Screening For Sera Biomarker In HBV-positive Hepatocellular Carcinoma And Prediction Of HBV Encoded MiRNA

Hepatocellular carcinoma (HCC) is one of the most predominant cancers andrepresents a leading cause of malignancy-related death in China. Due to the lack ofeffective prevention, detection or therapeutic strategy, the majority of HCC patientswere diagnosed when they reached medium or even terminal stage, therefore,screening potential biomarkers to effectively detect early tumor development is animportant step towards HCC diagnosis. MicroRNAs (miRNA) are a class ofendogenous, noncoding RNAs with~22nucleotides in length which have highstability in human serum or plasma due to protection from RNases. Studies haveshown that miRNAs play a role of oncogenes or tumor suppressor genes whichregulate gene expression by binding single or multiple target mRNA. Therefore,differential expression profile of miRNAs in different samples, suggesting theirpotential merit as diagnostic biomarkers.In this study, a total of77participants including52hepatocellular carcinomapatients without any chemotherapy treatment and25normal subjects were collectedfrom Hangzhou Xixi Hospital. For miRNA sequencing,5hepatitis B virus (HBV)positive hepatocellular carcinoma patients serum were pooled together and5healthycontrol samples were pooled together, respectively. There are172differentiallyexpressed miRNAs were found though analysis of high-throughput sequencing data.All samples were further studied for expression levels of selected miRNAs byreal-time quantitative polymerase chain reaction (RT-q PCR). We screened a group ofcandidate miRNAs (miR-182-5p,363-3p and378a-3p) in serum and found they weresignificantly up-regulated in HCC patients compared to healthy controls. Furthermore,these candidate miRNAs can also differentiate HCC patients with positive HBV DNAcopies from healthy. In conclusion, our results suggest that serum miR-182-5p,363-3p, and378a-3p might be used as potential noninvasive biomarkers for HCCdiagnosis, which is especially important for HBV-positive patients.To explore whether the hepatitis B virus encoded miRNA that involve inregulation of virus infection by targeting mRNA. Vir-Mir database was used to predict HBV genome encoded pre-miRNA, and the mature miRNA sequence was validatedby miRNA sequencing and RT-q PCR. The results show that six pre-miRNAs weredetected in hepatocellular carcinoma patients with positive HBV DNA level, and amature miRNA named hbv-miR-15467was validated, which provide new ideas forthe treatment of HBV-related liver disease.