Abstract 6034: Development of a pan-HLA-A02 restricted AFP targeting TCR for both TCR-T and TCR-bispecific therapy

Abstract Background: Alpha-fetoprotein (AFP) is a well-established tumor-associated antigen, particularly in hepatocellular carcinoma (HCC). Although the AFP specific TCR-T cell therapy has shown promising clinical efficacy in HCC, the HLA restriction limits the patient population. We previously identified a T-cell receptor (TCR) sequence specific to an HLA-A*02:03-restricted AFP158-166 antigen epitope. To broaden its therapeutic potential, we mutated the sequence of CDR regions of this TCR to recognize multiple major HLA-A*02 subtypes. Method: We developed a novel human T-cell-based functional display platform to optimize the original AFP-specific TCR sequence. This platform facilitated the identification of TCR variants with broadened HLA-A*02 subtype recognition. The optimized TCR was engineered into TCR-T cells which were subsequently tested for their ability to recognize and lyse tumor cells both in vitro and in vivo models. Comprehensive safety evaluations were conducted to assess off-target effects. Additionally, the platform was also able to enhance the affinity of TCRs to picomolar level, leading to the development of a TCR bispecific protein. Results: The original TCR could only recognize and kill AFP158-166 antigen presented by HLA-A*02:03. In contrast, the optimized TCR transduced T cells demonstrated the ability to recognize the same epitope presented by pan-HLA-A*02 molecules, including A*02:01, A*02:03, A*02:06, and A*02:07, significantly broadening their applicability. The pan-HLA-A*02 restricted TCR transduced T cells exhibited superior anti-tumor activity, as evidenced by stronger cytokines production and tumor cell lysis ability in vitro and the capability to completely eradicate tumors in vivo murine models. Toxicology studies revealed no off-target recognition of non-specific HLA or other antigens, indicating a favorable safety profile. Moreover, the high-affinity TCRs (-11M) were linked with an anti-CD3 scFv fragment to form a TCR bispecific protein, which demonstrated substantial anti-tumor efficacy in preclinical models. Conclusion: To this end, the sequence-optimized AFP-specific TCR-T cells demonstrated robust therapeutic efficacy and a favorable safety profile, expanding potential treatment options for over 60% of patients with pan-HLA-A*02. Moreover, the successful development of a TCR bispecific antibody highlights its promise as an innovative off-the-shelf therapeutic solution, offering improved clinical accessibility and versatility to address the needs of a broader patient population. Citation Format: Junjun Chu, Jiajia Du, Tingfeng Liang, Shifan Wu, Yaolong Chen, Chunguang Yang, Qingjia Yan, Lulu Zhang, Yanyan Han. Development of a pan-HLA-A02 restricted AFP targeting TCR for both TCR-T and TCR-bispecific therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6034.