核转录因子KLF4激活Wnt/β-catenin通路促进结肠癌肿瘤干细自我更新、转移复发的机制研究

Several studies link cancer treatment failures, metastases and recurrence to the cancer stem cell (CSC) self-renewal capacity. Moreover, Wnt/β-catenin pathway plays an critical role in maintaining self-renewal of CSCs. It was assumed that KLF4 may act as a tumor suppressor and inactivate the Wnt/β-catenin pathway in colon cancers, which based on the bulk cancer cells and clinical research. However, these studies did not uncover the relationship between KLF4 and CSCs. Our preliminary study indicated that in CSCs-enriched spheroid cells, there is a strong relationship between KLF4 and colon CSCs, and KLF4 may be involved in regulation of Wnt/β-catenin signaling pathway. However, the CD133+ subpopulation is about 3.5% but not 100%. More importantly, it has not been demonstrated that whether KLF4 activated Wnt/β-catenin pathway promotes the self-renewal, metastasis and recurrence of colon CSCs..Hence, in this study, we will focus on whether KLF4 regulate colon CSCs and finally involved in cancer progression and how is the relationship between KLF4 and Wnt/β-catenin pathway in purified colon CSCs by flow cytometry, limiting dilution assay, orthotopic transplantation model of colon CSCs, vivo imaging of CSCs, tandem affinity purification technology and surface plasmon resonance technology.

肿瘤干细胞（CSCs）强大的自我更新能力介导了肿瘤的转移复发。Wnt/β-catenin通路在维持CSCs自我更新过程中发挥重要作用。我们前期研究发现在富集结肠癌CSCs（CD133+细胞3.5%）的细胞球中KLF4的表达与细胞球的恶性行为及CSCs呈正相关性，且KLF4可能参与调控Wnt/β-catenin信号通路，提示KLF4在此研究模型中可能为原癌基因，且与CSCs密切相关。但国内外至今尚无发现KLF4与结肠癌CSCs和Wnt/β-catenin信号通路的直接证据。故本课题拟通过流式细胞术分选出纯化的结肠癌CSCs，并在CSCs、CSCs结肠癌原位移植瘤模型、药物诱导结肠癌裸鼠模型中上/下调KLF4的表达，再阻断/激活Wnt/β-catenin通路，采用多种生物学技术研究确认在CSCs中KLF4是否通过激活Wnt/β-catenin通路从而促进CSCs的自我更新、转移复发及具体机制。

核转录因子KLF4在结直肠癌（CRC）中被认为是抑癌基因，但我们前期发现在富集CRC肿瘤干细胞（CSCs）的细胞球中可能通过Wnt通路发挥原癌基因作用，但具体机制不明。本研究进一步在流式细胞术分选的ALDH+的CSCs中下调KLF4表达，检测细胞恶性行为变化及Wnt通路变化情况。我们首次发现KLF4可能通过Snail/EMT影响细胞侵袭迁移能力，可能通过Snail/E-cadherin/Wnt通路影响细胞自我更新、增殖、成瘤能力。本研究的完成证明了KLF4可能是能够同时调控CSCs自我更新、增殖、成瘤及侵袭转移的上游基因，为靶向CSCs的治疗提供了新的靶点。