SIRT1-USP29-Twist1轴诱导EMT调控乳腺癌肿瘤干细胞干性、转移及耐药的研究

转移和耐药是三阴性乳腺癌患者主要致死原因，而EMT是决定肿瘤干细胞干性、转移及耐药的关键环节。我们针对EMT起始阶段的重要转录因子Snail1发现了影响其蛋白稳定的CDK4/6-DUB3-Snail1轴（ Nat Commun, 2017,一作）。而EMT维持阶段的关键转录因子Twist1蛋白稳定性的调控机制尚不清楚。我们的预实验通过质谱鉴定出的去泛素化酶USP29可增加Twist1蛋白的稳定性从而促进EMT，而这一过程受去乙酰化酶SIRT1的调控，因此我们提出工作假设：SIRT1-USP29-Twist1轴诱导EMT从而增进乳腺癌肿瘤干细胞干性、转移及耐药。本课题旨在验证这一假设，并分别通过敲减SIRT1和USP29及特异性小分子抑制剂阻断SIRT1的活性阐明SIRT1-USP29-Twist1轴在EMT和上述EMT依赖性恶性表型的作用,为三阴性乳腺癌的精准干预提供新成药性靶点。

Triple negative breast cancer (TNBC) is the most malignant breast cancer subtype with a poor prognosis. Tumor metastasis and chemotherapeutic resistance is the primary cause of TNBC-related death. Epithelial-mesenchymal transition (EMT) is the central event to control stemness of breast cancer stem cell，metastasis and chemotherapeutic resistance. We previously demonstrated that CDK4/6-mediated activation of DUB3 is essential to deubiquitinate and stabilize Snail1, a key factor for the initiation of EMT. Furthermore,CDK4/6 inhibitor, palbociclib blocks breast cancer metastasis in TNBC model, without affecting primary tumor growth (Nat Commun 2017,first author). However, how to effectively target Twist1, a key factor for the maintenance of EMT in breast cancer remains unclear. In our preliminary study, we demonstrate that the deubiquitinase, USP29 is essential to deubiquitinate, stabilize Twist1 and therefore, promotes EMT and cell migration. Furthermore, the activity of SIRT1 is essential to maintain Twist1 protein level and cell migration through regulating USP29. Based on these results, our working hypothesis is that SIRT1 dependent activation of USP29 promotes EMT, metastasis, chemo-resistance and stemness of cancer stem cells through stabilizing Twist1. Overall, this study will establish the SIRT1-USP29-Twist1 axis as an important regulatory mechanism of stemness of cancer stem cells, metastasis and chemotherapeutic resistance. Knockdown of SIRT1 or the inhibition of SIRT1 by its specific inhibitor will be utilized to block this axis and the effect on metastasis, chemotherapeutic resistance and stemness of cancer stem cells will be investigated in the proposed study, which will provide a rationale for potential therapeutic interventions in the treatment of triple negative breast cancer.