{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T04:15:24Z","timestamp":1790309724610,"version":"4.1.0"},"reference-count":110,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2018,4,12]],"date-time":"2018-04-12T00:00:00Z","timestamp":1523491200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Chronic infection with the hepatitis B virus (HBV) is a global health concern and accounts for approximately 1 million deaths annually. Amongst other limitations of current anti-HBV treatment, failure to eliminate the viral covalently closed circular DNA (cccDNA) and emergence of resistance remain the most worrisome. Viral rebound from latent episomal cccDNA reservoirs occurs following cessation of therapy, patient non-compliance, or the development of escape mutants. Simultaneous viral co-infections, such as by HIV-1, further complicate therapeutic interventions. These challenges have prompted development of novel targeted hepatitis B therapies. Given the ease with which highly specific and potent nucleic acid therapeutics can be rationally designed, gene therapy has generated interest for antiviral application. Gene therapy strategies developed for HBV include gene silencing by harnessing RNA interference, transcriptional inhibition through epigenetic modification of target DNA, genome editing by designer nucleases, and immune modulation with cytokines. DNA-binding domains and effectors based on the zinc finger (ZF), transcription activator-like effector (TALE), and clustered regularly interspaced short palindromic repeat (CRISPR) systems are remarkably well suited to targeting episomal cccDNA. This review discusses recent developments and challenges facing the field of anti-HBV gene therapy, its potential curative significance and the progress towards clinical application.<\/jats:p>","DOI":"10.3390\/genes9040207","type":"journal-article","created":{"date-parts":[[2018,4,12]],"date-time":"2018-04-12T12:19:27Z","timestamp":1523535567000},"page":"207","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":65,"title":["Gene Therapy for Chronic HBV\u2014Can We Eliminate cccDNA?"],"prefix":"10.3390","volume":"9","author":[{"given":"Kristie","family":"Bloom","sequence":"first","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, Private Bag 3, Johannesburg, WITS 2050, South Africa"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohube Betty","family":"Maepa","sequence":"additional","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, Private Bag 3, Johannesburg, WITS 2050, South Africa"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Abdullah","family":"Ely","sequence":"additional","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, Private Bag 3, Johannesburg, WITS 2050, South Africa"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4149-8995","authenticated-orcid":false,"given":"Patrick","family":"Arbuthnot","sequence":"additional","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, Private Bag 3, Johannesburg, WITS 2050, South Africa"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2018,4,12]]},"reference":[{"key":"ref_1","unstructured":"WHO (2018, January 06). Global Hepatitis Report. Available online: http:\/\/www.who.int\/hepatitis\/ publications\/global-hepatitis-report2017\/en\/."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"541","DOI":"10.1001\/jama.1965.03080070025007","article-title":"A \u201cnew\u201d antigen in leukemia sera","volume":"191","author":"Blumberg","year":"1965","journal-title":"JAMA"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1002\/hep.28156","article-title":"Aasld guidelines for treatment of chronic hepatitis B","volume":"63","author":"Terrault","year":"2016","journal-title":"Hepatology"},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"EASL (2017). 2017 clinical practice guidelines on the management of hepatitis B virus infection. J. Hepatol., 67, 370\u2013398.","DOI":"10.1016\/j.jhep.2017.03.021"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/s12072-015-9675-4","article-title":"Asian-pacific clinical practice guidelines on the management of hepatitis B: A 2015 update","volume":"10","author":"Sarin","year":"2016","journal-title":"Hepatol. Int."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"S771","DOI":"10.1093\/infdis\/jix355","article-title":"Toward a cure for hepatitis B virus infection: Combination therapy involving viral suppression and immune modulation and long-term outcome","volume":"216","author":"Wu","year":"2017","journal-title":"J. Infect. Dis."