NPM1通过调控PD-L1表达抑制T细胞免疫活性和促进三阴乳腺癌发生发展的分子机制

Triple-negative breast cancer (TNBC) is characterized by its aggressive clinical behavior, and the lack of effective targeted agents. Anti-PD-1/ PD-L1 immunotherapy is a promising strategy for treatment of TNBC. Our previous research showed that expression of PD-L1 was higher in TNBC than other subtypes. Biotin-Streptavidin Agarose Pulldown assay discovered that NPM1 bound to PD-L1 promoter specifically in TNBC cells and activated PD-L1 transcription, thus inhibiting T cell activity in vitro. However, the molecular mechanism by which NPM1 regulates PD-L1 expression, tumor immune and TNBC growth remains unclear. Based on our primary results, we will further investigate the immune regulation activity of NPM1 in vivo by co-IP, protein mass spectrometry and ChIP assays. The prognostic value of NPM1 and its correlation with PD-L1 will be verified in patient tissues and clinical data. Together, this study is aimed to illuminate NPM1 as a novel transcription regulator of PD-L1 and a potential therapeutic target in TNBC.

三阴型乳腺癌侵袭性强、易复发转移、缺少有效的治疗靶点，预后差。近年来，PD-1/PD-L1单抗成为三阴型乳腺癌免疫治疗的新方式。前期研究我们证明PD-L1在三阴型乳腺癌中表达显著升高。通过生物素-链霉亲和素垂钓技术我们鉴定出三阴型乳腺癌细胞株中PD-L1的启动子结合蛋白NPM1，并证明NPM1可激活PD-L1转录及表达，从而抑制T细胞活性。但NPM1如何调控PD-L1表达和肿瘤免疫进而影响三阴型乳腺癌生长的分子机制及其临床意义，目前还不清楚。本项目我们拟①建立动物模型，体内证明NPM1对肿瘤免疫的调节作用；②通过co-IP,蛋白质谱，ChIP等方法阐明NPM1调控PD-L1转录的分子机制；③利用临床标本及生存资料，分析NPM1与PD-L1的相关性及其预后价值。本项目旨在阐明NPM1是三阴型乳腺癌中PD-L1的转录调控分子，为NPM1成为三阴型乳腺癌新的治疗靶点提供理论依据。

三阴型乳腺癌侵袭性强、易复发转移，缺少有效的治疗靶点。近年来，PD-1/PD-L1阻断成为三阴型乳腺癌免疫治疗的新方式。但是，PD-L1高表达的分子机理任然不清楚，我们发现PD-L1在三阴型乳腺癌中表达显著升高，并预测不良预后。通过生物素-链霉亲和素垂钓技术我们鉴定出三阴型乳腺癌细胞株中PD-L1的启动子结合蛋白NPM1，并证明NPM1可激活PD-L1转录及表达，从而抑制T细胞活性，促进肿瘤生长和转移。进一步的机制研究发现在TNBC中NPM1可与PARP1互作，这种互作可掩盖NPM1的C末端核酸结合结构域，因此阻碍其与PD-L1启动子的接触，即PARP1通过阻止NPM1结合PD-L1的启动子来抑制PD-L1表达，PARP1抑制剂Olaparib同样通过此机制上调PD-L1的表达。在体内，我们采用Olaparib同期联合PD-L1单克隆抗体治疗BRCA1缺陷的三阴型乳腺癌，发现联合用药方案可明显减小肿瘤体积，疗效优于单药，且Olaparib通过上调PD-L1引起的活性T细胞浸润减少可被PD-L1单克隆抗体逆转。敲减NPM1可以明显抑制肿瘤生长。项目揭示了NPM1与PARP1是三阴型乳腺癌中PD-L1的转录上调控的重要分子。阻断NPM1或者PARP1是一个潜在的治疗靶点，PARP1抑制剂和 PD-L1单克隆抗体联合用药治疗TNBC具有很好的协同效应。这为TNBC或者转移性的BRCA的治疗提供了新的治疗策略。在项目经费的支持下共发表SCI文章1篇。