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Updated: May 15, 2026
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Generation of Human Chimeric Antigen Receptor Regulatory T Cells
Published on: January 3, 2025
Efficient selection of genetically modified human T cells using methotrexate-resistant human dihydrofolate reductase
M Jonnalagadda1, C E Brown, W C Chang
1Departments of Cancer Immunotherapeutics and Tumor Immunology, and Hematology and Hematopoietic Cell Transplantation, Beckman Research Institute, City of Hope National Medical Center, Duarte, CA 91010, USA.
Gene Therapy
|January 11, 2013
Summary
Researchers developed a novel method to purify genetically modified T cells for cancer therapy. This system uses the drug methotrexate to select T cells expressing chimeric antigen receptors (CARs), ensuring product purity and potency for clinical use.
Area of Science:
- Immunology
- Gene Therapy
- Biotechnology
Background:
- Chimeric antigen receptor (CAR) T cell therapy is a promising cancer treatment.
- Efficient purification of genetically modified T cells is crucial for therapeutic efficacy and safety.
- Current purification methods are often complex and costly.
Purpose of the Study:
- To develop a simple and efficient method for purifying transgene-positive T cells.
- To create a selection system for T cells engineered to express CARs.
- To facilitate the manufacturing of clinical-grade T cell products.
Main Methods:
- Development of self-inactivating (SIN) lentiviral vectors for coordinated gene expression.
- Incorporation of a methotrexate-resistant human dihydrofolate reductase (DHFR) mutein (huDHFR(FS)).
- Utilizing methotrexate (MTX) as a selection agent to isolate modified T cells.
Main Results:
- Engineered T cells expressing huDHFR(FS) demonstrated resistance to lymphotoxic concentrations of MTX.
- Selected T cells maintained co-expression of CD19-specific CAR and EGFRt.
- The selection system proved effective in purifying genetically modified T cells.
Conclusions:
- The developed MTX-based selection system offers a promising approach for manufacturing clinical-grade T cell products.
- This method enhances T cell purification efficiency and consistency.
- The system has potential utility in advancing CAR T cell therapy manufacturing.