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"2427","DOI":"10.4254\/wjh.v7.i23.2427","article-title":"Antiviral therapy for chronic hepatitis B: Combination of nucleoside analogs and interferon","volume":"7","author":"Hagiwara","year":"2015","journal-title":"World J. Hepatol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1016\/0092-8674(86)90602-1","article-title":"Formation of the pool of covalently closed circular viral DNA in hepadnavirus-infected cells","volume":"47","author":"Tuttleman","year":"1986","journal-title":"Cell"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3350","DOI":"10.1128\/jvi.69.6.3350-3357.1995","article-title":"The covalently closed duplex form of the hepadnavirus genome exists in situ as a heterogeneous population of viral minichromosomes","volume":"69","author":"Newbold","year":"1995","journal-title":"J. Virol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1007\/BF01703079","article-title":"Hepatitis B virus genome is organized into nucleosomes in the nucleus of the infected cell","volume":"8","author":"Bock","year":"1994","journal-title":"Virus Genes"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"823","DOI":"10.1053\/j.gastro.2006.01.001","article-title":"Hepatitis B virus replication is regulated by the acetylation status of hepatitis B virus cccDNA-bound H3 and H4 histones","volume":"130","author":"Pollicino","year":"2006","journal-title":"Gastroenterology"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"19975","DOI":"10.1073\/pnas.0908365106","article-title":"Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function","volume":"106","author":"Belloni","year":"2009","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"E5715","DOI":"10.1073\/pnas.1518090112","article-title":"Mapping of histone modifications in episomal HBV cccDNA uncovers an unusual chromatin organization amenable to epigenetic manipulation","volume":"112","author":"Tropberger","year":"2015","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"386","DOI":"10.1038\/nature17170","article-title":"Hepatitis B virus X protein identifies the Smc5\/6 complex as a host restriction factor","volume":"531","author":"Decorsiere","year":"2016","journal-title":"Nature"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"S117","DOI":"10.1016\/j.jhep.2016.02.016","article-title":"New antiviral targets for innovative treatment concepts for hepatitis B virus and hepatitis delta virus","volume":"64","author":"Durantel","year":"2016","journal-title":"J. Hepatol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"e10","DOI":"10.1016\/S1473-3099(15)00436-3","article-title":"New therapeutic agents for chronic hepatitis B","volume":"16","author":"Brahmania","year":"2016","journal-title":"Lancet. Infect. Dis."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"131","DOI":"10.1016\/j.coph.2016.08.004","article-title":"Modulators of HBV capsid assembly as an approach to treating hepatitis B virus infection","volume":"30","author":"Cole","year":"2016","journal-title":"Curr. Opin. Pharmacol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"82","DOI":"10.1016\/j.antiviral.2015.06.020","article-title":"Core protein: A pleiotropic keystone in the HBV lifecycle","volume":"121","author":"Zlotnick","year":"2015","journal-title":"Antiviral Res."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"539","DOI":"10.2217\/17460913.3.5.539","article-title":"Thiazolides: A new class of drugs for the treatment of chronic hepatitis B and C","volume":"3","author":"Rossignol","year":"2008","journal-title":"Future Microbiol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"5660","DOI":"10.1021\/jm200696v","article-title":"Design, synthesis, and biological evaluation of triazolo-pyrimidine derivatives as novel inhibitors of hepatitis B virus surface antigen (HBsAg) secretion","volume":"54","author":"Yu","year":"2011","journal-title":"J. Med. Chem."