The Role And Mechanism Of Transcription Factor SOX6 In Clear Cell Renal Cell Carcinoma

Objective: Renal cell carcinoma (RCC) ranks the third most common urological neoplasms and accounts for approximately 3% of all malignancies. Of all the subtypes of RCC, clear cell renal cell carcinoma (ccRCC) occupies the most common seen subtype (80～90%) and is insensitive to chemotherapy and radiotherapy. Patients with late stage or metastatic ccRCC usually have a not promising prognosis, and prognostic predictors and therapeutic targets are urgent needed. Sry-related high-mobility-group box 6 (SOX6), a main member of SOXs, has been reported to involve in the regulation of cell fate and differentiation. Accumulating evidences have demonstrated that SOX6 is involved in regulation of carcinogenesis in various human malignancies, however, there is still no study that focused on SOX6 in ccRCC.Thus, this study aims to investigate the role and mechanism of SOX6 in ccRCC.Methods: Real-time polymerase chain reaction (PCR) and western blot assay were applied to detect the SOX6 expression in ccRCC tissues and correspondence adjacent tissues, and also in ccRCC cell lines (786-0, 769-P, OS-RC-2, and Caki-2) and human proximal tubule epithelial cell line (HK-2). Overexpression and knockdown of SOX6 were used to investigate the influence of SOX6 on cell proliferation and cell cycle of ccRCC in vitro and in vivo. SOX6AHMG plasmid, namely the high-mobility-group (HMG) domain was deleted in SOX6 plasmid, was constructed to investigate the role of HMG-domain in biological function of SOX6 in ccRCC. Luciferase assay, immunofluorescence and western blot assay were applied to detect the influence of SOX6 on the Wnt/β-catenin signaling pathway and its downstream target genes. Immunohistochemistry staining and survival analysis were used to analyze the association between SOX6 expression and clinicopathologic parameters and oncologic outcomes.Results: Compared with correspondence adjacent tissue, SOX6 expression levels were significantly downregulated in ccRCC tissue. Besides, the expression of SOX6 in ccRCC cell lines was lower than HK-2 cell line. Overexpressing SOX6 could significantly inhibit cell proliferation and G1/S transition in ccRCC cell lines, and vice versa. The further in vivo assay confirmed the xenograft tumor of SOX6 overexpressed slower growth, smaller, and lower weight than the control. The HMG-domain was crucial for the SOX6-mediated tumor suppression. When the HMG-domain was deleted, the ability of SOX6 in suppressing tumor growth was disappeared. SOX6 had a significant inhibitory effect on Wnt/β-catenin signaling and its downstream target genes (cMyc and Cyclin D1) in vitro and in vivo, which was in an HMG-domain dependent manner. The upregulation of SOX6 was significant associated with smaller tumor sizes, early tumor stage, and lower Fuhrman grades. Survival analysis suggested that patients with low SOX6 expression had a poorer cancer-specific survival than those with high SOX6 expression, and SOX6 could act as an independent prognostic factor for ccRCC.Conclusions: The present study firstly indicated that SOX6 acts as a novel tumor suppressor in ccRCC. Here, we demonstrated that SOX6 could inhibit tumor growth in vitro and in vivo by negatively regulating the Wnt/β-catenin signaling pathway in an HMG-domain dependent manner. The SOX6 levels significantly distinguished the survival of patients who underwent nephrectomy and provided an independent prognostic value in ccRCC. These findings might facilitate the understanding of the molecular mechanisms in human ccRCC pathogenesis and carry potential therapeutic implication.