Combining TAG-72CAR with anti-MUC16CAR improves the potency of ovarian cancer treatment.

Abstract

Ovarian cancer (OC) is typically associated with poor prognosis and high mortality. Chimeric antigen receptor (CAR) T-cell therapy offers a promising strategy, yet its application in solid tumours remains constrained by limited efficacy and on-target/off-tumour toxicities. We previously developed K101CAR T cells targeting CA125, the cleaved extracellular domain of MUC16, and demonstrated potent antitumour activity against OC. However, because MUC16 exhibits low-level expression in certain normal tissues, safety concerns remain regarding potential off-tumour effects. We designed a combinatorial CAR composed of two single-chain variable fragments (scFvs): (i) TAG-72 targeting the Sialyl-Tn antigen highly expressed on malignant cells, and (ii) the K101 scFv. The first unit is linked to a CD3ζ signalling domain, whereas the second is linked to a 4-1BB costimulatory domain, leading to a combinatorial CAR fully reactive against double-positive targets, Tz-KBBCAR. We demonstrate that Tz-KBBCAR T cells efficiently eliminate MUC16+/TAG-72+-, while sparing MUC16+/TAG-72-- cells in vitro and markedly suppress tumour progression in vivo. We also observed that compared to K101CAR T cells, Tz-KBBCAR T cells exhibit enhanced expansion, sustained viability, and a reduced exhaustion phenotype. Overall, our study presents an efficient combinatorial CAR T-cell strategy predicted to be safer than single CAR T-cell therapy for OC.