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Overcoming GNA/RNA base-pairing limitations using isonucleotides improves the pharmacodynamic activity of ESC+ GalNAc-siRNAs.

Authors :
Schlegel MK
Matsuda S
Brown CR
Harp JM
Barry JD
Berman D
Castoreno A
Schofield S
Szeto J
Manoharan M
Charissé K
Egli M
Maier MA
Source :
Nucleic acids research [Nucleic Acids Res] 2021 Nov 08; Vol. 49 (19), pp. 10851-10867.
Publication Year :
2021

Abstract

We recently reported that RNAi-mediated off-target effects are important drivers of the hepatotoxicity observed for a subset of GalNAc-siRNA conjugates in rodents, and that these findings could be mitigated by seed-pairing destabilization using a single GNA nucleotide placed within the seed region of the guide strand. Here, we report further investigation of the unique and poorly understood GNA/RNA cross-pairing behavior to better inform GNA-containing siRNA design. A reexamination of published GNA homoduplex crystal structures, along with a novel structure containing a single (S)-GNA-A residue in duplex RNA, indicated that GNA nucleotides universally adopt a rotated nucleobase orientation within all duplex contexts. Such an orientation strongly affects GNA-C and GNA-G but not GNA-A or GNA-T pairing in GNA/RNA heteroduplexes. Transposition of the hydrogen-bond donor/acceptor pairs using the novel (S)-GNA-isocytidine and -isoguanosine nucleotides could rescue productive base-pairing with the complementary G or C ribonucleotides, respectively. GalNAc-siRNAs containing these GNA isonucleotides showed an improved in vitro activity, a similar improvement in off-target profile, and maintained in vivo activity and guide strand liver levels more consistent with the parent siRNAs than those modified with isomeric GNA-C or -G, thereby expanding our toolbox for the design of siRNAs with minimized off-target activity.<br /> (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Details

Language :
English
ISSN :
1362-4962
Volume :
49
Issue :
19
Database :
MEDLINE
Journal :
Nucleic acids research
Publication Type :
Academic Journal
Accession number :
34648028
Full Text :
https://doi.org/10.1093/nar/gkab916