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A Nonviral Approach to Generate Transient Chimeric Antigen Receptor T Cells Using mRNA for Cancer Immunotherapy
Published on: February 21, 2025
Redirected antitumor activity of primary human lymphocytes transduced with a fully human anti-mesothelin chimeric
Evripidis Lanitis1, Mathilde Poussin, Ian S Hagemann
1Department of Obstetrics and Gynecology, Ovarian Cancer Research Center, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Abstract:
Cancer regression by gene-modified T cells bearing a chimeric antigen receptor (CAR) exodomain of mouse origin can be limited by the induction of transgene immunogenicity resulting in poor persistence and function in vivo. The development of functionally-active CAR of human origin can address this issue. Here, we constructed and evaluated fully human anti-mesothelin CARs comprised of a human mesothelin-specific single-chain antibody variable fragment (P4 scFv) coupled to T cell signaling domains. Primary human T cells expressing P4 CAR specifically produced proinflammatory cytokines, degranulated and exerted potent cytolytic functions when cultured with mesothelin-expressing tumors in vitro. P4 CAR T cells also mediated bystander killing of mesothelin-negative cancer cells during coculture. CAR reactivity was not abrogated by soluble tumor-secreted or recombinant mesothelin protein even at supraphysiological levels. Importantly, adoptive transfer of P4 CAR-expressing T cells mediated the regression of large, established tumor in the presence of soluble mesothelin in a xenogenic model of human ovarian cancer. Thus, primary human T cells expressing fully human anti-mesothelin CAR efficiently kill mesothelin-expressing tumors in vitro and in vivo and have the potential to overcome the issue of transgene immunogenicity that may limit CAR T cell trials that utilize scFvs of mouse origin.
Insights
Fully human anti-mesothelin CAR T cells show potent anti-cancer activity, overcoming immune rejection. These engineered T cells effectively target and eliminate mesothelin-expressing tumors in vitro and in vivo.
Area of Science:
- Immunology
- Oncology
- Biotechnology
Background:
- Chimeric antigen receptor (CAR) T cell therapy efficacy can be limited by immune responses against mouse-derived CAR components.
- Developing CARs with fully human components is crucial to enhance T cell persistence and function in cancer patients.
Purpose of the Study:
- To construct and evaluate fully human anti-mesothelin CARs to overcome transgene immunogenicity.
- To assess the in vitro and in vivo efficacy of these human CAR T cells against mesothelin-expressing cancers.
Main Methods:
- Construction of fully human anti-mesothelin CARs using a human mesothelin-specific single-chain variable fragment (P4 scFv).
- Assessment of primary human T cell effector functions (cytokine production, degranulation, cytotoxicity) against mesothelin-expressing tumor cells in vitro.
- Evaluation of P4 CAR T cell efficacy in a xenogenic ovarian cancer model, including bystander killing and response in the presence of soluble mesothelin.
Main Results:
- P4 CAR T cells demonstrated specific proinflammatory cytokine production, degranulation, and potent cytolytic activity against mesothelin-positive tumors in vitro.
- CAR reactivity remained unaffected by high levels of soluble mesothelin.
- Adoptive transfer of P4 CAR T cells led to the regression of established tumors in a human ovarian cancer xenograft model, even with soluble mesothelin present.
Conclusions:
- Fully human anti-mesothelin CAR T cells are effective against mesothelin-expressing tumors, both in vitro and in vivo.
- This approach has the potential to mitigate transgene immunogenicity issues encountered with mouse-derived CARs, improving CAR T cell therapy outcomes.
