IMMU-70. Novel multispecific CAR T cell therapy to address GBM antigen heterogeneity and immunosuppression

Abstract Glioblastoma (GBM) is the most lethal and common primary brain cancer in adults. Chimeric antigen receptor (CAR) T cells are being actively explored for GBM treatment. However, antigen heterogeneity and an immunosuppressive, myeloid-rich tumor microenvironment (TME) pose significant challenges for CAR T cell therapy. To address this, we engineered dual-function IL13Rα2-directed CAR T cells (CAR.mNb T cells) that broaden antigen coverage and engage CD16a-expressing natural killer (NK) cells and tumor-associated macrophages (TAMs) in the TME by simultaneously secreting a multispecific nanobody (mNb) targeting EGFR, EGFRvIII, and CD16a. In vitro, CAR.mNb-T cells retained potent cytotoxic activity against IL13Rα2-positive patient-derived xenograft (PDX) GBM cells. In co-cultures of NK cells and GBM cells, CAR.mNb T cell conditioned media, containing recombinant mNb protein, induced NK cell-mediated cytotoxicity against GBM cells. Most notably, significantly enhanced antitumor activity against GBM cells was seen in co-cultures with CAR.mNb T cells and NK cells, irrespective of IL13RA2 expression. In co-cultures with TAMs and GBM cells, CAR.mNb T cells activated macrophages and induced cellular phagocytosis against GBM cells, while also supporting T cell proliferation. The in vivo studies underway will evaluate the therapeutic potential of this platform in GBM to guide clinical translation. Together, these findings demonstrate that CAR.mNb T cells can target GBM cells and engage innate immune cells against multiple GBM antigens, providing a potential strategy to overcome antigen heterogeneity and immunosuppression in GBM.