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A Highly Effective and Long-Lasting Inhibition of miRNAs with PNA-Based Antisense Oligonucleotides

  • Oh, Su Young (Panagene Inc.) ;
  • Ju, YeongSoon (Panagene Inc.) ;
  • Park, Heekyung (Panagene Inc.)
  • Received : 2009.04.28
  • Accepted : 2009.08.26
  • Published : 2009.10.31

Abstract

MiRNAs are non-coding RNAs that play a role in the regulation of major processes. The inhibition of miRNAs using antisense oligonucleotides (ASOs) is a unique and effective technique for the characterization and subsequent therapeutic targeting of miRNA function. Recent advances in ASO chemistry have been used to increase both the resistance to nucleases and the target affinity and specificity of these ASOs. Peptide nucleic acids (PNAs) are artificial oligonucleotides constructed on a peptide-like backbone. PNAs have a stronger affinity and greater specificity to DNA or RNA than natural nucleic acids and are resistant to nucleases, which is an essential characteristic for a miRNA inhibitor that will be exposed to serum and cellular nucleases. For increasing cell penetration, PNAs were conjugated with cell penetrating peptides (CPPs) at N-terminal. Among the tested CPPs, Tat-modified peptide-conjugated PNAs have most effective function for miRNA inhibition. PNA-based ASO was more effective miRNA inhibitor than other DNA-based ASOs and did not show cytotoxicity at concentration up to 1,000 nM. The effects of PNA-based ASOs were shown to persist for 9 days. Also, PNA-based ASOs showed considerable stability at storage temperature. These results suggest that PNA-based ASOs are more effective ASOs of miRNA than DNA-based ASOs and PNA-based ASO technology, compared with other technologies used to inhibit miRNA activity can be an effective tool for investigating miRNA functions.

Keywords

Acknowledgement

Supported by : Small and Medium Business Administration (SMBA)

