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Life sciences · Journal article
Biomedicines · September 23, 2026
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Background: Ovarian cancer treatment is frequently complicated by postoperative recurrence associated with metastatic dissemination and acquired chemotherapy resistance, highlighting the need for new therapeutic approaches. Messenger RNA (mRNA)-based chimeric antigen receptor (CAR) T-cell therapy is a promising strategy for solid tumors because it enables transient CAR expression and may shorten the manufacturing process compared with lentiviral approaches. Given the potential relevance of rapid, transient CAR expression to locoregional or adjuvant applications, we evaluated an mRNA-based CAR-T cell approach targeting cancer antigen 125 (CA125) in vitro. Methods: CAR-T cells were generated from either activated or nonactivated (“resting”) lymphocytes obtained from healthy donors using mRNA electroporation. Their activity against target tumor cells was assessed by impedance-based real-time cell analysis (RTCA) using the xCELLigence platform. Results: We compared two mRNA CAR-T cell manufacturing strategies: “fast” CAR-T cells generated using a 3-day activation-based workflow and “ultra-fast” CAR-T cells generated from nonactivated T cells and functionally evaluated within 24 h after electroporation. “Fast” CAR-T cells exhibited strong in vitro activity, including at an effector-to-target ratio of 1:4, representing a high target-cell burden. “Ultra-fast” CAR-T cells were also functional but showed lower activity, with an observable effect requiring an effector-to-target ratio of at least 2.5:1. Conclusions: These findings support the feasibility of rapid mRNA-engineered anti-CA125 CAR-T cell manufacturing using either activated or nonactivated T cells. The two approaches differ in functional activity and may therefore be suitable for different manufacturing workflows and intended applications.