Targeting mutated KRAS by HLA-A*02:01 restricted anti-KRAS TCR-mimic CAR and bispecific T cell engager

Mutations in the KRAS proto-oncogene, particularly at codon 12, are among the most frequent genetic alterations in various cancers, and KRAS^G12V accounts for about 25% of all KRAS mutations observed in lung, pancreatic, and colorectal adenocarcinomas. Despite improved treatment regimes using targeted therapy and checkpoint inhibitors, cellular immunotherapy options for KRAS-mutated cancers remain elusive. We therefore developed two TCR-mimic (TCRm) anti-KRAS^G12V/HLA-A*02:01 chimeric antigen receptors (CARs) containing different hinge regions and, alternatively, a TCRm anti-KRAS^G12V/HLA-A*02:01 bispecific T cell engager (BiTE) to explore immunotherapy to the highly prevalent KRAS^G12V neoantigen. CAR-redirected or BiTE-exposed JNL-reporter cells demonstrated potent signaling capacity upon recognition of KRAS^G12V. Moreover, human CAR T and NK cells elicited IFN-γ release and cellular cytotoxicity upon encountering target cells pulsed with KRAS^G12V peptide, and the anti-KRAS^G12V Strep-tagII hinge CAR showed superior reactivity compared to a human IgG1-Fc hinge CAR. Similarly, a novel TCRm BiTE induced strong T cell immunity to KRAS^G12V. In contrast, we observed only very low CAR or BITE-mediated responses to naturally presented KRAS^G12V/HLA-A*02:01 complexes. In summary, this study demonstrates that the mutation-derived KRAS^G12V_5-14 peptide can be effectively targeted by TCRm CAR and BiTE-redirected T cells, suggesting that TCRm anti-KRAS^G12V CAR or BiTE represent promising formats to advance immunotherapy to mutated KRAS neoepitopes. KEY MESSAGES: Successful development of TCRm CAR and BiTE targeting mutated KRAS/HLA-A*02:01. Anti-KRAS^G12V TCRm CAR and BiTE induce potent immunity to KRAS^G12V neoepitope. Anti-KRAS/HLA-I TCRm CARs and BiTEs are novel therapeutics for cancer immunotherapy.