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1016\/j.antiviral.2016.07.004","article-title":"Nucleic acid polymers: Broad spectrum antiviral activity, antiviral mechanisms and optimization for the treatment of hepatitis B and hepatitis D infection","volume":"133","author":"Vaillant","year":"2016","journal-title":"Antiviral Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"e05570","DOI":"10.7554\/eLife.05570","article-title":"Sodium taurocholate cotransporting polypeptide is a functional receptor for human hepatitis B and D virus","volume":"3","author":"Yan","year":"2012","journal-title":"Elife"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"861","DOI":"10.1016\/j.jhep.2012.12.008","article-title":"The entry inhibitor myrcludex-B efficiently blocks intrahepatic virus spreading in humanized mice previously infected with hepatitis B virus","volume":"58","author":"Volz","year":"2013","journal-title":"J. Hepatol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"17589","DOI":"10.3390\/ijms160817589","article-title":"Progress and prospects of Anti-HBV gene therapy development","volume":"16","author":"Maepa","year":"2015","journal-title":"Int. J. Mol. Sci"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"383","DOI":"10.1002\/rmv.705","article-title":"Countering hepatitis B virus infection using RNAi: How far are we from the clinic?","volume":"21","author":"Ivacik","year":"2011","journal-title":"Rev. Med. Virol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"411","DOI":"10.1016\/j.ymthe.2005.10.013","article-title":"Effective inhibition of HBV replication in vivo by anti-HBx short hairpin RNAs","volume":"13","author":"Carmona","year":"2006","journal-title":"Mol. Ther."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"e91","DOI":"10.1093\/nar\/gkp446","article-title":"Efficient silencing of gene expression with modular trimeric Pol II expression cassettes comprising microRNA shuttles","volume":"37","author":"Ely","year":"2009","journal-title":"Nucleic Acids Res."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"6145","DOI":"10.1016\/j.bmc.2013.04.073","article-title":"Inhibition of hepatitis B virus replication in cultured cells and in vivo using 2\u2019-O-guanidinopropyl modified siRNAs","volume":"21","author":"Marimani","year":"2013","journal-title":"Bioorg. Med. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"537","DOI":"10.1038\/nature04791","article-title":"Fatality in mice due to oversaturation of cellular microRNA\/short hairpin RNA pathways","volume":"441","author":"Grimm","year":"2006","journal-title":"Nature"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"190","DOI":"10.1016\/j.omtn.2017.04.007","article-title":"Sustained inhibition of HBV replication in vivo after systemic injection of AAVs encoding artificial antiviral primary microRNAs","volume":"7","author":"Maepa","year":"2017","journal-title":"Mol. Ther. Nucleic Acids"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"E4244","DOI":"10.1073\/pnas.1409986111","article-title":"Involvement of the host DNA-repair enzyme TDP2 in formation of the covalently closed circular DNA persistence reservoir of hepatitis B viruses","volume":"111","author":"Koniger","year":"2014","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"638","DOI":"10.1016\/j.bbrc.2016.11.085","article-title":"Host factor PRPF31 is involved in cccDNA production in HBV-replicating cells","volume":"482","author":"Kinoshita","year":"2017","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"818","DOI":"10.1016\/j.jhep.2017.05.012","article-title":"Therapeutic editing of hepatocyte genome in vivo","volume":"67","author":"Lujambio","year":"2017","journal-title":"J. Hepatol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"311","DOI":"10.1016\/j.virusres.2017.01.003","article-title":"Advances with using CRISPR\/Cas-mediated gene editing to treat infections with hepatitis B virus and hepatitis C virus","volume":"244","author":"Moyo","year":"2018","journal-title":"Virus Res."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"3","DOI":"10.3851\/IMP1619","article-title":"Molecular genetics of HBV infection","volume":"15","author":"Locarnini","year":"2010","journal-title":"Antivir. Ther."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1038\/nbt0202-135","article-title":"Engineering polydactyl zinc-finger transcription factors","volume":"20","author":"Beerli","year":"2002","journal-title":"Nat. Biotechnol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"10570","DOI":"10.1073\/pnas.95.18.10570","article-title":"Foki dimerization is required for DNA cleavage","volume":"95","author":"Bitinaite","year":"1998","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"947","DOI":"10.1038\/mt.2010.20","article-title":"Zinc-finger nucleases as a novel therapeutic strategy for targeting hepatitis B virus DNAs","volume":"18","author":"Cradick","year":"2010","journal-title":"Mol. Ther."