SIRT1在结直肠癌干细胞干性维持中的作用及机制研究

Cancer stem cells (CSCs) play a crucial role in tumorigenesis, tumor development and tumor resistance to radiotherapy and chemotherapy. The mechanism studies of regulation of CSCs will contribute to the tumor prevention and therapy. In previous study, we found that SIRT1 was high expressed in colorectal cancer (CRC) tissue comparing with the pericarcinomatous tissue. Meanwhile, SIRT1 had a high expression in the colorectal CSCs with CD133 expression. In vivo and vitro studies showed that SIRT1 deficiency attenuated the stemness of CSCs. However, the mechanisms of this phenomenon are still unclear. Based on our previous research and the latest associated research progress, we attempt to carry out the following research proposal. 1. Firstly, by regulating SIRT1 expression via the gene over-expression and shRNA technologies, we will further confirm the influence of SIRT1 on stemness of CSCs. 2. Then we will screen the candidate stemness-associated pathways and thereby find out the SIRT1-regulated pathway and key molecules in stemness maintenance of colorectal CSCs. 3. We will further regulate the expressions of SIRT1 and screened factors to confirm the molecule mechanisms of SIRT1 in keeping the stemness of colorectal CSCs. 4. Finally, we will detect the expressions of SIRT1 and stemness-associated proteins regulated by SIRT1 in clinical CRC tissues. Implying the analysis of clinical and pathological data, the correlation between the expression of corresponding genes and the prognosis of CRC patients will be explored. This research will contribute to understanding the mechanisms of SIRT1 in keeping the stemness of colorectal CSCs, which provides a promising new target in CRC therapy and associated theoretical basis.

肿瘤干细胞（CSCs）被认为是肿瘤形成、发生发展乃至放化疗抵抗的根源，加深对其调控方式的研究将极大的推动肿瘤防治。我们前期研究发现，SIRT1在结直肠癌组织中表达高于癌旁，且在CD133+结直肠CSCs中高表达。体内外研究显示抑制SIRT1会降低CSCs干性，但具体机制仍不明确。本课题拟在此基础上，用过表达及shRNA技术调控SIRT1表达，进一步验证SIRT1对结直肠CSCs干性的影响；再筛选CSCs中受SIRT1调控的干性相关通路及其中显著变化的关键分子及调控方式；然后通过调控SIRT1及筛选出的关键分子表达，明确SIRT1调控结直肠CSCs干性的分子机制；最后在结直肠癌临床样本中验证SIRT1与其调控的干性相关蛋白表达的相关性，并结合临床资料分析与患者预后的关系。本研究有助于阐明SIRT1在结直肠CSCs调控中作用机制，为结直肠癌临床治疗提供理论依据和有前景的新的治疗靶标。

肿瘤干细胞CSCs的存在被认为是肿瘤难以治愈、易复发、发生转移的最主要的原因。前期研究发现，SIRT1在结直肠癌组织中高表达，且在肿瘤干细胞样细胞中表达明显升高。体内外研究显示抑制SIRT1会明显降低CSCs干性，为了进一步研究SIRT1结直肠癌肿瘤干细胞干性维持中的作用及分子机制，我们开展本课题。研究发现，SIRT1在原代富集的结直肠癌细胞干细胞样细胞中高表达，且抑制SIRT1可明显抑制HCT116细胞的体内成瘤能力。我们还发现，SIRT1在67%的结直肠癌转移病人中高表达，并且伴随着明显加快的转移发生时间。体内实验发现，调节SIRT1的活性能够显著控制结直肠癌细胞肝转移灶点的数目和大小。同时，抑制SIRT1表达，可以显著降低结直肠癌细胞CT26和SW620的EMT及侵袭迁移能力，过表达SIRT1可以明显增加SW480细胞的运动和迁移能力。进一步研究发现，mTORC1/4E-BP1是关键的调控轴线，抑制mTORC1可以逆转由SIRT1过表达引起的侵袭迁移能力增加。但同时我们发现，mTORC1/4E-BP1对结直肠癌侵袭迁移的调控作用并不是通过影响肿瘤干细胞干性发挥作用的。我们的研究发现，抑制WNT/β-catenin信号通路可以逆转由过表达SIRT1引起的结直肠癌干性增加，提示WNT/β-catenin才是SIRT1调控结直肠癌肿瘤干细胞干性的关键通路。本研究明确了SIRT1在调控结直肠癌肿瘤干细胞干性维持中的关键信号通路，同时阐明SIRT1分子与结直肠癌转移的关系及分子机制，为结直肠癌的临床治疗靶点提供理论基础。