Identification of TM4SF1 as a novel tumor-associated antigen in biliary tract cancers targetable by CAR-T cell therapy

Background & Aims: Biliary tract cancers (BTCs) are a rare set of genetically heterogenous and aggressive malignancies associated with delayed presentation and poor prognosis. Thus, there is an unmet clinical need for the development of novel therapeutic strategies. Identification of uniquely expressed cell-surface, tumor-associated antigens (TAAs) holds promise in epithelial tumors more generally, as they can serve as ligands for a variety of therapeutics, including CAR-T cell therapy. However, TAA identification in BTCs has been limited by expression in normal liver tissue. Here, we aim to characterize the cell-surface protein transmembrane 4 L six family member 1 (TM4SF1) as a potential BTC TAA targetable by CAR-T therapy. Methods: TM4SF1 expression in BTC relative to normal hepatic/biliary tissue was characterized at the mRNA level by analyzing TCGA datasets and at the protein level by immunohistochemistry of archival primary BTC tissue. Anti-TM4SF1 CAR-T cells were evaluated through in vitro coculture assays against human-derived BTC cell lines and in a cell line-derived xenograft (CDX) model. Results: We find TM4SF1 is significantly upregulated in BTC relative to normal hepatic/biliary tissue and across BTC anatomical subtypes at the mRNA and protein level. To validate TM4SF1 as a targetable TAA in BTC, we show anti-TM4SF1 CAR-T cells demonstrate robust growth inhibition of BTC cell lines in vitro and significant anti-tumor activity against a CDX in vivo. Conclusions: Together, our data provides support for TM4SF1 as a promising TAA in BTCs that can be used for the rational development of targeted therapeutic modalities, including CAR-T cells, in a patient population with significant unmet need.