Mechanisms Underlying Delayed loss of HBeAg and HBV DNA Following HBsAg Seroclearance in PEG-IFNα Treated Patients of chronic hepatitis B

A notable proportion of CHB patients undergoing PEG-IFNα based therapy experience lagged serum HBeAg and/or HBV DNA disappearance in patients achieving HBsAg loss. In this study, we explored the molecular mechanisms behind this clinical phenomenon, offering novel insights into the sustainability of chronic HBV infection. Two independent clinical cohorts were enrolled to validate this phenomenon. Then comprehensive analysis was performed using public datasets, coupled with a series of molecular biology experiments. Approximately 17-20% CHB patients underwent PEG-IFNα based therapy experienced seroclearance of HBsAg, while serum HBeAg and/or HBV DNA remained positive. These patients are more prone to serum HBsAg reappearance compared to those achieving complete virological response. Analysis of public datasets revealed that compared to the PC/BCP, the SP1/SP2 promoter displayed more pronounced inhibitory epigenetic modifications in HBeAg-negative patients and SP1/SP2 in-frame mutation peaked in immune active patients. In vitro experiments demonstrated that introduced SP1/SP2 inactive mutations would enhance PC/BCP transcriptional activity by a mechanism known as adjacent transcriptional interference. Furthermore, the deletion of L-HBsAg facilitated intracellular cccDNA replenishment. This study elucidates that under IFNα treatment and low viral load, transcriptional suppression of SP1/SP2 promoters through mutations and/or epigenetic changes would favor the maintenance of sustain chronic HBV infection, via enhancing the transcription activity of BCP to promote cccDNA replenishment.