分泌PD-1单链抗体的MesoCAR-T细胞治疗非小细胞肺癌的体内外研究

Secreted PD-1 scFv Enhanced Antitumor Activity of Chimeric Antigen Receptor T Cell Against Non-Small-Cell Lung Cancer

嵌合抗原受体T细胞(Chimeric Antigen Receptor T cell,CAR-T)疗法在血液瘤中的良好疗效未能延续到实体瘤的治疗中。程序性细胞死亡蛋白-1(Programmed Death-1,PD-1)免疫检查点的阻断可缓解肿瘤微环境(Tumor Microenvironment,TME)中CAR-T细胞受到的抑制作用。为了提高疗效,本文将一种分泌型PD-1单链抗体(single-chain variable Fragment,scFv)—E27,与靶向间皮素(Mesothelin,MSLN)的MesoCAR共表达。结果表明,分泌的PD-1 scFv阻断了CAR-T细胞表面的PD-1。在MSLN抗原的反复刺激下,E27-MesoCAR组的T细胞具有更强的细胞增殖能力,细胞的耗竭和凋亡也得到了明显缓解。进一步通过非小细胞肺癌(Non-Small-Cell Lung Cancer,NSCLC)细胞系来源的异种移植(Cell line-Derived Xenograft,CDX)小鼠的体内实验发现,与传统MesoCAR-T细胞相比,E27-MesoCAR-T细胞在体内也表现出更强的增殖能力和肿瘤抑制作用。本实验将全人源化序列应用于针对NSCLC的CAR-T和免疫检查点联合治疗中,避免了潜在的免疫原性,并为克服TME的免疫抑制提供了一种新的策略。

The success of Chimeric antigen receptor(CAR)T cell therapy in hematological malignancies has not extended to solid tumors.Blockade of the PD-1 immune checkpoint alleviates the suppression of CAR-T cells in the tumor microenvironment(TME).Herein,a mesothelin(MSLN)targeted CAR(MesoCAR)was co-expressed with secreted PD-1 single-chain variable fragments(scFv)E27.Our results showed that the secreted PD-1 scFv blocked PD-1 on the surface of CAR-T cells.After repeated stimulation with MSLN antigen,E27-MesoCAR-T cells were more proliferative,and the exhaustion and apoptosis of CAR-T cells were significantly alleviated.Furthermore,using a cell line-derived xenograft(CDX)NSG mouse model of non-small-cell lung cancer(NSCLC),we determined that compared with conventional MesoCAR-T cells,E27-MesoCAR-T cells showed superior tumorsuppressions because of increased expansion.This is the first time that fully human anti-MSLN CAR and humanized PD-1 scFv have been applied to the combination therapy of checkpoint and CAR-T cells for NSCLC to avoid potential immunogenicity.Our work provides a new strategy for overcoming the immunosuppression of TME.