The Functional Roles And Clinical Significance Of CircCCDC66 In Gastric Cancer

Objective:Gastric cancer(GC)is a kind of malignant tumor with the fourth morbidity and the third mortality worldwide.Therefore,there is an urgent need to further understand the molecular mechanism for GC development and progression and to seek for biomarkers for GC early diagnosis and prognosis prediction.CircRNAs play important roles in cancer development and progression and have the potential to serve as cancer biomarkers.Circular RNA CCDC66(circCCDC66)was previously found to be highly expressed in human colorectal cancer and could promote CRC growth and metastasis.However,its role in gastric cancer has not been reported yet.The aim of this study was to investigate the role of circCCDC66 in gastric cancer and to reveal the underlying mechanisms,providing new insights to the diagnosis and treatment of gastric cancer.Materials and Methods:(1)The expression of circCCDC66 in GC tissues and cell lines were examined by qRT-PCR.Then we collected the clinicopathological parameters.The correlation between expression level and clinicopathological characteristics was analyzed to explore the potential value of circCCDC66 in diagnosing gastric cancer.(2)The biological roles of circCCDC66 in GC cells proliferation and apoptosis were determined by cell counting,cell colony formation,flow cytometry,qRT-PCR and western blot.The role of circCCDC66 in GC cells growth was further confirmed by xenograft tumor mouse model.(3)Wound healing,transwell migration and matrigel invasion assays were performed to verify the influence of circCCDC66 on GC cells metastasis in vitro.Western blot and qRT-PCR were used to explore the effects of circCCDC66 on epithelial-mesenchymal transition(EMT)-related gene and protein expression,then explored the mechanism of circCCDC66 in gastric cancer metastasis.Results:(1)CircCCDC66 expression were elevated in both GC tissue and cell lines compared to adjacent normal tissues and control gastric epithelial cell line.The correlation between the expression level and clinicopathological parameters reveled that the upregulation of circCCDC66 in GC tissue was related to tumor stage and lymphatic metastasis.(2)CircCCDC66 knockdown significantly inhibited cells proliferation and induced cell apoptosis in GC cells.On the contrary,circCCDC66 overexpression has the opposite effects.In addition,circCCDC66 knockdown in GC cells suppressed tumor growth in nude mice.(3)Transwell migration and invasion assays showed circCCDC66 knockdown inhibited while overexpression promoted GC cells migration and invasion.Furthermore,as the qRT-PCR and western blot results showed that circCCDC66 might promote EMT through the activation of TGF-?/SMAD pathway in GC cells.Conclusion:CircCCDC66 was upregulated in GC tissue and cell lines,and promoted GC growth and metastasis by activating TGF-?/SMAD signaling pathway to promote EMT and could serve as a potential biomarker for GC.