},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Weber, N.D., Stone, D., Sedlak, R.H., De Silva Feelixge, H.S., Roychoudhury, P., Schiffer, J.T., Aubert, M., and Jerome, K.R. (2014). AAV-mediated delivery of zinc finger nucleases targeting hepatitis B virus inhibits active replication. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0097579"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1715","DOI":"10.1128\/AAC.41.8.1715","article-title":"Inducible expression of human hepatitis B virus (HBV) in stably transfected hepatoblastoma cells: A novel system for screening potential inhibitors of HBV replication","volume":"41","author":"Ladner","year":"1997","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1509","DOI":"10.1126\/science.1178811","article-title":"Breaking the code of DNA binding specificity of TAL-type III effectors","volume":"326","author":"Boch","year":"2009","journal-title":"Science"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1501","DOI":"10.1126\/science.1178817","article-title":"A simple cipher governs DNA recognition by TAL effectors","volume":"326","author":"Moscou","year":"2009","journal-title":"Science"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"720","DOI":"10.1126\/science.1215670","article-title":"Structural basis for sequence-specific recognition of DNA by TAL effectors","volume":"335","author":"Deng","year":"2012","journal-title":"Science"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"716","DOI":"10.1126\/science.1216211","article-title":"The crystal structure of TAL effector PthXo1 bound to its DNA target","volume":"335","author":"Mak","year":"2012","journal-title":"Science"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1889","DOI":"10.1038\/mt.2013.170","article-title":"Inactivation of hepatitis B virus replication in cultured cells and in vivo with engineered transcription activator-like effector nucleases","volume":"21","author":"Bloom","year":"2013","journal-title":"Mol. Ther."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"303","DOI":"10.1038\/mt.2013.212","article-title":"An efficient antiviral strategy for targeting hepatitis B virus genome using transcription activator-like effector nucleases","volume":"22","author":"Chen","year":"2014","journal-title":"Mol. Ther."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1563","DOI":"10.1016\/j.bbrc.2016.08.152","article-title":"Improved antiviral efficacy using TALEN-mediated homology directed recombination to introduce artificial primary mirnas into DNA of hepatitis B virus","volume":"478","author":"Dreyer","year":"2016","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"816","DOI":"10.1126\/science.1225829","article-title":"A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity","volume":"337","author":"Jinek","year":"2012","journal-title":"Science"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"e216","DOI":"10.1038\/mtna.2014.68","article-title":"Targeting hepatitis B virus with CRISPR\/Cas9","volume":"3","author":"Seeger","year":"2014","journal-title":"Mol. Ther. Nucleic Acids"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"110","DOI":"10.1016\/j.antiviral.2015.03.015","article-title":"Targeting hepatitis B virus cccDNA by CRISPR\/Cas9 nuclease efficiently inhibits viral replication","volume":"118","author":"Dong","year":"2015","journal-title":"Antiviral Res."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"13734","DOI":"10.1038\/srep13734","article-title":"Crispr\/cas9 nickase-mediated disruption of hepatitis B virus open reading frame S and X","volume":"5","author":"Karimova","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"196","DOI":"10.1016\/j.virol.2014.12.001","article-title":"Suppression of hepatitis B virus DNA accumulation in chronically infected cells using a bacterial CRISP\/Cas RNA-guided DNA endonuclease","volume":"476","author":"Kennedy","year":"2015","journal-title":"Virology"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"2252","DOI":"10.1099\/vir.0.000159","article-title":"Inhibition of hepatitis B virus by the CRISPR\/Cas9 system via targeting the conserved regions of the viral genome","volume":"96","author":"Liu","year":"2015","journal-title":"J. Gen. Virol."