Downregulation of SNX5 By KLF9 Leads to Clear Cell Renal Cell Carcinoma Progression By Inducing CD44 Internalization and Suppressing Epithelial-to-Mesenchymal Transition

Abstract Background: Aberrant expression of SNX5 can contribute to tumourigenesis, invasion, and metastasis of several human cancers. However, the clinicopathological and biological significance of SNX5 in clear cell renal cell carcinoma (ccRCC) remain unclear. The aim of this study was to examine the role of SNX5 in the progression of ccRCC.Methods: Immunohistochemical (IHC), Western blot, qRT-PCR, western blot, flow cytometry and immunofluorescence were used to detect the expression of indicated molecules. The biological role of SNX5 in ccRCC cells was evaluated by CCK8, colony formation, transwell assay, subcutaneous tumor formation as well as veil tail injection. ChIP assay and luciferase reporter assay were used to determine the direct binding of KLF9 to the promoter of the SNX5 gene.Results: SNX5 expression was downregulated in human ccRCC tissues. SNX5 expression was negatively correlated with tumor size, AJCC stage, tumor thrombus of inferior vena cava (IVC) and poor prognosis of ccRCC. Ectopic expression of SNX5 inhibited ccRCC cell proliferation and metastasis whereas knockdown of SNX5 increase these activities both in vitro and in vivo. Mechanistically, overexpression of SNX5 blocked internalization and intracellular trafficking of CD44 in ccRCC cells. Exogenous expression of CD44 partially rescued the inhibitory effects of SNX5 on the proliferation and invasion activity of ccRCC cells. Knockdown of SNX5 in ccRCC cells was associated with epithelial mesenchymal transition (EMT), including the down-regulation of E-cadherin, ZO-1 and Claudin-1 and the concomitant up-regulation of Snail and N-cadherin. In addition, SNX5 inhibited TGF-β-induced migration, invasion and EMT in ccRCC cells. Moreover, we observed a significant correlation between SNX5 expression and E-cadherin levels in ccRCC patients. In addition, KLF9 directly bound to the SNX5 promoter and increased SNX5 transcription. SNX5 expression was closely correlated with KLF9 expression in ccRCC. Moreover, we found that the combination of SNX5 and CD44 or E-cadherin or KLF9 was a more powerful predictor of poor prognosis than either parameter alone.Conclusion: Collectively, our data reveal a mechanism that KLF9-mediated SNX5 expression was associated with poor prognosis via trafficking of CD44 and promoting EMT in ccRCC. SNX5 may be a potential prognostic biomarker and therapeutic target for patients with ccRCC.