US 12,221,608 B2
Compositions, systems, and methods for regulation of hepatitis b virus through targeted gene repression
Brian Cosgrove, Durham, NC (US); Kendra Congdon, Durham, NC (US); Jason Dean, Seattle, WA (US); Veronica Gough, Durham, NC (US); Joshua B. Black, Durham, NC (US); and Britta Jones, Durham, NC (US)
Assigned to Tune Therapeutics, Inc., Seattle, WA (US)
Filed by Tune Therapeutics, Inc., Seattle, WA (US)
Filed on Aug. 18, 2023, as Appl. No. 18/452,508.
Claims priority of provisional application 63/531,309, filed on Aug. 7, 2023.
Claims priority of provisional application 63/472,236, filed on Jun. 9, 2023.
Claims priority of provisional application 63/399,634, filed on Aug. 19, 2022.
Prior Publication US 2024/0067968 A1, Feb. 29, 2024
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/113 (2010.01); C12N 9/22 (2006.01); C12N 15/11 (2006.01); C12N 15/88 (2006.01)
CPC C12N 15/1131 (2013.01) [C12N 9/22 (2013.01); C12N 15/11 (2013.01); C12N 15/88 (2013.01); C12N 2310/11 (2013.01); C12N 2310/20 (2017.05); C12N 2310/315 (2013.01); C12N 2310/321 (2013.01); C12N 2310/351 (2013.01)] 26 Claims
 
1. An epigenetic-modifying DNA-targeting system comprising a DNA-targeting module for repressing transcription of one or more Hepatitis B viral (HBV) genes, wherein the DNA-targeting module comprises: (a) a gRNA for targeting to a target site in a Hepatitis B viral DNA sequence, wherein the target site is in a regulatory element positioned between residue 1033 and a transcription start site of the HBx gene at residue base pair 1376 of a Hepatitis B viral sequence with reference to nucleotide positions of SEQ ID NO: 650; and (b) a fusion protein comprising a deactivated Streptococcus pyogenes Cas9 (dSpCas9) protein and at least one transcriptional repressor effector domain comprising a KRAB domain and a DNA methyltransferase 3 (DNMT3) domain that has DNA methyltransferase activity, wherein the one or more HBV genes are present in both a covalently closed circular DNA (cccDNA) form and in HBV viral DNA integrated in human genomic DNA, wherein the epigenetic-modifying DNA-targeting system is capable of repressing transcription of total Hepatitis B viral RNA transcript levels.