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"10833","DOI":"10.1038\/srep10833","article-title":"CRISPR\/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus","volume":"5","author":"Ramanan","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"9554","DOI":"10.3748\/wjg.v21.i32.9554","article-title":"Dual gRNAs guided CRISPR\/Cas9 system inhibits hepatitis B virus replication","volume":"21","author":"Wang","year":"2015","journal-title":"World J. Gastroenterol."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1038\/gt.2015.2","article-title":"Harnessing the clustered regularly interspaced short palindromic repeat (CRISPR)\/CRISPR-associated Cas9 system to disrupt the hepatitis B virus","volume":"22","author":"Zhen","year":"2015","journal-title":"Gene Ther."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"1253","DOI":"10.1111\/gtc.12437","article-title":"Highly multiplexed CRISPR-Cas9-nuclease and Cas9-nickase vectors for inactivation of hepatitis B virus","volume":"21","author":"Sakuma","year":"2016","journal-title":"Genes Cells"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"1258","DOI":"10.1038\/mt.2016.94","article-title":"Complete spectrum of CRISPR\/Cas9-induced mutations on HBV cccDNA","volume":"24","author":"Seeger","year":"2016","journal-title":"Mol. Ther."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1016\/j.virusres.2016.04.003","article-title":"CRISPR\/Cas9 produces anti-hepatitis B virus effect in hepatoma cells and transgenic mouse","volume":"217","author":"Zhu","year":"2016","journal-title":"Virus Res."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"1104","DOI":"10.7150\/ijbs.16064","article-title":"An effective molecular target site in hepatitis B virus S gene for Cas9 cleavage and mutational inactivation","volume":"12","author":"Li","year":"2016","journal-title":"Int. J. Biol. Sci."},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"440","DOI":"10.1038\/cr.2017.16","article-title":"A non-viral CRISPR\/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo","volume":"27","author":"Jiang","year":"2017","journal-title":"Cell Res."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"91","DOI":"10.3389\/fcimb.2017.00091","article-title":"Removal of integrated hepatitis B virus DNA using CRISPR-Cas9","volume":"7","author":"Li","year":"2017","journal-title":"Front. Cell. Infect. Microbiol."},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"7401","DOI":"10.1038\/s41598-017-07642-6","article-title":"ssAAVs containing cassettes encoding SaCas9 and guides targeting hepatitis B virus inactivate replication of the virus in cultured cells","volume":"7","author":"Scott","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"3090","DOI":"10.7150\/thno.18114","article-title":"The gRNA-miRNA-gRNA ternary cassette combining CRISPR\/Cas9 with RNAi approach strongly inhibits hepatitis B virus replication","volume":"7","author":"Wang","year":"2017","journal-title":"Theranostics"},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"7199","DOI":"10.3892\/mmr.2017.7518","article-title":"Efficient inhibition of duck hepatitis B virus DNA by the CRISPR\/Cas9 system","volume":"16","author":"Zheng","year":"2017","journal-title":"Mol Med Rep"},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"677","DOI":"10.1038\/nbt.2916","article-title":"Genome-wide analysis reveals characteristics of off-target sites bound by the Cas9 endonuclease","volume":"32","author":"Kuscu","year":"2014","journal-title":"Nat. Biotechnol."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"7473","DOI":"10.1093\/nar\/gku402","article-title":"CRISPR\/Cas9 systems have off-target activity with insertions or deletions between target DNA and guide RNA sequences","volume":"42","author":"Lin","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"822","DOI":"10.1038\/nbt.2623","article-title":"High frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells","volume":"31","author":"Fu","year":"2013","journal-title":"Nat. Biotechnol."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1016\/j.mrrev.2008.02.004","article-title":"Epigenetic interplay between histone modifications and DNA methylation in gene silencing","volume":"659","author":"Vaissiere","year":"2008","journal-title":"Mutat. Res."