OncoTargets and Therapy (Nov 2019)
CLK3 Is A Direct Target Of miR-144 And Contributes To Aggressive Progression In Hepatocellular Carcinoma
Abstract
Hua Li,1,* Xichun Cui,2,* Qiuyue Hu,2 Xiaolong Chen,2 Pengli Zhou3 1Department of Infectious Diseases, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, People’s Republic of China; 2Key Laboratory of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, People’s Republic of China; 3Department of Intervention, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450014, People’s Republic of China*These authors contributed equally to this workCorrespondence: Pengli ZhouDepartment of Intervention, The First Affiliated Hospital of Zhengzhou University, 1 Jianshe East Road, Erqi, Zhengzhou 450052, People’s Republic of ChinaEmail [email protected]: Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer with high incidence. The underlying molecular mechanisms of HCC development have been intensively studied. CLK3 (CDC Like Kinase 3) is a nuclear dual-specificity kinase and regulates gene splicing. We investigated the expression profile and functional role of CLK3 in HCC.Methods: Immunohistochemistry (IHC) and Western blot were performed to determine CLK3 expression in HCC tissues. Bioinformatics analysis using TCGA and GEO database was conducted to evaluate the relationship between CLK3 expression and HCC prognosis. Cell proliferation was assessed by CCK8, EdU and colony formation assays, while transwell and wound-healing assays were performed to investigate the cell migration and invasion in vitro. Xenograft nude mouse model was used to test the function of CLK3 on tumor growth in vivo. Luciferase reporter assay, Western blot and RT-qPCR were conducted to verify the miRNA that directly targeted CLK3.Results: CLK3 was markedly upregulated in HCC tissues, and the expression levels of CLK3 were closely associated with TNM stages and HCC prognosis. Functional analysis indicated that knockdown of CLK3 could suppress HCC cell growth, invasion and migration in vitro, and inhibit tumor development in vivo. Moreover, CLK3 was demonstrated as a direct target of miR-144 and miR-144 expression was inversely correlated with CLK3 expression in HCC. Enforced overexpression of miR-144 markedly inhibited the CLK3 expression while overexpression of CLK3 partially reversed the inhibitory function of miR-144 on HCC cell growth and metastasis. Mechanistically, we found that miR-144 overexpression inhibited Wnt/β-catenin signaling and the inhibition could be partly abolished by overexpression of CLK3.Conclusion: In summary, we demonstrate tumor suppressor miR-144 suppresses hepatocellular carcinoma development and metastasis via regulating CLK3 and Wnt/β-catenin signaling, indicating that miR-144/CLK3 could be used for HCC diagnosis and treatment.Keywords: miR-144, metastasis, HCC, CLK3, Wnt/β-catenin signaling
