Compositions and methods for epigenetic regulation of HBV gene expression
Inventors
Jaffe, Aron Brandon • Abubucker, Noorussahar • Anglero-Rodriguez, Yesseinia • Myer, Vic • Lombardo, Angelo Leone • Cappelluti, Martino Alfredo
Assignees
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Publication Number
US-12558437-B2
Publication Date
2026-02-24
Expiration Date
Abstract
This invention relates to compositions, methods, strategies, and treatment modalities related to the epigenetic modification of hepatitis B virus (HBV) genes.
Core Innovation
An epigenetic editing system is administered to a subject having detectable levels of HBV DNA and/or HBsAg and/or HBeAg in serum. The epigenetic editing system comprises a fusion protein, or a nucleic acid encoding the fusion protein, including a first dCas9 protein domain, a first DNMT domain, and an epigenetic repression domain that is a KRAB domain.
The dCas9 protein domain binds an HBV genome target region located within a region comprising nucleotides 1000-2448. In one embodiment, the target region overlaps with CpG Island II (CGI II) of the HBV genome, and in another embodiment the target region comprises nucleotides within 500 bps of a transcription start site of an HBx RNA.
The epigenetic editing system includes a first guide RNA (gRNA) comprising a region complementary to a strand of the target region, or one or more other nucleic acid molecules encoding the same. Administering results in reduction of the level of HBV DNA, HBsAg, and/or HBeAg in the serum of the subject, and the reduction is maintained for at least 35 days after administering.
Claims Coverage
The consolidated claim coverage includes two independent method claims. Both claims direct administering a KRAB-linked dCas9/DNMT epigenetic editing system with an HBV-targeting gRNA to reduce serum HBV markers for at least 35 days, with claim-specific target-region constraints within HBV nucleotides 1000-2448.
KRAB-linked dCas9 and DNMT fusion for epigenetic editing
A fusion protein, or a nucleic acid encoding the fusion protein, comprising a first dCas9 protein domain, a first DNMT domain, and an epigenetic repression domain, wherein the epigenetic repression domain is a KRAB domain.
HBV target region within nucleotides 1000-2448 overlapping CpG Island II
The dCas9 protein domain binds a first target region of an HBV genome located within a region comprising nucleotides 1000-2448, wherein the first target region overlaps with CpG Island II (CGI II) of the HBV genome.
HBx transcription start site proximity within the HBV target region
The dCas9 protein domain binds a first target region of an HBV genome located within a region comprising nucleotides 1000-2448, wherein the first target region comprises nucleotides within 500 bps of a transcription start site of an HBx RNA.
gRNA complementary to the HBV target region
A first guide RNA (gRNA) comprising a region complementary to a strand of the first target region, or one or more other nucleic acid molecules encoding the same.
Serum HBV marker reduction maintained after administering
Administering the epigenetic editing system results in a reduction of the level of HBV DNA, the level of HBsAg, and/or the level of HBeAg in the serum of the subject, wherein the reduction is maintained for at least 35 days after the administering.
Both independent claims require a KRAB-domain epigenetic repression function fused to dCas9 and a DNMT domain, together with an HBV-targeting gRNA that binds an HBV target region within nucleotides 1000-2448. One claim requires overlap with CpG Island II, while the other requires proximity to the HBx transcription start site; both claims require reduction of serum HBV markers that is maintained for at least 35 days after administering.
Stated Advantages
Reduction of serum HBV DNA and/or HBsAg and/or HBeAg after administering the epigenetic editing system.
The reduction of serum HBV markers is maintained for at least 35 days after administering.
Documented Applications
A method of administering an epigenetic editing system to a subject with detectable HBV DNA and/or HBsAg and/or HBeAg in serum to reduce those serum markers.
In vivo and in vitro evaluation against HBV in AAV-HBV and Tg-HBV mouse models, including reductions measured by HBsAg, HBV DNA, and HBeAg readouts.
Evaluation in HepG2.2.15 cells for cellular methylation/off-target specificity using LNPs.
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