免疫检验点分子PD-1沉默的CART19细胞的抗肿瘤功能及机制研究

Cancers figure among the leading causes of morbidity and mortality worldwide. In recent years, CART cells have shown big potential in the field of tumor therapy. Especially in B cell malignancies, CART19 cells have proved their efficacy. However, in patients with malignancies, immune checkpoint molecule PD-1 was expressed upon T cells, which could lead to the immune anergy of T cells. Our previous research found that, 30% of the cultured CART19 cells expressed PD-1, after co-culture with tumor cells, the rate was even over 45%. Thus, it is probable that the CART19 cells reinfused into patients could turn into anergy because of the inhibition signal transduced by PD-1. The intercept of the PD-1 signal might release CART19 cells from the inhibition, and enhance the anti-tumor activity of CART19 cells. Thus, we aim to create PD-1 silenced CART19 cells, perform multi-layered reseaches to reveal its anti-tumor function and mechanism, in order to illustrating the regulation of PD-1 pathway in CART19 cells, and providing PD-1 silenced CART19 cells as a new potential drug for cancer therapy.

癌症是人类发病及死亡的主要原因之一，近年来多项临床试验证明了CART细胞在肿瘤治疗领域的良好前景。尤其是在B细胞肿瘤中，CART19细胞显示了一定的抗肿瘤作用。然而，在肿瘤病人中，免疫检验点分子PD-1常表达于T细胞表面，导致T细胞免疫耐受。我们的前期实验发现，培养获得的CART19细胞PD-1表达率约为30%，与肿瘤细胞共培养后，表达率＞45%。这提示我们，回输的CART19细胞在病人体内可能受到PD-1的负调控，导致部分免疫耐受。因此，抑制该信号可能解除肿瘤细胞对CART19细胞的负调控，增强CART19细胞抗肿瘤活性。因此，我们拟制备一种免疫检验点分子PD-1沉默的CART19细胞，并从分子、细胞、动物模型以及临床病理样本多个层次对其抗肿瘤功能及机制进行研究，阐明PD-1通路在CART19细胞中的调控作用，为临床提供一种新型的免疫检验点分子PD-1沉默的CART19细胞潜力药物。

阻断程序性死亡受体-1（PD-1）被认为是改善T细胞功能的有效策略，并且许多进行的临床试验正在对其探索。然而，PD-1阻断带来的效果是否持久并非是定论。为了确定持久的PD-1阻断对T细胞功能的影响，我们构建了PD-1稳定阻断的嵌合抗原受体修饰的T（CART）细胞，并研究了它们的抗肿瘤功能以及相关特性。根据短期体外结果，我们再次证实了PD-1阻断可以增强对程序性死亡 - 配体1（PD-L1）介导的免疫抑制的抵抗。然而，PD-1被稳定阻断的CART细胞在体外或体内实验中未呈现更好的抗肿瘤功能。我们发现，持久的PD-1阻断可能损害CART细胞的抗肿瘤潜力通过抑制T细胞的增殖活性。此外，我们观察到PD-1阻断会加速T细胞的早期分化并削弱其持久性。这些结果表明PD-1可能在维持T细胞的正常增殖和分化中起重要作用，并且PD-1的长期阻断将通过抑制增殖活性而损害T细胞的抗肿瘤功能。