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Chromatin Accessibility-Guided Targeting Identifies Structurally Constrained Regions in HBV cccDNA and Suppresses

Lianghao Kong1, Sadahiro Iwabuchi2,3, Ying-Yi Li1,4

  • 1Department of Clinical Laboratory Medicine, Kanazawa University Graduate School of Medical Sciences, Kanazawa-shi 920-0942, Ishikawa, Japan.

International Journal of Molecular Sciences
|August 13, 2026
PubMed

Insights

Investigating hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) chromatin accessibility revealed specific vulnerable regions. Targeting these sites with CRISPR/Cas9 reduced HBV DNA, offering a new strategy for HBV cure.

Area of Science:

  • Virology
  • Epigenetics
  • Molecular Biology

Background:

  • Covalently closed circular DNA (cccDNA) is the stable hepatitis B virus (HBV) genome form, crucial for viral replication and a key obstacle to HBV eradication.
  • Understanding cccDNA's interaction with host cell machinery, particularly chromatin, is vital for developing effective therapies.

Purpose of the Study:

  • To map chromatin accessibility patterns of HBV cccDNA in infected hepatocytes at single-molecule resolution.
  • To identify specific cccDNA regions vulnerable to therapeutic targeting based on chromatin accessibility.
  • To evaluate the efficacy of targeting these regions using a CRISPR/Cas9-based approach.

Main Methods:

  • Single-molecule resolution analysis of chromatin accessibility on HBV cccDNA in HBV-infected hepatocytes.
  • CRISPR/Cas9 gene editing to target identified accessibility-associated regions on cccDNA.
  • Quantification of HBV DNA-related signals, including cccDNA fractions and total HBV DNA, to assess therapeutic impact.

Main Results:

  • Most HBV cccDNA copies showed limited chromatin accessibility around nucleotides 800-1000, overlapping key viral gene regions.
  • A subset of cccDNA displayed detectable accessibility at this site, indicating heterogeneous chromatin states.
  • Targeting the accessibility-associated region with CRISPR/Cas9 led to significant reductions in HBV DNA levels and cccDNA enrichment.

Conclusions:

  • Chromatin accessibility profiling can identify novel, vulnerable regions within HBV cccDNA for therapeutic intervention.
  • This study provides a proof-of-concept for an accessibility-informed strategy to target and reduce HBV.
  • Further research into chromatin-associated vulnerabilities of HBV cccDNA is warranted for advancing HBV cure strategies.

Keywords:
CRISPR/Cas9cccDNAchromatin accessibilityhepatitis B virus

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