463 Tandem CAR T Cells Targeting EGFRvIII and IL-13R⍺2 are Effective Against Heterogeneous Glioblastoma

INTRODUCTION: Chimeric antigen receptor (CAR) T cells have achieved remarkable responses in patients with hematological malignancies; however, the potential of this therapeutic platform for solid tumors like glioblastoma (GBM) has been limited, due in large part to the targeting of single antigens in a heterogenous disease. Strategies that allow CAR T cells to engage multiple antigens concomitantly may broaden therapeutic responses and mitigate the effects of immune escape. METHODS: We generated a TanCART construct with dual specificity consisting of both the humanized anti-EGFRvIII scFv and IL-13 zetakine translated in tandem separated by a flexible glycine-serine linker. We also employed two monospecific CAR T cells, CART-EGFRvIII and CART-IL-13Rɑ2 as controls. We used both standard immunological assays and multiple orthotopic preclinical models including patient-derived xenograft to demonstrate efficacy of this approach against heterogenous tumors. RESULTS: Tandem CAR T cells displayed enhanced cytotoxicity in vitro against heterogeneous GBM populations, including patient-derived brain tumor cultures (P < 0.05). Compared to CAR T cells targeting single antigens, dual antigen engagement through the tandem construct was necessary to achieve long-term, complete and durable responses in orthotopic murine models of heterogeneous GBM, including patient-derived xenografts (P < 0.05). CONCLUSIONS: We demonstrate that TanCART is effective against heterogenous tumors in the brain. To our knowledge, this is the first report of a tandem CAR T cell targeting EGFRvIII and IL-13Rɑ2 simultaneously. These data lend further credence to the development of multi-specific CAR T cells in the treatment of GBM and other cancers.