Characterization of TROP-2 bispecific T cell engagers for immunotherapy of triple negative breast and bladder cancer.

T cell-based immunotherapy has markedly expanded the therapeutic options in numerous cancers. However, these approaches still achieve only limited clinical benefit in triple negative breast cancer (TNBC) and bladder cancer. Although immune checkpoint inhibitors improve outcomes for a subset of patients, no T cell-redirecting therapies such as CAR-T cells or bispecific antibodies (bsAbs) have been approved for either indication. Trophoblast cell surface antigen 2 (TROP-2) is highly expressed across several epithelial cancers including TNBC and bladder cancer, but has been primarily exploited as a target for antibody drug conjugates (ADCs) with limited exploration in T cell-engaging constructs. Here, we report on the generation and characterization of a panel of TROP-2×CD3 bsAbs containing clinically validated TROP-2 binders and CD3 binders with distinct affinities. All bsAbs induced robust T cell activation, cytokine secretion and sustained T cell expansion, resulting in potent T cell-mediated cytotoxicity against TNBC and bladder cancer cells with either high or low levels of TROP-2 expression. Notably, combining a TROP-2 binder with enhanced tumor selectivity and a low-affinity CD3 binder increased discrimination between high and very low TROP-2-expressing cells while (reducing cytokine release without compromising anti-tumor efficacy. Thus, TROP-2-directed bsAbs can achieve effective tumor cell killing without over dependence on antigen density, in contrast to ADC-based approaches. Our results support further development of TROP-2×CD3 bsAbs as immunotherapy for solid tumors with heterogeneous or low TROP-2 expression.