},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"398","DOI":"10.1002\/hep.29133","article-title":"PRMT5 restricts hepatitis B virus replication through epigenetic repression of covalently closed circular DNA transcription and interference with pregenomic RNA encapsidation","volume":"66","author":"Zhang","year":"2017","journal-title":"Hepatology"},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"720","DOI":"10.4161\/epi.6.6.15815","article-title":"HBc binds to the CpG islands of HBV cccDNA and promotes an epigenetic permissive state","volume":"6","author":"Guo","year":"2011","journal-title":"Epigenetics"},{"key":"ref_72","doi-asserted-by":"crossref","unstructured":"Zhang, Y., Mao, R., Yan, R., Cai, D., Zhang, Y., Zhu, H., Kang, Y., Liu, H., Wang, J., and Qin, Y. (2014). Transcription of hepatitis B virus covalently closed circular DNA is regulated by CpG methylation during chronic infection. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0110442"},{"key":"ref_73","first-page":"1491","article-title":"The epigenetic control of hepatitis B virus modulates the outcome of infection","volume":"6","author":"Koumbi","year":"2015","journal-title":"Front. Microbiol."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"2066","DOI":"10.1002\/hep.29479","article-title":"Epigenetic regulation of hepatitis B virus covalently closed circular DNA: Implications for epigenetic therapy against chronic hepatitis B","volume":"66","author":"Hong","year":"2017","journal-title":"Hepatology"},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1111\/1348-0421.12345","article-title":"Interaction of the hepatitis B virus X protein with the lysine methyltransferase SET and MYND domain-containing 3 induces activator protein 1 activation","volume":"60","author":"Hayashi","year":"2016","journal-title":"Microbiol. Immunol."},{"key":"ref_76","doi-asserted-by":"crossref","unstructured":"Lubyova, B., Hodek, J., Zabransky, A., Prouzova, H., Hubalek, M., Hirsch, I., and Weber, J. (2017). PRMT5: A novel regulator of hepatitis B virus replication and an arginine methylase of HBV core. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0186982"},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"2811","DOI":"10.3892\/or.2017.5518","article-title":"HBx represses RIZ1 expression by DNA methyltransferase 1 involvement in decreased miR-152 in hepatocellular carcinoma","volume":"37","author":"Zhao","year":"2017","journal-title":"Oncol. Rep."},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"e186","DOI":"10.1038\/mtna.2014.38","article-title":"The CRISPR\/Cas9 system facilitates clearance of the intrahepatic HBV templates in vivo","volume":"3","author":"Lin","year":"2014","journal-title":"Mol. Ther. Nucleic Acids"},{"key":"ref_79","first-page":"111","article-title":"Hepatitis B virus can be inhibited by DNA methyltransferase 3a via specific zinc-finger-induced methylation of the X promoter","volume":"79","author":"Xirong","year":"2014","journal-title":"Biochemistry"},{"key":"ref_80","doi-asserted-by":"crossref","first-page":"378","DOI":"10.1177\/1087057112463066","article-title":"Creation of a six-fingered artificial transcription factor that represses the hepatitis B virus HBx gene integrated into a human hepatocellular carcinoma cell line","volume":"18","author":"Zhao","year":"2013","journal-title":"J. Biomol. Screen."},{"key":"ref_81","doi-asserted-by":"crossref","first-page":"S192","DOI":"10.1016\/S1525-0016(16)34092-8","article-title":"483. Epigenetic silencing of hepatitis B cccDNA in vitro and in vivo using AAV-delivered engineered repressor transcription activator-like effector","volume":"23","author":"Moyo","year":"2015","journal-title":"Mol. Ther."},{"key":"ref_82","doi-asserted-by":"crossref","unstructured":"Liu, F., Campagna, M., Qi, Y., Zhao, X., Guo, F., Xu, C., Li, S., Li, W., Block, T.M., and Chang, J. (2013). \u03b1-interferon suppresses hepadnavirus transcription by altering epigenetic modification of cccDNA minichromosomes. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003613"},{"key":"ref_83","doi-asserted-by":"crossref","first-page":"333","DOI":"10.1016\/j.antiviral.2012.10.003","article-title":"Anti-hepatitis B virus activities of \u03b1-DDB-FNC, a novel nucleoside-biphenyldicarboxylate compound in cells and ducks, and its anti-immunological liver injury effect in mice","volume":"96","author":"Yang","year":"2012","journal-title":"Antiviral Res."