M9657, a bispecific tumor-targeted conditional anti-CD137 agonist, induces MSLN-dependent antitumor immunity without liver inflammation

The costimulatory receptor CD137 (also known as TNFRSF9 or 4-1BB) is crucial insustaining effective cytotoxic T cell immune responses. Agonistic anti-CD137 mAb cancerimmunotherapies are being investigated in clinical trials. First-generation CD137-agonistmonotherapies, utomilumab and urelumab, were unsuccessful due to low antitumor efficacy orhepatotoxicity, respectively, mediated by the epitope recognized on CD137 and Fc-gamma-R ligand-dependent CD137 activation. M9657 was engineered as a tetravalent bispecific antibody(mAb2 TM) in a human IgG1 backbone with LALA mutations to reduce binding to the Fc-gammareceptors. Here we report that M9657 selectively binds to mesothelin (MSLN) and CD137 withsimilar affinity in humans and cynomolgus monkeys. In a cellular functional assay, M9657enhanced CD8+ T cell-mediated cytotoxicity and cytokine release in the presence of tumor cells,which was dependent on both MSLN expression and TCR/CD3 activation (Signal 1). BothFS122m, a murine surrogate with the same protein structure as M9657, and chimeric M9657, amodified M9657 antibody with the Fab portion replaced with an anti-murine MSLN motif,demonstrated in vivo antitumor efficacy against various tumors in wild-type or human CD137knock-in (KI) mice, accompanied by activated CD8+ T cell infiltration in the tumormicroenvironment. The antitumor immunity of M9657 and FS122m depended on MSLNexpression density and the mAb2 TM structure. Compared with 3H3, a murine surrogate ofurelumab, FS122m and chimeric M9657 displayed significantly lower on-target/off-tumortoxicity. Taken together, M9657 exhibits a promising developability profile as a tumor-targetedimmune agonist with potent anticancer activity without systemic immune activation andassociated hepatotoxicity.