氧化磷酸化促进休眠膀胱癌细胞表达PD-L1介导免疫逃逸的机制研究

Dormant tumor cells can escape immune killing, contributing to tumor recurrence and metastasis, but the mechanism underlying dormant tumor cells immune tolerance is not clear. Our previous studies showed that the immunocheckpoint protein PD-L1 was up-regulated in dormant bladder cancer cells. These dormant cells showed enhanced oxidative phosphorylation (OXPHOS) metabolism and inhibition of OXPHOS could inhibit the gene expression of PD-L1 and moreover, promote PD-L1 protein degradation. More importantly, inhibition of OXPHOS enhanced the killing effect of T cells on dormant bladder cancer cells. This project aims to investigate the effect of OXPHOS on the expression of PD-L1 in dormant bladder cancer cells, and to explore the reversal effect of inhibiting OXPHOS on dormant bladder cancer cells immune tolerance in vitro and in vivo, with following specific mechanism focus: (1) by investigating the activating effect of OXPHOS on Bmp4-SMAD-IRF-1 signaling pathway, to clarify the promoting effect of OXPHOS on PD-L1 gene expression in dormant bladder cancer cells; (2) through investigating the effect of OXPHOS on endoplasmic reticulum-related degradation (ERAD) pathway, to clarify the inhibitory effect of OXPHOS on ERAD-mediated PD-L1 degradation. The completion of this project will further clarify the mechanism of immune escape of dormant bladder cancer cells, provide a theoretical basis for the effective elimination of dormant bladder cancer cells and help to succeed in prevention of tumor recurrence and metastasis.

休眠肿瘤细胞能够耐受免疫杀伤，是肿瘤复发和转移的根源，但其免疫耐受机制还不明确。我们前期研究发现休眠膀胱癌细胞PD-L1表达增高，且细胞的氧化磷酸化（OXPHOS）显著增强，而抑制OXPHOS既能抑制PD-L1表达，又可以促进PD-L1降解，增强T细胞对休眠膀胱癌细胞的杀伤。本项目拟在前期基础上，通过体内外研究进一步明确休眠膀胱癌细胞中OXPHOS对PD-L1表达的促进作用，以及抑制OXPHOS增强T细胞对休眠膀胱癌细胞的杀伤作用。并从以下两方面阐明机制：（1）通过研究OXPHOS对Bmp4-SMAD-IRF-1信号通路的活化，明确OXPHOS促进休眠膀胱癌细胞PD-L1表达的机制；（2）通过研究OXPHOS对内质网相关降解途径的抑制，明确OXPHOS 减少PD-L1降解的机制。本项目的完成将进一步阐明休眠膀胱癌细胞免疫逃逸的机制，为有效清除休眠膀胱癌细胞、防止肿瘤复发和转移提供理论依据

休眠肿瘤细胞能够耐受免疫杀伤，是肿瘤复发和转移的根源，但其免疫耐受机制还不明确。我们前期研究发现休眠膀胱癌细胞（DBC）细胞程序性死亡-配体1(PD-L1)表达增高，且细胞的氧化磷酸化（OXPHOS）显著增强。本项目研究了休眠膀胱癌细胞的代谢特点、自噬及其与PD-L1表达之间的关系,通过细胞培养、细胞转染、Real-time PCR、Western Blot、Seahorse、流式细胞技术、免疫荧光等方法，发现DBC中OXPHOS和自噬增强，并可相互促进，DBC胞内（而非胞膜）PD-L1水平升高；OXPHOS和自噬相互协同，抑制PD-L1降解。胞内PD-L1通过调控衰老相关基因P53、P21表达，抑制DBC衰老，为了明确DBC代谢特点和抗衰老相关分子机制提供了研究数据，提示联合抑制OXPHOS和自噬可能是靶向休眠肿瘤细胞的新思路。