Gene Copy Number Variations In The Leukocyte Genome Of Hepatocellular Carcinoma Patients With Integrated Hepatitis B Virus DNA

Background & Aims:Hepatitis B virus(HBV)chronically infects over 250 million people worldwide and chronically infected individuals are at an increased risk for hepatocellular carcinoma(HCC),which represents the third leading cause of cancer-related mortality worldwide.Moreover,studies have shown that the integration of HBV DNA into the human liver cell genome is believed to promote HBV-related carcinogenesis.And the integration of HBV DNA into the human genome is a paramount mechanism for altering the function of endogenous genes or inducing chromosomal instability during HBV-HCC.This study aimed to quantify the integration of HBV DNA into the leukocyte genome in HBV-HCC patients in order to identify potential biomarkers for HBV-related diseases.Methods:Whole-genome comparative genomic hybridization(CGH)chip array analyses were performed to screen gene copy number variations(CNV)in the leukocyte genome,and the results were confirmed by quantitative polymerase chain reaction(qPCR).Amplification products were analyzed on 1% agarose gels.CD3+ T cells,CD3-CD19+ B cells,CD3-CD56+ natural killer cells and CD3-CD14+ monocytes were isolated from PBMCs using a MoFlo XDP high-speed flow cytometry sorter.Finally,we used receiver operating characteristic curves to calculate the diagnostic accuracy of candidate biomarkers.Results:The commonly detected regions included chromosome arms 19 p,5q,1q and 15 p,where 200 copy number gain events and 270 copy number loss events were noted.In particular,gains were observed in 5q35.3(OR4F3)and 19p13.3(OR4F17)in 90% of the samples.Successful homologous recombination of OR4F3 and the HBV P gene was demonstrated,and the amplification at 5q35.3 is potentially associated with the integration of HBV P gene into natural killer cells isolated from peripheral blood mononuclear cells(PBMCs).Receiver operating characteristic(ROC)curve analysis indicated that the combination of OR4F3 and OR4F17 a novel potential biomarker of HBV-related diseases.Conclusions:These findings revealed that HBV P gene integration in the 5' untranslated region of OR4F3 and gene copy number variations in the leukocyte genome occurred in HBV-HCC patients.Furthermore,our study also suggested that OR4F3 and OR4F17 could serve as candidate biomarkers for HBV-related diseases.