转录因子YB-1与PD-L1正反馈互作促进乳腺癌转移的机制研究

The occurrence of EMT in cancer cells can always accompanied with immune escape, but the molecular mechanism is not clear. It is known that YB-1 and PD-L1 are involved in the regulation of EMT and immune escape, respectively. Recently, we have obtained preliminary results that YB-1 and PD-L1 were co-expressed in metastatic breast cancer cells, and knockdown of PD-L1 can down-regulate the phosphorylation of PI3K/AKT pathway and YB-1. Moreover, interference with YB-1 could inhibited the expression of PD-L1 and enhanced the sensitivity of breast cancer cells to T cells. Sequence analysis revealed a potential binding site of YB-1 on 5 '-UTR of PD-L1 mRNA, suggesting that YB-1 may involved in the regulation of PD-L1 translation process. The above data have led to the formation of our central hypothesis of this project, there is positive feedback interaction between YB-1 and PD-L1, which jointly promote EMT and immune escape of breast cancer cells and enhance the metastasis of breast cancer. To test our central hypothesis, we have planned the following experiments: 1) study the relationship between YB-1 and PD-L1; 2) confirmed that PD-L1 could activate YB-1 through the PI3K/AKT pathway and promote EMT; 3) proved that YB-1 could feedback regulate the expression of PD-L1 and promote immune escape. This study will provide scientific basis for elucidation the molecular mechanism and search for new therapeutic targets of breast cancer metastasis.

肿瘤细胞发生EMT的同时可伴随产生免疫逃逸，但其中的分子机制并不十分清楚。已知YB-1与PD-L1分别参与调控EMT和免疫逃逸，近期我们发现，在转移乳腺癌细胞中YB-1与PD-L1同时高表达，且敲低PD-L1可下调PI3K/AKT通路及下游YB-1的磷酸化；而干扰YB-1可抑制PD-L1的表达并增强乳腺癌细胞对T细胞的敏感性。序列分析发现PD-L1 mRNA的5’-UTR上存在YB-1潜在结合位点，提示YB-1可能参与调控PD-L1的翻译过程。由此推测，YB-1与PD-L1存在正反馈互作，共同促进乳腺癌细胞EMT及免疫逃逸，增强乳腺癌的转移。本项目拟：1）阐明YB-1与PD-L1的表达具有关联性；2）明确PD-L1可通过PI3K/AKT通路激活YB-1并促进EMT；3）探明YB-1可反馈调节PD-L1的表达促进免疫逃逸。本研究为阐明乳腺癌转移的分子机制和寻找新的治疗靶点提供科学依据。

程序性细胞死亡1 (PD-1)通过抑制T细胞受体下游的信号转导，帮助肿瘤细胞逃避免疫反应，从而抑制T效应因子的功能。 然而，细胞内PD-1在癌细胞中的作用和机制尚不清楚。 在本研究中，我们发现PD-1在三阴性乳腺癌（TNBC）患者和细胞系中异常上调。且TNBC中的PD-1可以在体外和体内促进乳腺癌的生长和转移。进一步的研究表明，PD-1对TNBC细胞生长的影响依赖于细胞本征-PD-1/PD-L1通路，而不依赖于适应性免疫。 此外，我们进一步发现TNBC细胞内PD-1/PD-L1通路的激活受基因表达调节剂YB-1的调控。 从机制上看，蛋白质降解分析、mRNA翻译活性分析、CLICK化学和l-叠氮多聚丙氨酸(AHA)入组分析、免疫沉淀试验和双荧光素酶报告基因试验结果表明，YB-1通过翻译激活途径促进PD-1的表达。 我们提供的体外和体内证据表明，沉默YB-1在TNBC细胞中的表达可以抑制细胞增殖、肿瘤发生和转移。 然而，这种抑制可以通过同时外源性表达PD-1和PD-L1蛋白来挽救。综上，我们的研究结果确定了PD-1/PD-L1轴在肿瘤生长和转移中的TNBC细胞固有功能; 研究发现PD-1/PD-L1是YB -1介导的TNBC增殖和转移的重要效应因子。