The Expression And Clinical Significance Of Circ-Foxo3 In Renal Cell Carcinoma

Background:Renal cell carcinoma is a common urinary system tumor.In recent years,the incidence has increased at a rate of 2% per year.Although the screening and detection methods for renal cell carcinoma have continued to improve over the past decade,the early detection rate of small renal cell carcinoma has been significantly higher than that of symptomatic renal cell carcinoma,but about 20% to 30% of patients with renal cell carcinoma have already developed locally advanced manifestations such as lymph nodes metastasis or thrombus formation in veins at the early stage of diagnosis.CircRNA has been a hot area in recent tumor research.More and more studies have shown that circRNA plays a crucial role in tumorigenesis and development by acting as a miRNA sponge.Circ-Foxo3 has been reported to be significantly decreased in many tumor tissues and can regulate the cellular functions of tissue cells while suppress cancer cell progression.However,the molecular functions of circ-Foxo3 in renal cell carcinoma have rarely been explored.Objective:To detect the expression level of circ-Foxo3 in renal cell carcinoma and corresponding normal tissue adjacent to cancer and further analyze its potential clinical significance.Methods:Quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the expression level of circ-Foxo3 in renal cell carcinoma cell lines and normal renal tubular cell lines.A total of 26 pairs of tumors and adjacent normal renal tissue(>3cm from the edge of the tumor)of RCC patients who underwent radical nephrectomy in Department of Urology,Nanjing General Hospital of Nanjing Military Command from May 2016 to October 2016 were collected.After diagnosis by pathology,there are 20 cases of clear cell renal cell carcinoma(ccRCC),3 cases of papillary renal cell carcinoma,2 cases of chromophobic renal cell carcinoma,1 case of Xp11.2translocation/TFE3 fusion gene renal cell carcinoma.One of the samples wasrandomly selected for microarray analysis.Then,qRT-PCR was used to detect the expression level of circ-Foxo3 in RCC tissues and paired paracancerous tissues,and the difference of expression of circ-Foxo3 in RCC and corresponding normal tissues was analyzed and its clinical significance was analyzed at the same time.Finally,the correlation between circ-Foxo3 expression and clinical pathological features in RCC samples and the correlation between circ-Foxo3 expression and pathological grade(Furhman grade)in ccRCC samples were analyzed.Results:1.qRT-PCR detection of circ-Foxo3 transcription level.Among them HK2 is normal tubular epithelial cells,and A498 and Caki-1 are two human renal cancer cell lines.circ-Foxo3 transcription was significantly down-regulated in A498 and Caki-1cells compared to HK2 cells(all P<0.001).2.Microarray validation showed that circ-Foxo3 was significantly down-regulated in renal cancer tissues compared with normal tissues.In addition,binding sites exist for hsa-miR-138-5p,hsa-miR-143-3p,hsa-miR-185-3p,hsa-miR-30c-1-3p,and hsa-miR-762.3.qRT-PCR was used to detect the expression of circ-Foxo3 in all renal cell carcinoma samples and their corresponding paracancerous tissues.The results showed that the relative expression in renal cell carcinoma tissues was lower than that in adjacent normal tissues,and the difference was statistically significant.Significance(P<0.001).The relative expression level of 20 ccRCC samples was also lower than that of the corresponding normal tissue adjacent to the cancer.The difference was statistically significant(P<0.001).4.The receiver operating characteristic curve(ROC)analysis showed that the expression level of circ-Foxo3 could effectively identify the RCC group and its control group [AUC=0.858(95%CI:0.759,0.957)] with high specificity and sensitivity..Circ-Foxo3 expression levels can also be identified in the ccRCC group and its control group [AUC 0.805(95% CI: 0.671,0.939)],with similar specificity and sensitivity.5.To further divide the renal cell carcinoma into circ-Foxo3 high expressiongroup(n = 4)and low expression group(n = 22),we use the cutoff value of 0.0018 of circ-Foxo3,The square test showed that there was no significant correlation between circ-Foxo3 expression in RCC tissues and clinicopathological characteristics such as AJCC staging,patients' age,sex and tumor morphology.(all P>0.05).The cutoff value of circ-Foxo3 expression in 20 cases of ccRCC was 0.0018.The ccRCC patients were divided into circ-Foxo3 high expression group(n=4)and low expression group(n=16).There was no significant correlation between circ-Foxo3 expression levels and Furhman grading(P>0.05).Conclusions:1.circ-Foxo3 was highly expressed in RCC tissues and cell lines,which may play an important role in the occurrence and progression of RCC;2.circ-Foxo3 can be used as a potential biomarker for diagnosis of RCC.