method of claim 39 or 40, wherein the immunoresponsive cell is a T cell.

42. The method of claim 41, wherein T cell is selected from the group consisting of a helper T cell, a cytotoxic T cell, a memory T cell, a central memory T cell, a stem-like memory T cell, an effector memory T cell, a regulatory T cell, a tumor-infiltrating lymphocyte (TIL), a Natural killer T cells, a y5 T cell, a CD4+T cells, and a CD8+T cell.

43. The method of claim 42, wherein the T cell is a CD4+T cell or a CD8+T cell.

44. The method of claim 39 or 40, wherein the immunoresponsive cell is obtained from a sample of the subject.

45. The method of claim 44, wherein the sample is selected from bone marrow, thymus, tumor biopsy, tumor fragments, enzymatically digested tumor tissue, dissociated / suspended cells, a lymph node sample, or a bodily fluid sample46. The method of claim 45, wherein the bodily fluid sample is a blood sample, an ascites sample, or a lymph sample.

47. The method of claim 5, wherein the subject is a human subject.

48. A composition comprising a cell comprising a CD19-targeted chimeric receptor, wherein the CD19-targeted chimeric receptor comprises an extracellular antigen-binding domain, a transmembrane domain, and an intracellular domain, wherein the extracellular antigenbinding domain comprises a heavy chain variable region (VH) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 10, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 11, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 12, and a light chain variable region (VL) comprising a CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 13, a CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 14, and a CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 15.

49. The composition of claim 48, wherein the extracellular antigen-binding domain comprises a scFv.

50. The composition of claim 48, wherein a) the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 16; and b) the heavy chain variable region comprises an amino acid sequence that is at least about 80% identical to the amino acid sequence set forth in SEQ ID NO: 17.

51. The composition of claim 50, wherein a) the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 16; and b) the heavy chain variable region comprises the amino acid sequence set forth in SEQ ID NO: 17.

52. The composition of claim 48, wherein the transmembrane domain comprises a CD28 polypeptide.

53. The composition of claim 48, wherein the intracellular signaling domain comprises a CD3(^ polypeptide comprising a native IT AMI, an ITAM2 variant consisting of two loss-of-function mutations, and an ITAM3 consisting of two loss-of-function mutations.

54. The composition of claim 53, wherein the native ITAM1 consists of the amino acid sequence set forth in SEQ ID NO: 28, the ITAM2 variant consists of the amino acid sequence set forth in SEQ ID NO: 34, and the ITAM3 variant consists of the amino acid sequence set forth in SEQ ID NO: 38.

55. The composition of claim 53, wherein the intracellular signaling domain comprises a CD3(^ polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 40.

56. The composition of claim 53, wherein the intracellular signaling domain further comprises a CD28 polypeptide.

57. The composition of claim 48, wherein the chimeric receptor comprises the amino acid sequence set forth in SEQ ID NO: 45.

58. The composition of claim 48, wherein the chimeric receptor is expressed from a vector.

59. The composition of claim 58, wherein the vector is a retroviral vector.

60. The composition of claim 48, wherein the chimeric receptor is constitutively expressed on the surface of the cell.

61. The composition of claim 48, wherein the cell is an immunoresponsive cell.

62. The composition of claim 61, wherein the cell is a cell of the lymphoid lineage or a cell of the myeloid lineage.

63. The composition of claim 61, wherein the cell is selected from the group consisting of a T cell, a Natural Killer (NK) cell, a stem cell from which lymphoid cells may be differentiated, and a stem cell from which myeloid cells may be differentiated.

64. The composition of claim 63, wherein the cell is a T cell.

65. The composition of claim 64, wherein the T cell is selected from the group consisting of helper T cells, cytotoxic T cells, memory T cells, regulatory T cells, tumor-infiltrating lymphocyte (TIL), Natural Killer T cells, mucosal associated invariant T cells, and y5 T cells.

66. The composition of claim 63, wherein the cell is an NK cell.

67. The composition of claim 63, wherein the cell is derived from a stem cell.

68. The composition of claim 67, wherein the stem cell is a pluripotent stem cell.

69. The composition of claim 68, wherein the pluripotent stem cell is an embryoid stem cell or an induced pluripotent stem cell.