References

  1. Abes, S., Turner, J.J., Ivanova, G.D., Owen, D., Williams, D., Arzumanov, A., Clair, P., Gait, M.J., and Lebleu, B. (2007). Efficient splicing correction by PNA conjugation to an R6-penetratin delivery peptide. Nucleic Acids Res. 35, 4495-4502 https://doi.org/10.1093/nar/gkm418
  2. Boutla, A., Delidakis, C., and Tabler, M. (2003). Developmental defects by antisense-mediated inactivation of micro-RNAs 2 and 13 in Drosophila and the identification of putative target genes. Nucleic Acids Res. 31, 4973-4980 https://doi.org/10.1093/nar/gkg707
  3. Braasch, D.A., and Corey, D.R. (2002). Novel antisense and peptide nucleic acid strategies for controlling gene expression. Biochemistry 41, 4503-4510 https://doi.org/10.1021/bi0122112
  4. Bushati, N., and Cohe, S.M. (2007). microRNA functions. Annu. Rev. Cell Dev. Biol. 23, 175-205 https://doi.org/10.1146/annurev.cellbio.23.090506.123406
  5. Davis, S., Lollo, B., Freier, S., and Esau, C. (2006). Improved targeting of miRNA with antisense oligonucleotides. Nucleic Acids Res. 34, 2294-2304 https://doi.org/10.1093/nar/gkl183
  6. Drygin, D., Barone, S., and Bennett, F. (2007). Sequence-dependent cytotoxicity of second-generation oligonucleotides. Nucleic Acids Res. 32, 6585-6594 https://doi.org/10.1093/nar/gkh997
  7. Esau, C.C. (2008). Inhibition of microRNA with antisense oligonucleotides. Methods 44, 55-60 https://doi.org/10.1016/j.ymeth.2007.11.001
  8. Esquela-Kerscher, A., and Slack, F.J. (2006). Oncomirs-microRNAs with a role in cancer. Nature Rev. 6, 259-269 https://doi.org/10.1038/nrc1840
  9. Fabani, M.M., and Gait, M.J. (2008). miR-122 targeting with LNA/2′- O-methyl oligonucleotide mixmers, peptide nucleic acids (PNA), and PNA-peptide conjugates. RNA 14, 1-11
  10. Kang, I.G., Hwang, Y.J., Ha, J.S., Kim, D.G., and Kim, S.T. (2005). Expression of apoptosis, Bax and Bcl-2 in nasal polyps. J. Clin. Otolaryngol. 16, 264-269
  11. Kim, V.N. (2005). Small RNAs: classification, biogenesis, and function. Mol. Cells 19, 1-15 https://doi.org/10.1016/j.molcel.2005.05.026
  12. Koning, M.C., Marel, G.A., and Overhand, M. (2003). Synthetic developments towards PNA-peptide conjugates. Curr. Opin. Chem. Biol. 7, 734-740 https://doi.org/10.1016/j.cbpa.2003.10.006
  13. Larsen, H.J., Bentin, T., and Nielsen, P.E. (1999). Antisense properties of peptide nucleic acid. Biochim. Biophy. Acta 1489, 159-166 https://doi.org/10.1016/S0167-4781(99)00145-1
  14. Lebedeva, I., and Stein, C.A. (2001). Antisense oligonucleotides: promise and reality. Annu. Rev. Pharmacol. Toxicol. 41, 403-419 https://doi.org/10.1146/annurev.pharmtox.41.1.403
  15. Lee, H., Jeon, J.H., Lim, J.C., Choi, H., Yoon, Y., and Kim, S.K. (2007). Peptide nucleic acid synthesis by novel amide formation. Org. Lett. 9, 3291-3293 https://doi.org/10.1021/ol071215h
  16. Naguibneva, I., Ameyar-Zazoua, M., Nonne, N., Polesskaya, A., Ait-Si-Ali, S., Groisman, R., Souidi, M., Pritchard, L.L., and Harel-Bellan, A. (2006). An LNA-based loss-of-function assay for micro-RNAs. Biomed. Pharm. 60, 633-638 https://doi.org/10.1016/j.biopha.2006.07.078
  17. Orom, U.A., Kauppinen, S., and Lund, A.H. (2006). LNA-midified oligonucleotides mediate specific inhibition of microRNA function. Gene 372, 137-141 https://doi.org/10.1016/j.gene.2005.12.031
  18. Pellestor, F., and Paulasova, P. (2004). The peptide nucleic acids, efficient tools for molecular diagnosis (Review). Int. J. Mol. Med. 13, 521-525
  19. Rayburn, E.R., and Zhang, R. (2008). Antisense, RNAi, and gene silencing strategies for therapy: Mission possible or impossible? Drug Discov. Today 13, 513-521 https://doi.org/10.1016/j.drudis.2008.03.014
  20. Swayze, E.E., Siwkowski, A.M., Wancewicz, E.V., Migawa, M.T., Wyrzykiewicz, T.K., Hung, G., Monia, B.P., and Bennett, C.F. (2007). Antisense oligonucleotides containing locked nucleic acid improve potency but cause significant hepatotoxicity in animals. Nucleic Acids Res. 35, 687-700 https://doi.org/10.1093/nar/gkl1071
  21. Uhlmann, E., Will, D.W., Breipohl, G., Langner, D., and Ryte, A. (1996). Synthesis and properties of PNA/DNA chimeras. Angew. Chem. Int. Ed. Engl. 35, 2632-2635 https://doi.org/10.1002/anie.199626321
  22. Wang, B., Doench., J.G., and Novina, C.D. (2007). Analysis of microRNA effector functions in vitro. Methods 43, 91-104 https://doi.org/10.1016/j.ymeth.2007.04.003
  23. Watts, J.K., Deleavey, G.F., and Damha, M.J. (2008). Chemically modified siRNA: tools and applications. Drug Discov. Today 13, 842-855 https://doi.org/10.1016/j.drudis.2008.05.007
  24. Weiler, J., Hunziker, J., and Hall, J. (2006). Anti-miRNA oligonucleotides (AMOs): ammunition to target miRNAs implicated in human disease? Gene Ther. 13, 496-502 https://doi.org/10.1038/sj.gt.3302654
  25. Wittung, P., Kajanus, J., Edwards, K., Haaima, G., Nielsen, P.E., Norden, B., and Malmstrom B.G. (1995). Phospholipid membrane permeability of peptide nucleic acid. FEBS Lett. 375, 27-29 https://doi.org/10.1016/0014-5793(95)01153-6
  26. Xia, L., Zhang, D., Du, R., Pan, Y., Zhao, L., Sun, S., Hong, L., Liu, J., and Fan, D. (2008). miR-15b and miR-16 modulate multidrug resistance by targeting BCL2 in human gastric cancer cells. Int. J. Cancer 123, 372-379 https://doi.org/10.1002/ijc.23501

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  30. Toward conditional control of Smac mimetic activity by RNA‐templated reduction of azidopeptides on PNA or 2′‐OMe‐RNA vol.112, pp.12, 2009, https://doi.org/10.1002/bip.23466