Targeting the hepatitis B cccDNA with a sequence-specific ARCUS nuclease to eliminate hepatitis B virus in vivo

Persistence of chronic hepatitis B (CHB) is attributed to main-tenance of the intrahepatic pool of the viral covalently closed circular DNA (cccDNA), which serves as the transcriptional template for all viral gene products required for replication. Current nucleos(t)ide therapies for CHB prevent virus produc-tion and spread but have no direct impact on cccDNA or expres-sion of viral genes. We describe a potential curative approach using a highly specific engineered ARCUS nuclease (ARCUS-POL) targeting the hepatitis B virus (HBV) genome. Transient ARCUS-POL expression in HBV-infected primary human he-patocytes produced substantial reductions in both cccDNA and hepatitis B surface antigen (HBsAg). To evaluate ARCUS-POL in vivo, we developed episomal adeno-associated virus (AAV) mouse and non-human primate (NHP) models contain-ing a portion of the HBV genome serving as a surrogate for cccDNA. Clinically relevant delivery was achieved through systemic administration of lipid nanoparticles containing AR-CUS-POL mRNA. In both mouse and NHP, we observed a sig-nificant decrease in total AAV copy number and high on-target indel frequency. In the case of the mouse model, which supports HBsAg expression, circulating surface antigen was durably reduced by 96%. Together, these data support a gene-editing approach for elimination of cccDNA toward an HBV cure.

Automated summary

This study discovered a highly specific engineered ARCUS nuclease (ARCUS-POL) targeting the hepatitis B virus (HBV) genome, which significantly reduces both the viral covalently closed circular DNA (cccDNA) and hepatitis B surface antigen (HBsAg) in liver cells. Using mouse and non-human primate models, the researchers demonstrated that ARCUS-POL can modify target genes, resulting in a durable reduction of HBsAg levels. These findings provide a theoretical basis for the application of gene editing in HBV treatment.