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Updated: May 15, 2026
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Generation of CAR T Cells for Adoptive Therapy in the Context of Glioblastoma Standard of Care
Published on: February 16, 2015
Generation and Characterization of CAR-T Cells
Sarah Ash1, Greta Maria Paola Giordano Attianese1, Paris Kosti1
1Department of Fundamental Oncology, Ludwig Institute for Cancer Research Lausanne Branch, University of Lausanne, Lausanne, Switzerland.
Methods in Molecular Biology (Clifton, N.J.)
|May 13, 2026
Summary
This study details protocols for engineering next-generation chimeric antigen receptor (CAR)-T cells for solid tumors. It covers lentivirus/retrovirus production, T cell manipulation, in vitro assays, and in vivo mouse models for enhanced cancer therapy.
Area of Science:
- Immunology
- Molecular Biology
- Biotechnology
Background:
- Chimeric antigen receptors (CARs) are synthetic receptors enabling T-cell activation against specific antigens.
- Current clinical CAR-T therapies primarily use second-generation (2G) CARs, necessitating advancements for solid tumor treatment.
- Next-generation CAR-T cells aim to enhance function and safety, particularly for overcoming solid tumor microenvironment challenges.
Purpose of the Study:
- To present optimized laboratory protocols for developing and evaluating advanced CAR-T cell therapies.
- To detail methods for lentivirus and retrovirus production, T cell engineering, and characterization.
- To provide a framework for preclinical assessment of CAR-T cells in solid tumor models.
Main Methods:
- Development of switchable CARs for remote control of T cell activity.
- Implementation of non-viral (CRISPR/Cas9, base editing, transposons) and viral (lentivirus, retrovirus) gene-editing tools.
- Establishment of protocols for CAR-T cell production, purification, activation, transduction, expansion, and in vitro/in vivo assays.
Main Results:
- Detailed protocols for lentivirus and retrovirus production and titration are provided.
- Methods for purification, activation, transduction, and expansion of mouse and human CAR-T cells are described.
- In vitro assays for assessing CAR-T cell efficacy (transduction, proliferation, cytotoxicity) and in vivo study designs are presented.
Conclusions:
- The presented protocols facilitate the development and preclinical evaluation of enhanced CAR-T cells for solid tumor treatment.
- These methods are adaptable for T cell receptor (TCR)-engineered T cells.
- The protocols support comprehensive analysis from CAR-T cell production to in vivo efficacy studies.
