Human lung explants as a predictive platform for evaluating the on-target, off-tumor toxicity of T cell bispecifics
T cell bispecifics (TCBs) are potent immunotherapies with proven efficacy in hematological cancers but limited success in solid tumors, where on-target, off-tumor toxicity has restricted their therapeutic index. Clinical discontinuation of TCB formulations against epithelial cell adhesion molecule (EpCAM TCB, Solitomab) and preclinical termination of a TCB against folate receptor alpha (FolR1-TCB) highlight the urgent need for predictive preclinical models to evaluate such toxicities. Conventional models, including cell lines and organoids, lack the immune and stromal complexity of the tumor microenvironment (TME), and while patient-derived explants preserve native tissue architecture and heterogeneity, quantification of TCB-mediated target cell killing has not been previously achieved in this system. Here, we establish a human lung explant model to assess TCB activity in matched tumor and normal adjacent tissues. Using a multi-modal approach combining flow cytometry, cytokine profiling, and multiplexed immunofluorescence, we show that EpCAM and FolR1 TCBs drive robust T cell activation and, critically, quantifiable epithelial cell killing above background levels. Normal lung explants exhibited consistently higher levels of killing than tumor counterparts, reflecting the on-target, off-tumor toxicities observed preclinically and clinically with the tested TCBs. In contrast, colon explants displayed poor viability ex vivo, limiting their suitability for assessing TCB-induced killing despite measurable immune activation. Our findings establish lung explants as a predictive and clinically relevant preclinical model that uniquely enables simultaneous quantification of TCB-mediated T cell activation and target cell killing. This model captures key features of TCB mechanism of action and recapitulates clinically observed toxicities, supporting its application for preclinical evaluation of TCB drug candidates and advancing the development of immunotherapies for solid tumors.