抑癌基因锌指蛋白 471 通过抑制 AKT 和 Wnt/β-连环蛋白信号通路抑制乳腺癌的生长和转移。

BACKGROUND: Zinc-finger protein 471 (ZNF471) is a member of the Krüppel-associated box domain zinc finger protein (KRAB-ZFP) family. ZNF471 is methylated in squamous cell carcinomas of tongue, stomach and esophageal. However, its role in breast carcinogenesis remains elusive. Here, we studied its expression, functions, and molecular mechanisms in breast cancer. METHODS: We examined ZNF471 expression by RT-PCR and qPCR. Methylation-specific PCR determined its promoter methylation. Its biological functions and related molecular mechanisms were assessed by CCK-8, clonogenicity, wound healing, Transwell, nude mice tumorigenicity, flow cytometry, BrdU-ELISA, immunohistochemistry and Western blot assays. RESULTS: ZNF471 was significantly downregulated in breast cell lines and tissues due to its promoter CpG methylation, compared with normal mammary epithelial cells and paired surgical-margin tissues. Ectopic expression of ZNF471 substantially inhibited breast tumor cell growth in vitro and in vivo, arrested cell cycle at S phase, and promoted cell apoptosis, as well as suppressed metastasis. Further knockdown of ZNF471 verified its tumor-suppressive effects. We also found that ZNF471 exerted its tumor-suppressive functions through suppressing epithelial-mesenchymal transition, tumor cell stemness and AKT and Wnt/β-catenin signaling. CONCLUSIONS: ZNF471 functions as a tumor suppressor that was epigenetically inactivated in breast cancer. Its inhibition of AKT and Wnt/β-catenin signaling pathways is one of the mechanisms underlying its anti-cancer effects.

背景：锌指蛋白 471（ZNF471）是 Krüppel 相关盒结构域锌指蛋白（KRAB-ZFP）家族的成员。ZNF471 在舌、胃和食管鳞状细胞癌中发生甲基化。然而，其在乳腺癌发生中的作用仍不清楚。在这里，我们研究了其在乳腺癌中的表达、功能和分子机制。

方法：我们通过 RT-PCR 和 qPCR 检查 ZNF471 的表达。甲基化特异性 PCR 确定其启动子甲基化。通过 CCK-8、集落形成、划痕愈合、Transwell、裸鼠致瘤性、流式细胞术、BrdU-ELISA、免疫组织化学和 Western blot 检测评估其生物学功能和相关分子机制。

结果：与正常乳腺上皮细胞和配对的手术切缘组织相比，由于其启动子 CpG 甲基化，ZNF471 在乳腺癌细胞系和组织中表达显著下调。ZNF471 的异位表达显著抑制了乳腺癌细胞在体外和体内的生长，将细胞周期阻滞在 S 期，并促进细胞凋亡，同时抑制转移。进一步敲低 ZNF471 验证了其肿瘤抑制作用。我们还发现，ZNF471 通过抑制上皮-间充质转化、肿瘤细胞干性和 AKT 和 Wnt/β-catenin 信号通路发挥其肿瘤抑制功能。

结论：ZNF471 作为一种肿瘤抑制因子，在乳腺癌中被表观遗传失活。其对 AKT 和 Wnt/β-catenin 信号通路的抑制是其抗癌作用的机制之一。