DMG-20. A potent and safe ‘Gated’ CAR-T cell therapy approach for DIPG

Abstract Diffuse intrinsic pontine glioma (DIPG) are highly aggressive and difficult-to-treat brain tumors in children. Recent advances in Chimeric Antigen Receptor (CAR) expressing T cell therapy suggest application for brain tumors, including DIPG. We recently identified CD99 as a highly expressed antigen in DIPG cells and an immunotherapy target in DIPG. Though the monovalent CD99 CAR-T cells cleared the tumor initially, CD99-driven fratricide limited its long-term anti-tumor efficacy against DIPG. DIPG cells uniquely express the combination of CD99 and B7-H3 at high levels. Therefore, to overcome CD99-driven fratricide, we developed a bicistronic CAR construct that employs Boolean logic to ‘gate’ the function of CAR-T cells according to ‘AND’ rules, where two distinct antigens are necessary for CAR-T cell activation. In our model, we used CD99 CAR (based on our new antibody-based scFV) in conjunction with B7-H3 CAR (enoblituzumab antibody-based scFv). The “AND” CAR-T cells 1) showed enhanced tumor cell killing when co-cultured with DIPG tumor cells; 2) the AND gating protected the T cells from CD99-driven fratricide and demonstrated no functionality against either single antigen-­expressing cells, suggesting the specificity of the AND CAR-T in targeting dual-antigen-expressing tumor cells; and 3) a single low dose of “AND” CAR-T cells delivered either IV or intrathecally completely cleared DIPG in the murine models with a significant increase in animal survival while either antigen targeting monovalent CAR-T cells showed only temporary clearance of tumor as the tumor relapsed after treatment. In addition, during the meeting, we will discuss the AND CAR-T cell’s long-term persistence, its safety in protecting single antigen-expressing cells using humanized mouse models, the mechanism of enhanced efficacy against heterogeneously expressed tumor target antigens, its efficacy when combined with radiation and discuss other factors governing its potential clinical applicability to treat DIPG.