Methods for producing autologous t cells useful to treat b cell malignancies and other cancers and compositions thereof

A cell population and cell technology, applied in the direction of receptors/cell surface antigens/cell surface determinants, cell culture active agents, chemical instruments and methods, etc., can solve problems that are difficult to commercially apply and time-consuming

Pending Publication Date: 2017-02-22
凯德药业公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The process for producing autologous engineered T cells for use in cancer therapy is time-consuming (10-24 days), involves two cycles of retroviral t

Method used

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  • Methods for producing autologous t cells useful to treat b cell malignancies and other cancers and compositions thereof
  • Methods for producing autologous t cells useful to treat b cell malignancies and other cancers and compositions thereof
  • Methods for producing autologous t cells useful to treat b cell malignancies and other cancers and compositions thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1: Preparation of ex vivo genetically modified autologous cells

[0067] figure 1 A schematic diagram of an exemplary T cell production process ("improved" process) according to one embodiment is provided. This improved process also includes improvements to the traditionally used process for producing T cells (the "previous" process) (see the figure 2 ), while maintaining T cell product characteristics. Specifically, the improved process is a closed process that surprisingly avoids the use of serum. Moreover, this improved process uses a single cycle of transduction to produce a population of transduced T cells. Furthermore, cells subjected to a total expansion time of 6 days showed a more naïve immunophenotypic profile when using this procedure compared to cells subjected to an expansion time of 10 days. This process enables reproducible production of products with target numbers of transduced T cells expressing a chimeric antigen receptor (CAR) such as CD...

Embodiment 2

[0081] Example 2: Growth performance of T cells expanded in cell culture bags

[0082]Embodiments described herein provide a method for efficient production of engineered autologous T cell therapy within 6 days. The following improvements have been achieved over the existing technology: the process time has been shortened to 6 days instead of the 24, 14 or 10 days previously used (this reduces the number of tests (including RCR tests) required for product release); T cell products, including a higher proportion of naive T cells, achieve increased potency and effectiveness; larger numbers of cells can be used to initiate culture to compensate for shorter manufacturing times; closed systems are used to perform the methods described in the present invention identification of serum-free culture conditions that support T cell growth; single-cycle retroviral transduction in culture bags; cell culture activation and expansion in culture bags rather than vials; The product. These im...

Embodiment 3

[0102] Example 3: Development of transduction conditions in a closed system

[0103] Previously, transduction of PBMCs was performed in 6-well plates treated with non-tissue culture. These plates were incubated at 2-8°C with 10 μg / mL of Coating was performed overnight, or at room temperature for 2 hours. After incubation, remove Plates were blocked with 2.5% HSA for 30 minutes, followed by washing with HBSS+5mM HEPES. In the plate-based process, retroviral vectors are applied to coated wells and spun down in a centrifuge, followed by removal of approximately 75% of the viral supernatant, followed by addition of cells by spinnoculation for transduction .

[0104]In the present invention, three studies were performed to optimize the method for transducing PBMC in closed cell culture bags concentrations and to determine whether HSA washing and virus supernatant removal affected transduction. The first experiment was in Origen PermaLife TM PL07 culture bag, in which cel...

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Abstract

Provided herein are methods for manufacturing T cells. In certain embodiments, methods for manufacturing T cells which express a cell surface receptor that recognizes a specific antigenic moiety on the surface of a target cell are provided. Such methods may include, but are not limited to, steps of (1 ) enriching a population of lymphocytes obtained from a donor subject; (2) stimulating the population of lymphocytes with one or more T-cell stimulating agents to produce a population of activated T cells, wherein the stimulation is performed in a closed system using serum-free culture medium; (3) transducing the population of activated T cells with a viral vector comprising a nucleic acid molecule which encodes the cell surface receptor, using a single cycle transduction to produce a population of transduced T cells, wherein the transduction is performed in a closed system using serum-free culture medium; and (4) expanding the population of transduced T cells for a predetermined time to produce a population of engineered T cells, wherein the expansion is performed in a closed system using serum-free culture medium. Also provided herein are populations of engineered T cells produced by the methods described herein and pharmaceutical compositions thereof.

Description

[0001] Cross References to Related Applications [0002] This application claims the benefit of US Provisional Patent Application 61 / 935,833, filed February 4, 2014, which is incorporated herein by reference in its entirety. [0003] Statement of Government Interest [0004] This invention was created under a Cooperative Research and Development Agreement with the National Cancer Institute (NCI), an agency of the US Department of Health and Human Services. The US Government has certain rights in this invention. Background technique [0005] The process for producing autologous engineered T cells for use in cancer therapy is time-consuming (10-24 days), involves two cycles of retroviral transduction, and is poorly suited for commercial application (see Kochenderfer et al., Blood 2012119:2709-2720; Johnson et al., Blood. 2009;114(3):535-546). Therefore, it is desirable to develop improved methods of the T cell manufacturing process to overcome these limitations. Contents of...

Claims

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Application Information

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IPC IPC(8): C12N5/0783C12N5/10A61K35/17A61P35/00
CPCC12N2500/90C12N2501/2302C12N2501/515C12N2510/00A61K38/1774C12N5/0636A61P35/00C07K2319/03C07K14/7051A61K39/4611A61K39/4631A61K39/464412A61K35/17A61K39/0011A61K39/39558A61K2039/5156A61K2039/5158C07K16/2896C07K2319/74
Inventor 马克·贝特尔史蒂芬·A·费德曼史蒂芬·A·罗森伯格
Owner 凯德药业公司
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