70. The composition of claim 48, wherein the cell is selected from a CD57+T cell, a TIM3+T cell, a KLRGT T cell, a CTLA4' T cell, a LAG3' T cell, a PD1+T cell, a CD57+TIM3+T cell, a CD57+KLRGT T cell, a CD57+CTLA4' T cell, a CD57+LAG3' T cell, a CD57+PD1+T cell, a TIM3+KLRGT T cell, a TIM3+CTLA4' T cell, a TIM3+LAG3' T cell, a TIM3+PD1+T cell, a KLRGT CTLA4' T cell, a KLRGT LAG3' T cell, a KLRGT PD1+T cell, a CTLA4' LAG3’ T cell, a CTLA4' PD1+T cell, a LAG3' PD1+T cell, a CD57+TIM3+KLRGT T cell, a CD57+TIM3+CTLA4' T cell, a CD57+TIM3+LAG3' T cell, a CD57+TIM3+PD1+T cell, a CD57+KLRGT CTLA4' T cell, a CD57+KLRGT LAG3' T cell, a CD57+KLRGT PD1+T cell, a CD57+CTLA4' LAG3' T cell, a CD57+CTLA4' PD1+T cell, a CD57+LAG3' PD1+T cell, a TIM3+KLRGT CTLA4' T cell, a TIM3+KLRGT LAG3' T cell, a TIM3+CTLA4' LAG3’ T cell, a TIM3+KLRGT PD1+T cell, a TIM3+CTLA4' LAG3' T cell, a TIM3+CTLA4’ PD1+T cell, a TIM3+LAG3' PD1+T cell, a KLRGT CTLA4' LAG3' T cell, a KLRGT CTLA4’ PD1+T cell, a KLRGT LAG3' PD1+T cell, a CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' T cell, a CD57+TIM3+KLRGT LAG3' T cell, a CD57+TIM3+KLRGT PD1+T cell, a CD57+TIM3+CTLA4' LAG3' T cell, a CD57+TIM3+CTLA4' PD1+T cell, a CD57+TIM3+LAG3' PD1+T cell, a CD57+KLRGT CTLA4' LAG3' T cell, a CD57+KLRGT CTLA4' PD1+T cell, a CD57+KLRGT LAG3' PD1+T cell, a CD57+CTLA4' LAG3’ PD1+T cell, a TIM3+KLRGT CTLA4' LAG3' T cell, a TIM3+KLRGT CTLA4' LAG3’ T cell, a TIM3+KLRGT CTLA4' PD1+T cell, a TIM3+KLRGT LAG3' PD1+T cell, a TIM3+CTLA4’ LAG3' PD1+T cell, a KLRGT CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4’ LAG3' T cell, a CD57+TIM3+KLRGT CTLA4' PD1+T cell, a CD57+TIM3+KLRGT LAG3' PD1+T cell, a CD57+TIM3+CTLA4' LAG3' PD1+T cell, a CD57+KLRGT CTLA4’ LAG3' PD1+T cell, a TIM3+KLRGT CTLA4' LAG3' PD1+T cell, a CD57+TIM3+KLRGT CTLA4' LAG3' PD1+T cell, or a combination thereof.

71. The composition of claim 48, comprising at least about 5% of a CD57+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+T cell comprising the CD 19- targeted chimeric receptor, at least about 5% of a KLRGT T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGT T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+PD1+T cell comprising the CD19-targeted chimeric receptor, atleast about 5% of a TIM3+KLRGP T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' T cell comprising the CD 19- targeted chimeric receptor, at least about 5% of a CD57+TIM3+PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP CTLA4' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGP LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a TIM3+KLRGP PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGP LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP CTLA4' T cell comprising the CD 19- targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGP LAG3' T cellcomprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+LAG3' PD1+T cell comprising the CD 19- targeted chimeric receptor, at least about 5% of a CD57+KLRG1" CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRGT CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG1" LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGT CTLA4' LAG3' T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a TIM3+KLRGT CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGT CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRG1" LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a KLRGT CTLA4' LAG3' PD1+T cell comprising the CD19- targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGT CTLA4' LAG3' T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRG1" CTLA4' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGT LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+KLRG1 ’ CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a TIM3+KLRGT CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, at least about 5% of a CD57+TIM3+KLRGT CTLA4' LAG3' PD1+T cell comprising the CD19-targeted chimeric receptor, or a combination thereof.

