The Mary & John Knight Translational Ovarian Cancer Research Unit, London Regional Cancer Program, London, Ontario, Canada.
Department of Anatomy & Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, Ontario, Canada.
Mol Cancer Res. 2020 Mar;18(3):488-500. doi: 10.1158/1541-7786.MCR-19-0530. Epub 2019 Nov 19.
Epithelial ovarian cancer (EOC) spreads by direct dissemination of malignant cells and multicellular clusters, known as spheroids, into the peritoneum followed by implantation and growth on abdominal surfaces. Using a spheroid model system of EOC metastasis, we discovered that Liver kinase B1 (LKB1), encoded by the gene, and its canonical substrate AMP-activated protein kinase (AMPK) are activated in EOC spheroids, yet only LKB1 is required for cell survival. We have now generated -knockout cell lines using normal human FT190 cells and three EOC cell lines, OVCAR8, HeyA8, and iOvCa147. KO did not affect growth and viability in adherent culture, but it decreased anchorage-independent growth of EOC cells. EOC spheroids lacking LKB1 had markedly impaired growth and viability, whereas there was no difference in normal FT190 spheroids. To test whether LKB1 loss affects EOC metastasis, we performed intraperitoneal injections of OVCAR8-, HeyA8-, and iOvCa147-KO cells, and respective controls. LKB1 loss exhibited a dramatic reduction on tumor burden and metastatic potential; in particular, OVCAR8-KO tumors had evidence of extensive necrosis, apoptosis, and hypoxia. Interestingly, LKB1 loss did not affect AMPKα phosphorylation in EOC spheroids and tumor xenografts, indicating that LKB1 signaling to support EOC cell survival in spheroids and metastatic tumor growth occurs via other downstream mediators. We identified the dual-specificity phosphatase DUSP4 as a commonly upregulated protein due to LKB1 loss; indeed, knockdown in HeyA8-KO cells partially restored spheroid formation and viability. IMPLICATIONS: LKB1 possesses key tumor-promoting activity independent of downstream AMPK signaling during EOC metastasis.
上皮性卵巢癌 (EOC) 通过恶性细胞和多细胞簇（称为球体）的直接扩散传播到腹膜中，然后在腹部表面种植和生长。我们使用 EOC 转移的球体模型系统发现，基因编码的肝激酶 B1 (LKB1)及其典型底物 AMP 激活的蛋白激酶 (AMPK) 在 EOC 球体中被激活，但只有 LKB1 是细胞存活所必需的。我们现在使用正常的人 FT190 细胞和三种 EOC 细胞系 OVCAR8、HeyA8 和 iOvCa147 生成了 -敲除细胞系。 KO 对贴壁培养中的生长和活力没有影响，但它降低了 EOC 细胞的无锚定依赖性生长。缺乏 LKB1 的 EOC 球体的生长和活力明显受损，而正常的 FT190 球体则没有差异。为了测试 LKB1 缺失是否影响 EOC 转移，我们进行了 OVCAR8、HeyA8 和 iOvCa147-KO 细胞及其相应对照的腹腔内注射。 LKB1 缺失表现出肿瘤负担和转移潜力的急剧降低；特别是，OVCAR8-KO 肿瘤有广泛坏死、凋亡和缺氧的证据。有趣的是，LKB1 缺失不影响 EOC 球体和肿瘤异种移植物中的 AMPKα 磷酸化，表明 LKB1 信号支持 EOC 细胞在球体和转移性肿瘤生长中的存活是通过其他下游介质发生的。我们发现双特异性磷酸酶 DUSP4 是由于 LKB1 缺失而普遍上调的蛋白质之一；事实上，在 HeyA8-KO 细胞中敲低部分恢复了球体的形成和活力。意义：在 EOC 转移过程中，LKB1 具有独立于下游 AMPK 信号的关键促肿瘤活性。
