Ma Keke, Wang Jie, Jiang Min, He Juanhua, Li Fangyang, Lu Dan, Su Chao, Chai Yan, Jin Wenjing, Chen Yu, Zhang Catherine W H, Ma Xiaopeng, Tan Hui, Gao George F, Tan Shuguang
Department of Infectious Diseases, Shenzhen Children's Hospital, Shenzhen, 518026, China.
CAS Key Laboratory of Pathogen Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China.
Adv Sci (Weinh). 2025 Aug;12(31):e00181. doi: 10.1002/advs.202500181. Epub 2025 Jun 5.
T cell receptor (TCR) based immunotherapy is an attractive strategy to target a wide range of intra-tumoral antigens and elicit robust tumor cytotoxicity. However, engineering soluble TCR engagers that preserve physiological affinity is crucial for universal TCR drug development, yet remains challenging. In the present study, multiple TCR engagers featuring diverse architectures based on the KRAS-G12V specific 1-2C TCR in the context of HLA-A11:01 is designed and evaluated. Notably, a soluble tandem double single-chain TCR (STanD-scTCR) engager, comprising two repeated single-chain variable fragment (scFv) TCRs, exhibit enhanced binding avidity and potent T-cell activation. Through site-directed mutagenesis, T96F mutation (T96F-TCR) within the TCR β chain is identified, which substantially augment T cell reactivity while maintaining physiological affinity and minimizing off-target cross-reactivity. The T96F-mutated STanD-scTCR engager demonstrates improved antigen sensitivity, promotes multi-functional T-cell responses, and facilitates immune synapse formation between T cells and target cells. In a xenograft tumor model harboring the KRAS-G12V mutation, the TCR engager displays substantial tumor suppression efficacy. These findings underscore the therapeutic potential of 1-2C STanD-scTCR engage in targeting KRAS-G12V mutations in the context of HLA-A11:01. Furthermore, the engineering strategies employ in the development of STanD-scTCR engager provide an invaluable for future designs of TCR engager drugs.
基于T细胞受体（TCR）的免疫疗法是一种有吸引力的策略，可靶向多种肿瘤内抗原并引发强大的肿瘤细胞毒性。然而，构建保留生理亲和力的可溶性TCR衔接器对于通用TCR药物开发至关重要，但仍然具有挑战性。在本研究中，设计并评估了多种基于HLA-A11:01背景下KRAS-G12V特异性1-2C TCR的具有不同结构的TCR衔接器。值得注意的是，一种可溶性串联双链单链TCR（STanD-scTCR）衔接器，由两个重复的单链可变片段（scFv）TCR组成，表现出增强的结合亲和力和强大的T细胞激活能力。通过定点诱变，在TCRβ链内鉴定出T96F突变（T96F-TCR），该突变在保持生理亲和力并最小化脱靶交叉反应性的同时，大幅增强了T细胞反应性。T96F突变的STanD-scTCR衔接器表现出更高的抗原敏感性，促进多功能T细胞反应，并促进T细胞与靶细胞之间的免疫突触形成。在携带KRAS-G12V突变的异种移植肿瘤模型中，TCR衔接器显示出显著的肿瘤抑制功效。这些发现强调了1-2C STanD-scTCR衔接器在HLA-A11:01背景下靶向KRAS-G12V突变的治疗潜力。此外，在STanD-scTCR衔接器开发中采用的工程策略为未来TCR衔接器药物的设计提供了宝贵的借鉴。
