Controlling miRNA-like off-target effects of an siRNA with nucleobase modifications

Scott R. Suter, Alexi Ball-Jones, Madeline M. Mumbleau, Rachel Valenzuela, Jose Ibarra-Soza, Hassan Owens, Andrew J Fisher, Peter A. Beal

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

SiRNAs can cause unintended gene silencing due to miRNA-like effects because of the similarity in function of an siRNA guide strand and a miRNA. Here we evaluate the effect on miRNA-like off targeting of introducing the adenosine derivative 7-EAA and triazoles prepared from 7-EAA at different positions in an siRNA guide strand. We find that a sterically demanding triazole placed in the RNA duplex major groove at position six of the guide strand dramatically reduces miRNA-like off targeting potency. A high-resolution structure of an RNA duplex bearing a novel, major-groove localized triazole is reported, which suggests that modified triazoles could be disrupting the hAgo2-guide-target RNA ternary complex. Five different triazole modifications were tested at the guide strand 6-position for effects on on-target and miRNA-like off target knockdown potency. A 7-EAA triazole bearing a benzylamine substituent displayed on-target knockdown activity as potent as the native siRNA, while having an IC50 against a miRNA-like off target >100-fold higher. Melting temperature studies revealed no obvious correlation between potency in knockdown assays and a modification's effect on duplex stability. These results, along with known structures of hAgo2-guide-target ternary complexes, are used to rationalize the effect of 7-EAA triazoles on miRNA-like off target effects.

Original languageEnglish (US)
Pages (from-to)10029-10036
Number of pages8
JournalOrganic and Biomolecular Chemistry
Volume15
Issue number47
DOIs
StatePublished - Jan 1 2017

ASJC Scopus subject areas

  • Biochemistry
  • Physical and Theoretical Chemistry
  • Organic Chemistry

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    Suter, S. R., Ball-Jones, A., Mumbleau, M. M., Valenzuela, R., Ibarra-Soza, J., Owens, H., Fisher, A. J., & Beal, P. A. (2017). Controlling miRNA-like off-target effects of an siRNA with nucleobase modifications. Organic and Biomolecular Chemistry, 15(47), 10029-10036. https://doi.org/10.1039/c7ob02654d