Novel assay to detect integrated HBV DNA in urine of chronic hepatitis B patients hepatitis B virus (HBV) infection is a global public health concern

The CDC estimates that up to 2 million americans are chronically infected with HBV. HBV infection causes acute and chronic hepatitis, the latter often leading to severe liver diseases such cirrhosis and HBV-associated hepatocellular carcinoma (HBV-HCC). HBV-HCC accounts for 50% of HCC cases worldwide. HBV is a DNA virus, and its partial genome can integrate into a host chromosome. Although HBV DNA replication can be suppressed effectively by nucleoside analogs, integrated viral DNA and covalently closed circular HBV DNA (CCCDNA) persist in hepatocytes even after prolonged antiviral therapy. Continuous antiviral treatment is necessary to keep viral replication suppressed due to its ineffectiveness in eliminating the "remnant" HBV DNA (i.e., integrated HBV DNA and CCCDNA) fully. Currently, the treatment goal is to achieve a functional cure, defined as the clearance of hepatitis B surface antigen (HBSAG), without definitively eliminating integrated or CCCDNA. No clinical test, however, is currently capable of identifying this "cured" population. The current thinking is that after prolonged effective anti-HBV therapy, integrated HBV DNA is the major species of residual HBV DNA in liver and most HBSAG in HBEAG-negative patients is derived from this DNA. The ability to detect integrated DNA in patients with chronic hepatitis B is an urgent unmet need. The goal of this application is to develop a noninvasive urine test that can detect and characterize integrated HBV DNA, and be used to monitor antiviral treatment efficacy and identify "cured" subjects who can be taken off the lifelong therapy. In the phase I feasibility study, we will improve our current prototype HBV-host junction detection assay and develop a computation pipeline, hbvjseq, for comprehensive profiling of HBV integration and HBV genetics (aim 1). In aim 2, we will perform comparative analyses to correlate quantitative and qualitative characteristics of integrated DNA in urine and plasma with the levels of HBSAG and HBV pregenomic RNA (PGRNA) in hepatitis B E-antigen (HBEAG)-negative patients. Results of these analyses will be used to determine if cell-free integrated HBV DNA can be developed as a biomarker to identify "cured" patients in HBEAG-negative populations, warranting a phase II study to bring the test to the clinic for disease management.