胰腺癌是最致命的恶性肿瘤之一，缺乏治疗方案使其更加致命。嵌合抗原受体T细胞（CAR-T）免疫疗法彻底改变了癌症治疗，在治疗血液恶性肿瘤方面取得了巨大突破，但其在治疗实体癌方面的成功仍然有限，主要是由于缺乏肿瘤特异性抗原。另一方面，漫长的传统制造过程带来了挑战，需要 2 至 6 周的时间，并影响患者的治疗结果。 CD276最近已成为抗实体癌症治疗的潜在治疗靶点。在这里，我们研究了CD276 CAR-T和快速制造的CAR-T对抗胰腺癌的功效。在本研究中，CD276 CAR-T是由携带376.96 scFv序列、CD8铰链和跨膜结构域、4-1BB的CAR结构制备的和 CD3ze 细胞内结构域。此外，创新开发了CD276快速制造CAR-T（命名为CD276 Dash CAR-T），通过缩短体外培养时间，减少CAR-T制造时间。我们评估了CD276 CAR-T的抗肿瘤功效，并通过检测CAR的免疫表型、杀伤能力、扩增能力和肿瘤根除效果，进一步比较了Dash CAR-T和常规CAR-T在体外和体内的功能评估-T.我们发现CD276在多种实体癌细胞系中强表达，并且CD276 CAR-T可以有效杀死这些实体癌细胞。此外，Dash CAR-T 在 48-72 小时内成功制造，并随后进行了功能验证。在体外，与传统CAR-T相比，CD276 Dash CAR-T具有低分化表型和强大的增殖能力。在体内异种移植小鼠模型中，CD276 Dash CAR-T 显示出增强的抗胰腺癌功效和 T 细胞扩增。此外，除高剂量组外，小鼠体重均保持稳定，小鼠状态正常。在本研究中，我们证明CD276 CAR-T在体外和体内均表现出强大的抗胰腺癌细胞活性。更重要的是，我们证明了在更短的时间内生成的 CD276 Dash CAR-T 的制造可行性、可接受的安全性和卓越的抗肿瘤功效。上述研究结果表明，CD276 Dash CAR-T 免疫疗法可能是一种新颖且有前景的胰腺癌治疗策略。© 2024。作者。

Pancreatic cancer is one of the most lethal malignancies and the lack of treatment options makes it more deadly. Chimeric Antigen Receptor T-cell (CAR-T) immunotherapy has revolutionized cancer treatment and made great breakthroughs in treating hematological malignancies, however its success in treating solid cancers remains limited mainly due to the lack of tumor-specific antigens. On the other hand, the prolonged traditional manufacturing process poses challenges, taking 2 to 6 weeks and impacting patient outcomes. CD276 has recently emerged as a potential therapeutic target for anti-solid cancer therapy. Here, we investigated the efficacy of CD276 CAR-T and rapidly-manufactured CAR-T against pancreatic cancer.In the present study, CD276 CAR-T was prepared by CAR structure carrying 376.96 scFv sequence, CD8 hinge and transmembrane domain, 4-1BB and CD3ζ intracellular domains. Additionally, CD276 rapidly-manufactured CAR-T (named CD276 Dash CAR-T) was innovatively developed by shortening the duration of ex vitro culture to reduce CAR-T manufacturing time. We evaluated the anti-tumor efficacy of CD276 CAR-T and further compared the functional assessment of Dash CAR-T and conventional CAR-T in vitro and in vivo by detecting the immunophenotypes, killing ability, expansion capacity and tumor-eradicating effect of CAR-T.We found that CD276 was strongly expressed in multiple solid cancer cell lines and that CD276 CAR-T could efficiently kill these solid cancer cells. Moreover, Dash CAR-T was successfully manufactured within 48-72 h and the functional validation was carried out subsequently. In vitro, CD276 Dash CAR-T possessed a less-differentiated phenotype and robust proliferative ability compared to conventional CAR-T. In vivo xenograft mouse model, CD276 Dash CAR-T showed enhanced anti-pancreatic cancer efficacy and T cell expansion. Besides, except for the high-dose group, the body weight of mice was maintained stable, and the state of mice was normal.In this study, we proved CD276 CAR-T exhibited powerful activity against pancreatic cancer cells in vitro and in vivo. More importantly, we demonstrated the manufacturing feasibility, acceptable safety and superior anti-tumor efficacy of CD276 Dash CAR-T generated with reduced time. The results of the above studies indicated that CD276 Dash CAR-T immunotherapy might be a novel and promising strategy for pancreatic cancer treatment.© 2024. The Author(s).