IMMU-17. REDUCING THE EX VIVO MANUFACTURING TIME OF EGFRVIII-SPECIFIC CHIMERIC ANTIGEN RECEPTOR (CAR) T CELLS IMPROVES PHENOTYPE AND DEMONSTRATES POTENT ANTI-GLIOMA FUNCTION IN VIVO

摘要

Chimeric antigen receptor (CAR) T-cell therapy for glioma has been developed that targets EGFRvIII, HER2, EphA2, and IL13Rα2 antigens. Epidermal growth factor receptor variant III (EGFRvIII) is an attractive target for CAR therapy because its expression is tumor-restricted. Non-viral manufacturing of CAR T cells via Sleeping Beauty transposition is cost effective and reduces risk of insertional mutagenesis from viral transduction. However, the current gold standard methodology requires ex vivo expansion on artificial antigen-presenting cells (AaPCs) for 4 weeks to eliminate competing NK cells from culture and to generate sufficient cell numbers for clinical application. We monitored phenotypic changes of EGFRvIII-specific CAR T cells over the course of ex vivo manufacturing and found significantly increased expression of the exhaustion markers such as PD-1, PD-L1, TIM-3, and LAG-3 after two weeks of culture, which continued to rise over time. To reduce the culture time required to generate the CAR T cell population, we selected for T cells in peripheral blood mononuclear cells prior to CAR modification (to eliminate the NK cell population) and were able to generate a CAR+ T cell population with comparable CAR expression and cell numbers in two weeks—half the time required by the standard protocol and with lower expression of exhaustion markers. We treated mice bearing established orthotopic EGFRvIII+ U87 gliomas with EGFRvIII-specific CAR T cells derived using the expedited approach and showed significant suppression of tumor growth after one treatment. Thus, T-cell selection prior to CAR modification produces CAR T cells with an improved phenotype and potent in vivo function in less time, which will decrease the wait time for patients.