ΔNp63α在未折叠蛋白反应介导的肿瘤转移中的作用和机制研究

Malignant tumor metastasis is the main cause of death in people with cancer, understanding the regulation mechanism of tumor metastasis and the role of key proteins have important theoretical and clinical significance. It has been reported that unfolded protein response (UPR) is an important cause of tumor metastasis, however, its downstream target proteins and their signaling remain unclear. Our previous studies have shown that p53 family member ΔNp63α plays a critical role in inhibiting tumor metastasis. Oncogenic signaling pathways including RAS, PI3K and HER2 promote the metastasis of malignant tumors by inhibiting the transcriptional expression of ΔNp63α. In this project, clinical database analysis revealed that ΔNp63α gene expression was negatively correlated with GRP78, the master regulator of UPR. The expression of ΔNp63α was upregulated after knockdown of IRE1α or ATF6α, or suppression of IRE1α/XBP1 and ATF6α by small molecular inhibitor, resulted in cell migration and invasion were suppressed. Restoration of ΔNp63α expression could inhibit XBP1s-mediated tumor lung metastasis. This project aims to elucidate the role and mechanism of ΔNp63α in UPR-mediated breast cancer metastasis, and provide potential targets for breast cancer diagnosis and treatment.

恶性肿瘤转移是导致肿瘤病人死亡的主要原因，深入研究肿瘤转移的调控机制及其关键蛋白具有重要理论意义和临床意义。未折叠蛋白反应（UPR）是肿瘤转移的重要诱因，然而其下游关键作用蛋白及其调控通路并不完全清楚。我们前期研究表明：p53家族成员ΔNp63α是抑制肿瘤转移的重要蛋白，RAS、PI3K和HER2等关键癌信号通路抑制ΔNp63α表达，进而促进恶性肿瘤的转移。本项目前期工作证明，临床数据中UPR核心调节因子GRP78与ΔNp63α基因表达呈显著负相关；激活UPR可下调ΔNp63α促进细胞迁移；敲低UPR关键蛋白IRE1α或ATF6α，或抑制IRE1α/XBP1和ATF6α通路可上调ΔNp63α蛋白表达，导致细胞迁移和侵袭能力下降；回补ΔNp63α可明显抑制XBP1s介导的乳腺癌肺转移。本项目拟阐明ΔNp63α在UPR介导的乳腺癌转移中作用及机制，为临床乳腺癌诊治提供潜在靶点。

肿瘤快速生长需要消耗大量氧气，实体瘤内部由于血管的缺乏导致局部缺氧而呈低氧状态。然而，低氧促进肿瘤转移的分子机制还不完全清楚。ΔNp63α是细胞-细胞或细胞-基质粘附中的重要调节因子，我们前期研究证明ΔNp63α是重要的肿瘤转移抑制因子。本研究中，我们发现低氧下调ΔNp63α的表达进而促进细胞迁移，而异源表达ΔNp63α抑制低氧诱导的细胞迁移。低氧通过增加H3K27me3修饰抑制ΔNp63α的表达，而该过程不依赖HIF1α。进一步研究发现低氧通过激活非折叠蛋白反应（unfold protein response, UPR）中的IRE1α-XBP1s增加H3K27me3修饰而抑制ΔNp63α的表达促进细胞迁移。已有研究报道H3K27me3修饰和H3K27ac修饰在基因表达中起相反的作用。我们发现低氧通过HDAC2下调H3K27ac修饰抑制ΔNp63α的表达。此外，我们发现低氧促进XBP1s蛋白通过其C末端与HDAC2和EZH2直接相互作用形成阻遏复合物；XBP1s直接与ΔNp63启动子结合，导致ΔNp63启动子上H3K27ac修饰减少而H3K27me3修饰增加，从而抑制ΔNp63α表达。抑制EZH2活性或敲减EZH2的表达均能阻断H3K27me3的修饰，上调ΔNp63α表达，进而抑制低氧或XBP1s诱导的细胞迁移。抑制HDAC2的活性或敲减HDAC2的表达均可上调ΔNp63α的表达，从而抑制低氧或XBP1s诱导的细胞迁移。临床数据库分析发现人乳腺癌中，p63的mRNA水平与XBP1，HDAC2或EZH2的mRNA水平呈负相关。人乳腺癌组织芯片中p63的蛋白水平与XBP1s和H3K27me3呈负相关。此外，在人乳腺癌患者中，p63低表达或XBP1高表达的乳腺癌患者总生存期较差。总之，这些结果揭示了XBP1s/HDAC2/EZH2-ΔNp63α轴在缺氧诱导的细胞迁移和肿瘤转移中的重要作用。