title: Hepatotoxicity of high affinity gapmer antisense oligonucleotides is mediated by RNase H1 dependent promiscuous reduction of very long pre-mRNA transcripts.
abstract: High affinity antisense oligonucleotides (ASOs) containing bicylic modifications (BNA) such as locked nucleic acid (LNA) designed to induce target RNA cleavage have been shown to have enhanced potency along with a higher propensity to cause hepatotoxicity. In order to understand the mechanism of this hepatotoxicity, transcriptional profiles were collected from the livers of mice treated with a panel of highly efficacious hepatotoxic or non-hepatotoxic LNA ASOs. We observed highly selective transcript knockdown in mice treated with non-hepatotoxic LNA ASOs, while the levels of many unintended transcripts were reduced in mice treated with hepatotoxic LNA ASOs. This transcriptional signature was concurrent with on-target RNA reduction and preceded transaminitis. Remarkably, the mRNA transcripts commonly reduced by toxic LNA ASOs were generally not strongly associated with any particular biological process, cellular component or functional group. However, they tended to have much longer pre-mRNA transcripts. We also demonstrate that the off-target RNA knockdown and hepatotoxicity is attenuated by RNase H1 knockdown, and that this effect can be generalized to high affinity modifications beyond LNA. This suggests that for a certain set of ASOs containing high affinity modifications such as LNA, hepatotoxicity can occur as a result of unintended off-target RNase H1 dependent RNA degradation.
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA sburel@isisph.com.
authors.forename: Sebastien A
authors.initials: SA
authors.lastname: Burel
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Hart
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Patrick
authors.initials: P
authors.lastname: Cauntay
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Jill
authors.initials: J
authors.lastname: Hsiao
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Todd
authors.initials: T
authors.lastname: Machemer
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Melanie
authors.initials: M
authors.lastname: Katz
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Andy
authors.initials: A
authors.lastname: Watt
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Huynh-Hoa
authors.initials: HH
authors.lastname: Bui
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Husam
authors.initials: H
authors.lastname: Younis
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Mahyar
authors.initials: M
authors.lastname: Sabripour
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Susan M
authors.initials: SM
authors.lastname: Freier
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Gene
authors.initials: G
authors.lastname: Hung
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Amy
authors.initials: A
authors.lastname: Dan
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: T P
authors.initials: TP
authors.lastname: Prakash
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Punit P
authors.initials: PP
authors.lastname: Seth
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Eric E
authors.initials: EE
authors.lastname: Swayze
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: C Frank
authors.initials: CF
authors.lastname: Bennett
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Stanley T
authors.initials: ST
authors.lastname: Crooke
authors.affiliation: Isis Pharmaceuticals, Carlsbad, CA 921010, USA.
authors.forename: Scott P
authors.initials: SP
authors.lastname: Henry
mesh_terms: D000410:Alanine Transaminase / Q000097:blood / Q000235:genetics; D000818:Animals; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D046228:Microarray Analysis; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism / Q000633:toxicity*; D012322:RNA Precursors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D016914:Ribonuclease H / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D059467:Transcriptome / Q000187:drug effects
chemical_list: D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D012322:RNA Precursors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C477371:locked nucleic acid; D000410:Alanine Transaminase; D016914:Ribonuclease H; C078856:ribonuclease HI