{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T00:59:27Z","timestamp":1760057967495,"version":"build-2065373602"},"reference-count":54,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2025,3,5]],"date-time":"2025-03-05T00:00:00Z","timestamp":1741132800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"South African Medical Research Council"},{"name":"Poliomyelitis Research Foundation"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Microorganisms"],"abstract":"<jats:p>Available interventions for the management of chronic hepatitis B (hepB) exhibit limited efficacy and barriers to vaccination against the hepatitis B virus (HBV) have hampered prophylaxis programmes. Development of potent therapeutics capable of functional cure of chronic hepB thus remains a relevant medical objective. RNA interference (RNAi) can be exploited to effect potent and specific silencing of target genes through the introduction of RNA sequences that mimic the natural activators of the pathway. To achieve a therapeutic effect, artificial primary microRNAs (pri-miRNAs) have been used extensively to target various viruses, including HBV. To date artificial pri-miRNAs have exclusively been produced from DNA expression cassettes. Although this achieves impressive silencing, eventual translation of this platform to the clinic is complicated by the requirement for viral vectors to deliver DNA. Consequently, clinical translation has been slow. Recently, the use of in vitro transcribed RNA, specifically to produce mRNA vaccines at industrial scale, has gained significant interest. We therefore sought to evaluate the feasibility of using in vitro transcribed artificial pri-miRNAs for the inhibition of HBV gene expression. Artificial HBV-targeting pri-miR-31 sequences, which are highly effective when expressed in cells from a DNA template, demonstrated modest silencing of viral replication when incorporated into mRNA that was transcribed in vitro. Off-target effects were also observed. Characterisation revealed that intracellular processing of the artificial pri-miRNAs was inefficient and non-specific effects were caused by stimulation of the interferon response. Nevertheless, optimised nuclear delivery of the artificial pri-miRNAs should improve their processing and achieve better anti-hepB efficacy.<\/jats:p>","DOI":"10.3390\/microorganisms13030604","type":"journal-article","created":{"date-parts":[[2025,3,5]],"date-time":"2025-03-05T09:37:52Z","timestamp":1741167472000},"page":"604","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["In Vitro Transcribed Artificial Primary MicroRNA for the Inhibition of Hepatitis B Virus Gene Expression in Cultured Cells"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1987-4942","authenticated-orcid":false,"given":"Creanne","family":"Shrilall","sequence":"first","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, Infectious Diseases and Oncology Research Institute (IDORI), Faculty of Health Sciences, University of the Witwatersrand, Johannesburg 2050, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"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, Infectious Diseases and Oncology Research Institute (IDORI), Faculty of Health Sciences, University of the Witwatersrand, Johannesburg 2050, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1205-2436","authenticated-orcid":false,"given":"Abdullah","family":"Ely","sequence":"additional","affiliation":[{"name":"Wits\/SAMRC Antiviral Gene Therapy Research Unit, Infectious Diseases and Oncology Research Institute (IDORI), Faculty of Health Sciences, University of the Witwatersrand, Johannesburg 2050, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,3,5]]},"reference":[{"key":"ref_1","unstructured":"World Health Organization (2017). 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