72. The composition of claim 48, which is a pharmaceutical composition further comprising a pharmaceutically acceptable carrier.

73. The composition of claim 48, comprising between about 1 x 106and about 5 * 108cells.

74. The composition of claim 48, comprising between about 1 x 106and about 1 x 108cells.

75. The composition of claim 48, comprising between about 1 x 106and about 5 x 107cells.

76. The composition of claim 48, comprising about 2.5 x 107cells.

77. A method of reducing tumor burden in a subject, comprising administering to the subject a composition of claim 48.

78. The method of claim 77, wherein the method reduces the number of tumor cells, reduces tumor size, and / or eradicates the tumor in the subject.

79. A method of increasing or lengthening survival of a subject having a neoplasm, comprising administering to the subject a composition of claim 48.

80. A method of treating and / or preventing a neoplasm in a subject, comprising administering to the subject a composition of claim 48.

81. The method of any one of claims 77-80, wherein the tumor and / or neoplasm is associated with CD 19.

82. The method of any one of claims 77-80, wherein the tumor and / or neoplasm is a blood cancer.

83. The method of claim 82, wherein the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas.

84. The method of claim 83, wherein the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed- phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia.

85. The method of claim 84, wherein the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma.

86. The method of any one of claims 77-80, wherein the tumor and / or neoplasm is a B cell malignancy.

87. The method of claim 86, wherein the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma.

88. The method of any one of claims 77-80, wherein the tumor and / or neoplasm is B cell lymphoma.

89. The method of claim 88, wherein the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma.

90. The method of any one of claims 77-80, wherein the subject is a human subject.

91. A method of reducing tumor burden in a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy ifi) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample, and / or vi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition of any one of claims 48-76.

92. The method of claim 91, wherein the method reduces the number of tumor cells, reduces tumor size, and / or eradicates the tumor in the subject.

93. A method of increasing or lengthening survival of a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy if i) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample, and / or vi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition of claim 48.

94. A method of treating and / or preventing a neoplasm in a subject that is non-responsive or has a reduced likelihood to respond to an adoptive cell therapy, comprising: a) measuring the expression level of at least one of CD57, TIM3, KLRG1, CTLA4, LAG3 and PD1 in an immunoresponsive cell of a subject; b) comparing the expression level to a reference sample; c) identifying the subject as non-responsive or as having a reduced likelihood to respond to an adoptive cell therapy if i) the expression level of CD57 is reduced compared to the reference sample, ii) the expression level of TIM3 is reduced compared to the reference sample, iii) the expression level of KLRG1 is increased compared to the reference sample, iv) the expression level of CTLA4 is increased compared to the reference sample, v) the expression level of LAG3 is increased compared to the reference sample and / orvi) the expression level of PD1 is reduced compared to the reference sample; and d) administering to the subject a composition of claim 48.

95. The method of any one of claims 91-94, wherein the tumor and / or neoplasm is associated with CD 19.

96. The method of any one of claims 91-94, wherein the tumor and / or neoplasm is a blood cancer.

97. The method of claim 96, wherein the blood cancer is selected from the group consisting of multiple myeloma, leukemia, and lymphomas.

98. The method of claim 97, wherein the leukemia is selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed- phenotype acute leukemia (MLL), hairy cell leukemia, and B cell prolymphocytic leukemia.

99. The method of claim 97, wherein the lymphoma is Hodgkin’s lymphoma or non-Hodgkin’s lymphoma.

100. The method of any one of claims 91-94, wherein the tumor and / or neoplasm is a B cell malignancy.

101. The method of claim 100, wherein the B cell malignancy is selected from the group consisting of B cell non-Hodgkin lymphomas (NHL), B cell Hodgkin's lymphomas, B cell acute lymphocytic leukemia (ALL), B cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter’s transformation, and CNS lymphoma.

102. The method of any one of claims 91-94, wherein the tumor and / or neoplasm is B cell lymphoma.

103. The method of claim 102, wherein the B cell lymphoma is relapsed or refractory (R / R) B cell lymphoma.

104. The method of any one of claims 91-94, wherein the subject is a human subject.

105. A kit for performing the method of any one of claims 1-47.

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