盐霉素诱导转移性卵巢癌细胞的间充质-上皮性转换及其机理研究

Since epithelial-mesenchymal transition (EMT) of cancer cells located at the periphery of primary tumors is essential to ovarian cancer metastasis, searching drugs that can block this process has potential clinical significance. Salinomycin has been proved to be toxic to mesenchymal-like cells selectively in many types of cancers. Our preliminary work found that salinomycin induced the expression of epithelial markers in mesenchymal-like ovarian cells while down-regulating the expression of LEF1, Fibronectin, Cyclin D1, all of which are downstream molecules of Wnt pathway. However, the influence of salinomycin on the EMT of ovarian cancer is still unclear. Our study plans to use different metastatic ovarian cancer cells to elucidate: ①whether salinomycin could inhibit the EMT of ovarian cancer cells and then induce their MET; ②whether salinomycin could block the invasion and metastasis of ovarian cancer cells in vitro and in vivo; ③whether Wnt/Fzd/LRP complex is the important and specific target of salinomycin. We try to prove: salinomycin, inducing MET and inhibiting metastatic ability of ovarian cancer cells through blockage of Wnt/Fzd/LRP complex, could be a new promising drug for the treatment of metastatic ovarian cancers.

位于卵巢癌原发灶边缘处的癌症细胞所发生的上皮-间充质转换（EMT）是卵巢癌转移的关键步骤，因此，寻找阻断EMT的药物具有潜在的临床意义。盐霉素被证实在多种癌症中选择性靶向间充质样干细胞。本研究前期工作发现：盐霉素在间充质样卵巢癌细胞中诱导了上皮性分子的表达并下调了Wnt通路的下游分子- - LEF1，Fibronectin及Cyclin D1。但盐霉素对卵巢癌EMT的影响及其作用机理尚不清楚。因此，本研究拟用不同转移能力的卵巢癌细胞为研究对象，探讨①盐霉素是否具有抑制卵巢癌细胞EMT并进而发生MET的能力；②盐霉素是否可以抑制卵巢癌细胞体内及体外的侵袭及转移；③Wnt/Fzd/LRP复合体是否是盐霉素发挥该作用的重要而特异性的靶点。尝试证实：盐霉素通过抑制Wnt/Fzd/LRP复合体诱导转移性卵巢癌细胞的MET、影响其侵袭转移能力，从而为转移性卵巢癌的治疗提供一个新的有意义的药物。

盐霉素是一种单羧酸聚醚类抗生素，被广泛的应用于鸡的抗球虫病治疗当中。在乳腺癌的研究中发现盐霉素可以靶向杀伤乳腺癌干细胞。而上皮间充质转换（epithelial-mesenchymal transiton, EMT）是一系列癌症细胞获得干性的重要机制。因此，盐霉素对乳腺癌干细胞的抑制作用很可能是通过抑制EMT来实现的。本课题通过体外实验探讨盐霉素对上皮性卵巢癌细胞侵袭和转移能力的影响及其作用机制。结果发现：用盐霉素处理上皮性卵巢癌细胞A2780和SK-OV-3之后，细胞的上皮性标志物E-cadherin和Keratin的表达明显上调，而间充质性标志物N-cadherin和Vimentin的表达明显下调。同时，细胞的增殖、迁移和侵袭能力均受到明显的抑制。这提示盐霉素能够抑制上皮性卵巢癌细胞的EMT。通过对Wnt/β-catenin信号通路的相关分子的表达进行检测，我们也探讨了盐霉素可能的作用机制。我们发现在盐霉素处理后，卵巢癌细胞中Wnt/β-catenin信号通路相关分子及Wnt靶基因的表达均明显下调。而且免疫荧光检测发现盐霉素可以抑制β-catenin向胞核内的迁移。使用GSK-3β抑制剂进一步反向验证了盐霉素能够抑制Wnt/β-catenin信号通路。本课题证实了盐霉素可以通过抑制上皮性卵巢癌细胞发生EMT而抑制其迁移和侵袭能力，而盐霉素抑制上皮性卵巢癌细胞发生EMT主要是通过抑制Wnt/β-catenin信号通路来完成的。这为卵巢癌转移的治疗提供了重要的实验基础。