},{"key":"ref_84","doi-asserted-by":"crossref","unstructured":"Zhang, Y., Li, C., Zhang, Y., Zhu, H., Kang, Y., Liu, H., Wang, J., Qin, Y., Mao, R., and Xie, Y. (2013). Comparative analysis of CpG islands among HBV genotypes. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0056711"},{"key":"ref_85","doi-asserted-by":"crossref","first-page":"103","DOI":"10.1111\/j.1365-2893.2007.00905.x","article-title":"Hepatitis B viral DNA is methylated in liver tissues","volume":"15","author":"Vivekanandan","year":"2008","journal-title":"J. Viral Hepat."},{"key":"ref_86","doi-asserted-by":"crossref","first-page":"10478","DOI":"10.1038\/srep10478","article-title":"Comprehensive DNA methylation analysis of hepatitis B virus genome in infected liver tissues","volume":"5","author":"Jain","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_87","doi-asserted-by":"crossref","first-page":"1227","DOI":"10.1086\/529437","article-title":"Comprehensive genetic and epigenetic analysis of occult hepatitis B from liver tissue samples","volume":"46","author":"Vivekanandan","year":"2008","journal-title":"Clin. Infect. Dis."},{"key":"ref_88","doi-asserted-by":"crossref","first-page":"700","DOI":"10.1086\/655398","article-title":"DNA methylation of hepatitis B virus (HBV) genome associated with the development of hepatocellular carcinoma and occult HBV infection","volume":"202","author":"Kaur","year":"2010","journal-title":"J. Infect. Dis."},{"key":"ref_89","doi-asserted-by":"crossref","first-page":"1177","DOI":"10.1002\/jmv.21525","article-title":"Evidence that methylation of hepatitis B virus covalently closed circular DNA in liver tissues of patients with chronic hepatitis B modulates HBV replication","volume":"81","author":"Guo","year":"2009","journal-title":"J. Med. Virol."},{"key":"ref_90","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1038\/nmeth.3733","article-title":"Editing the epigenome: Technologies for programmable transcriptional modulation and epigenetic regulation","volume":"13","author":"Thakore","year":"2016","journal-title":"Nature methods"},{"key":"ref_91","doi-asserted-by":"crossref","first-page":"1799","DOI":"10.1097\/QAD.0b013e32832ec1dc","article-title":"Expression of latent human immunodeficiency type 1 is induced by novel and selective histone deacetylase inhibitors","volume":"23","author":"Archin","year":"2009","journal-title":"AIDS"},{"key":"ref_92","doi-asserted-by":"crossref","first-page":"30","DOI":"10.1172\/JCI69738","article-title":"New and emerging HDAC inhibitors for cancer treatment","volume":"124","author":"West","year":"2014","journal-title":"J. Clin. Investig."},{"key":"ref_93","doi-asserted-by":"crossref","first-page":"4816","DOI":"10.1128\/jvi.74.10.4816-4823.2000","article-title":"Liver-specific alpha 2 interferon gene expression results in protection from induced hepatitis","volume":"74","author":"Aurisicchio","year":"2000","journal-title":"J. Virol."},{"key":"ref_94","doi-asserted-by":"crossref","first-page":"10111","DOI":"10.1128\/JVI.78.18.10111-10121.2004","article-title":"Helper-dependent adenoviral vector-mediated delivery of woodchuck-specific genes for alpha interferon (IFN-\u03b1) and IFN-\u03b3: IFN-\u03b1 but not IFN-\u03b3 reduces woodchuck hepatitis virus replication in chronic infection in vivo","volume":"78","author":"Fiedler","year":"2004","journal-title":"J. Virol."},{"key":"ref_95","doi-asserted-by":"crossref","first-page":"668","DOI":"10.1038\/sj.gt.3302449","article-title":"Liver-specific expression of interferon \u03b3 following adenoviral gene transfer controls hepatitis b virus replication in mice","volume":"12","author":"Dumortier","year":"2005","journal-title":"Gene Ther."},{"key":"ref_96","doi-asserted-by":"crossref","first-page":"68","DOI":"10.1016\/j.ymthe.2005.02.017","article-title":"IFN-\u03b1 gene therapy for woodchuck hepatitis with adeno-associated virus: Differences in duration of gene expression and antiviral activity using intraportal or intramuscular routes","volume":"12","author":"Berraondo","year":"2005","journal-title":"Mol. Ther."},{"key":"ref_97","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1016\/j.jhep.2015.02.048","article-title":"Liver-directed gene therapy of chronic hepadnavirus infection using interferon alpha tethered to apolipoprotein A-I","volume":"63","author":"Berraondo","year":"2015","journal-title":"J. Hepatol."