microRNAs (miRNAs) are small regulatory RNAs that are important in development and disease1,2,3 and therefore represent a potential new class of targets for therapeutic intervention4. Despite recent progress in silencing of miRNAs in rodents5,6, the development of effective and safe approaches for sequence-specific antagonism of miRNAs in vivo remains a significant scientific and therapeutic challenge. Moreover, there are no reports of miRNA antagonism in primates. Here we show that the simple systemic delivery of a unconjugated, PBS-formulated locked-nucleic-acid-modified oligonucleotide (LNA-antimiR) effectively antagonizes the liver-expressed miR-122 in non-human primates. Acute administration by intravenous injections of 3 or 10 mg kg-1 LNA-antimiR to African green monkeys resulted in uptake of the LNA-antimiR in the cytoplasm of primate hepatocytes and formation of stable heteroduplexes between the LNA-antimiR and miR-122. This was accompanied by depletion of mature miR-122 and dose-dependent lowering of plasma cholesterol. Efficient silencing of miR-122 was achieved in primates by three doses of 10 mg kg-1 LNA-antimiR, leading to a long-lasting and reversible decrease in total plasma cholesterol without any evidence for LNA-associated toxicities or histopathological changes in the study animals. Our findings demonstrate the utility of systemically administered LNA-antimiRs in exploring miRNA function in rodents and primates, and support the potential of these compounds as a new class of therapeutics for disease-associated miRNAs.
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All primate studies were performed at the St Kitts Biomedical Research Foundation, St Kitts, West Indies. We thank A. Konge, A. Koustrup, B. Nordbo, H. W. Høvring, J. J. Jørgensen, K. R. Nielsen, L. Bang, H. Brostrøm, O. Olsen, R. Sølberg, U. Steinmeier, J. Staruk, C. Kent, E. Nisbett, M. Struharik and R. Valles for technical assistance. This study was supported by grants from the Danish National Advanced Technology Foundation, the Danish Medical Research Council and the Lundbeck Foundation (to S.K.) and by grants from the National Institutes of Health (to P.S.). The Wilhelm Johannsen Centre for Functional Genome Research was established by the Danish National Research Foundation.
Author Contributions J.E. and M. Lindow contributed equally to this study. J.E., M. Lindow, M. Lawrence, U.B., S.O., M.H., A.P., S.S. and E.M.S. performed experiments and contributed data. H.F.H. performed the synthesis of all oligonucleotides. J.E., M. Lindow, M. Lawrence, M. Lindholm, H.F.H., P.K., S.G., P.S., E.M.S. and S.K. designed experiments and discussed the data. S.K. supervised the study and wrote the paper together with J.E. and M. Lindow.
J.E., M. Lindow, A.P., S.O., M. Lindholm, M.H., H.F.H., P.K., E.M.S. and S.K. are employees of Santaris Pharma, a clinical stage biopharmaceutical company that develops RNA-based therapeutics.
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Elmén, J., Lindow, M., Schütz, S. et al. LNA-mediated microRNA silencing in non-human primates. Nature 452, 896–899 (2008). https://doi.org/10.1038/nature06783
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