Related Experiment Video
Updated: Aug 10, 2026
Non-Viral Engineering of Primary Human T Cells via Homology-Mediated End-Joining Targeted Integration of Large DNA Templates
Published on: May 9, 2025
Simultaneous gene engineering of T cells with multiple neoantigen-specific T cell receptors for adoptive cell
Melinda Magna1, Lior Levy1, Krisztian Csomos2
1Surgery Branch, Center for Cancer Research, National Cancer Institute, NIH, 10 Center Drive, Bethesda, MD 20892, USA.
Abstract:
T cell receptor (TCR) T cell therapy targeting tumor-specific antigens and neoantigens can mediate impressive tumor regressions in solid tumors. However, solid epithelial tumors are characterized by inherent tumor heterogeneity that can lead to tumor escape and thwart TCR T cell therapy. Targeting multiple cancer antigens in a single TCR T product may help overcome the antigen heterogeneity among metastatic deposits. However, the time and expense of preparing multiple TCR-transduced T cell populations under good manufacturing practice (GMP) for patient treatment has limited this approach to target 1 or 2 antigens. Here, we design strategies to enhance TCR T cell therapy efficacy by simultaneously targeting multiple neoantigens. We developed two novel manufacturing processes to introduce multiple neoantigen-specific TCRs into T cells, using gamma-retroviral delivery. These strategies resulted in multipotent neoantigen-reactive TCR T cell products, which demonstrated functionality against multiple tumor neoantigens and displayed cytotoxicity against heterogeneous tumor cells in vitro. High-dimensional single-cell and single-clone T cell analysis confirmed the presence of multiple TCR-expressing T cells in the products and demonstrated functional recognition of multiple neoantigens. This study, thus, addresses the challenges of tumor heterogeneity and immune escape mechanisms in solid tumors, paving the way for more effective TCR T cell therapies in cancer treatment.
Insights
This study developed novel manufacturing processes to create T cell receptor (TCR) T cell therapies targeting multiple neoantigens simultaneously. These enhanced TCR T cell products effectively combat tumor heterogeneity and immune escape in solid tumors.
Area of Science:
- Immunology
- Oncology
- Biotechnology
Background:
- T cell receptor (TCR) T cell therapy shows promise for solid tumors but struggles with tumor heterogeneity and immune escape.
- Current limitations include the time and cost of preparing multiple TCR T cell products for targeting several antigens.
Purpose of the Study:
- To design and develop strategies for enhancing TCR T cell therapy efficacy by simultaneously targeting multiple neoantigens.
- To overcome challenges posed by tumor heterogeneity and immune escape in solid tumors.
Main Methods:
- Developed two novel gamma-retroviral manufacturing processes to introduce multiple neoantigen-specific TCRs into T cells.
- Utilized high-dimensional single-cell and single-clone T cell analysis to confirm product composition and functionality.
Main Results:
- Generated multipotent neoantigen-reactive TCR T cell products capable of targeting multiple tumor neoantigens.
- Demonstrated in vitro cytotoxicity against heterogeneous tumor cells, confirming the efficacy of the developed products.
- Confirmed the presence of multiple TCR-expressing T cells and their functional recognition of multiple neoantigens.
Conclusions:
- The novel manufacturing strategies successfully created multi-neoantigen targeting TCR T cell products.
- These products address critical challenges in solid tumor treatment, including heterogeneity and immune escape.
- This approach paves the way for more effective TCR T cell therapies in cancer treatment.
