EZH2 Inhibition in Ewing Sarcoma Upregulates GD2 Expression for Targeting with Gene-Modified T Cells

Kailayangiri S, Altvater B, Lesch S, Balbach S, Göttlich C, Kühnemundt J, Mikesch JH, Schelhaas S, Jamitzky S, Meltzer J, Farwick N, Greune L, Fluegge M, Kerl K, Lode HN, Siebert N, Müller I, Walles H, Hartmann W, Rossig C

Abstract

Chimeric antigen receptor (CAR) engineering of Tcells allows one to specifically target tumor cells via cell surface antigens. A candidate target in Ewing sarcoma is the ganglioside GD2, but heterogeneic expression limits its value. Here we report that pharmacological inhibition of Enhancer of Zeste Homolog 2 (EZH2) at doses reducing H3K27 trimethylation, but not cell viability, selectively and reversibly induces GD2 surface expression in Ewing sarcoma cells. EZH2 in Ewing sarcoma cells directly binds to the promoter regions of genes encoding for two key enzymes of GD2 biosynthesis, and EZH2 inhibition enhances expression of these genes. GD2 surface expression in Ewing sarcoma cells is not associated with distinct invitro proliferation, colony formation, chemosensitivity, or invivo tumorigenicity. Moreover, disruption of GD2 synthesis by gene editing does not affect its invitro behavior. EZH2 inhibitor treatment sensitizes Ewing sarcoma cells to effective cytolysis by GD2-specific CAR gene-modified Tcells. In conclusion, we report a clinically applicable pharmacological approach for enhancing efficacy of adoptively transferred GD2-redirected Tcells against Ewing sarcoma, by enabling recognition of tumor cells with low or negative target expression.