Hybridization-mediated off-target effects of splice-switching antisense oligonucleotides

Scharner, J., Ma, W. K., Zhang, Q., Lin, K. T., Rigo, F., Bennett, C. F., Krainer, A. R. (December 2019) Hybridization-mediated off-target effects of splice-switching antisense oligonucleotides. Nucleic Acids Res, 48 (2). ISSN 0305-1048 (Public Dataset)

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URL: https://www.ncbi.nlm.nih.gov/pubmed/31802121
DOI: 10.1093/nar/gkz1132

Abstract

Splice-switching antisense oligonucleotides (ASOs), which bind specific RNA-target sequences and modulate pre-mRNA splicing by sterically blocking the binding of splicing factors to the pre-mRNA, are a promising therapeutic modality to treat a range of genetic diseases. ASOs are typically 15-25 nt long and considered to be highly specific towards their intended target sequence, typically elements that control exon definition and/or splice-site recognition. However, whether or not splice-modulating ASOs also induce hybridization-dependent mis-splicing of unintended targets has not been systematically studied. Here, we tested the in vitro effects of splice-modulating ASOs on 108 potential off-targets predicted on the basis of sequence complementarity, and identified 17 mis-splicing events for one of the ASOs tested. Based on analysis of data from two overlapping ASO sequences, we conclude that off-target effects are difficult to predict, and the choice of ASO chemistry influences the extent of off-target activity. The off-target events caused by the uniformly modified ASOs tested in this study were significantly reduced with mixed-chemistry ASOs of the same sequence. Furthermore, using shorter ASOs, combining two ASOs, and delivering ASOs by free uptake also reduced off-target activity. Finally, ASOs with strategically placed mismatches can be used to reduce unwanted off-target splicing events.

Item Type: Paper
Subjects: Investigative techniques and equipment > cloning > PCR
Investigative techniques and equipment > assays > cloning > PCR
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > antisense
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > mRNA dynamics
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > oligonucleotide
CSHL Authors:
Communities: CSHL labs > Krainer lab
Depositing User: Adrian Gomez
Date: 5 December 2019
Date Deposited: 11 Dec 2019 15:38
Last Modified: 22 Jan 2020 19:35
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  • Supplement: https://academic.oup.com/nar/advance-article/doi/10.1093/nar/gkz1132/5658446#supplementary-data
URI: https://repository.cshl.edu/id/eprint/38735

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