},{"key":"ref_98","doi-asserted-by":"crossref","first-page":"2663","DOI":"10.1128\/JVI.02384-08","article-title":"Treatment of chronic viral hepatitis in woodchucks by prolonged intrahepatic expression of interleukin-12","volume":"83","author":"Crettaz","year":"2009","journal-title":"J. Virol."},{"key":"ref_99","doi-asserted-by":"crossref","first-page":"12266","DOI":"10.1128\/JVI.01597-09","article-title":"Semliki forest virus expressing interleukin-12 induces antiviral and antitumoral responses in woodchucks with chronic viral hepatitis and hepatocellular carcinoma","volume":"83","author":"Liu","year":"2009","journal-title":"J. Virol."},{"key":"ref_100","doi-asserted-by":"crossref","first-page":"8563","DOI":"10.1128\/JVI.01030-16","article-title":"Complementary effects of interleukin-15 and alpha interferon induce immunity in hepatitis b virus transgenic mice","volume":"90","author":"Otano","year":"2016","journal-title":"J. Virol."},{"key":"ref_101","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1053\/j.gastro.2007.11.002","article-title":"T cells redirected against hepatitis B virus surface proteins eliminate infected hepatocytes","volume":"134","author":"Bohne","year":"2008","journal-title":"Gastroenterology"},{"key":"ref_102","doi-asserted-by":"crossref","first-page":"103","DOI":"10.1016\/j.jhep.2010.10.025","article-title":"Engineering virus-specific T cells that target HBV infected hepatocytes and hepatocellular carcinoma cell lines","volume":"55","author":"Gehring","year":"2011","journal-title":"J. Hepatol."},{"key":"ref_103","doi-asserted-by":"crossref","first-page":"3177","DOI":"10.1172\/JCI93024","article-title":"Lymphocytes transiently expressing virus-specific T cell receptors reduce hepatitis B virus infection","volume":"127","author":"Kah","year":"2017","journal-title":"J. Clin. Investig."},{"key":"ref_104","doi-asserted-by":"crossref","first-page":"456","DOI":"10.1053\/j.gastro.2013.04.047","article-title":"T cells expressing a chimeric antigen receptor that binds hepatitis B virus envelope proteins control virus replication in mice","volume":"145","author":"Krebs","year":"2013","journal-title":"Gastroenterology"},{"key":"ref_105","doi-asserted-by":"crossref","unstructured":"Wisskirchen, K., Metzger, K., Schreiber, S., Asen, T., Weigand, L., Dargel, C., Witter, K., Kieback, E., Sprinzl, M.F., and Uckert, W. (2017). Isolation and functional characterization of hepatitis B virus-specific T-cell receptors as new tools for experimental and clinical use. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0182936"},{"key":"ref_106","doi-asserted-by":"crossref","first-page":"103","DOI":"10.1007\/s00430-014-0379-5","article-title":"Therapeutic vaccination and immunomodulation in the treatment of chronic hepatitis B: Preclinical studies in the woodchuck","volume":"204","author":"Kosinska","year":"2015","journal-title":"Med. Microbiol. Immunol."},{"key":"ref_107","doi-asserted-by":"crossref","first-page":"415","DOI":"10.1038\/nm.4313","article-title":"Refining strategies to translate genome editing to the clinic","volume":"23","author":"Cornu","year":"2017","journal-title":"Nat. Med."},{"key":"ref_108","doi-asserted-by":"crossref","unstructured":"Hardee, L.C., Ar\u00e9valo-Soliz, M.L., Hornstein, D.B., and Zechiedrich, L. (2017). Advances in non-viral DNA vectors for gene therapy. Genes, 8.","DOI":"10.3390\/genes8020065"},{"key":"ref_109","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1038\/gt.2016.90","article-title":"Delivering efficient liver-directed AAV-mediated gene therapy","volume":"24","author":"Baruteau","year":"2017","journal-title":"Gene Ther."},{"key":"ref_110","doi-asserted-by":"crossref","first-page":"475","DOI":"10.1038\/mt.2016.1","article-title":"Nuclease target site selection for maximizing on-target activity and minimizing off-target effects in genome editing","volume":"24","author":"Lee","year":"2016","journal-title":"Mol. Ther."}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/9\/4\/207\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T15:00:26Z","timestamp":1760194826000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/9\/4\/207"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2018,4,12]]},"references-count":110,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2018,4]]}},"alternative-id":["genes9040207"],"URL":"https:\/\/doi.org\/10.3390\/genes9040207","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2018,4,12]]}}}