Sirtuin 2 inhibitor AGK2 exerts antiviral effects by inducing epigenetic suppression of hepatitis B virus covalently closed circular DNA through recruitment of repressive histone lysine methyltransferases and reduction of cccDNA
Chronic hepatitis B virus (HBV) infection remains a significant global health challenge because existing treatments, including interferon-α and nucleos(t)ide analogs, are unable to completely eliminate the virus. This limitation is primarily due to the persistence of covalently closed circular DNA (cccDNA) and integrated HBV DNA within infected cells. Previous studies have indicated that AGK2, a selective inhibitor of SIRT2, can suppress HBV replication by altering critical cellular signaling pathways. This research focused on further investigating the anti-HBV properties of AGK2, with particular attention to its effects on the epigenetic regulation of cccDNA.
The study employed HBV-transfected and HBV-infected cell models to evaluate how AGK2 influences viral replication. Several molecular parameters were measured to understand the mechanisms behind AGK2’s antiviral effects. These included the expression levels of SIRT2 and acetylated α-tubulin, which were assessed by immunoblotting techniques. The formation of HBV core particles was analyzed to determine the impact on viral assembly. Additionally, synthesis of HBV RNA and DNA was monitored to assess viral replication at the transcriptional and genomic levels. Levels of cccDNA were also quantified to evaluate the direct effect on the viral minichromosome.
Epigenetic changes induced by AGK2 were examined using chromatin immunoprecipitation assays. These experiments focused on the deposition of transcriptionally repressive histone modifications on cccDNA. AGK2 treatment resulted in decreased expression of SIRT2 and an increase in acetylated α-tubulin, which is consistent with SIRT2 inhibition. Furthermore, the synthesis of HBV RNA and DNA was significantly reduced following AGK2 exposure. Notably, AGK2 caused a reduction in cccDNA levels, which suggests it impacts the stability or formation of this viral reservoir.
One of the key findings was that AGK2 enhanced the enrichment of repressive histone marks such as H4K20me1, H3K27me3, and H3K9me3 on cccDNA. These histone modifications are associated with gene silencing and are deposited by specific histone lysine methyltransferases including PR-Set7, EZH2, SETDB1, and SUV39H1. The increased presence of these markers indicates a shift toward a more transcriptionally inactive chromatin state on the viral genome. In parallel, there was a decrease in the recruitment of RNA polymerase II and acetylated histone H3 to cccDNA, further supporting the conclusion that AGK2 promotes transcriptional repression of HBV.
Overall, AGK2 suppresses HBV replication not only through direct antiviral effects but also by inducing epigenetic modifications that silence cccDNA transcription. These findings suggest that targeting SIRT2 and related epigenetic regulators with AGK2 may represent a promising strategy for achieving a functional cure for chronic hepatitis B infection.