title: siRNA off-target effects can be reduced at concentrations that match their individual potency.
abstract: Small interfering RNAs (siRNAs) are routinely used to reduce mRNA levels for a specific gene with the goal of studying its function. Several studies have demonstrated that siRNAs are not always specific and can have many off-target effects. The 3' UTRs of off-target mRNAs are often enriched in sequences that are complementary to the seed-region of the siRNA. We demonstrate that siRNA off-targets can be significantly reduced when cells are treated with a dose of siRNA that is relatively low (e.g. 1 nM), but sufficient to effectively silence the intended target. The reduction in off-targets was demonstrated for both modified and unmodified siRNAs that targeted either STAT3 or hexokinase II. Low concentrations reduced silencing of transcripts with complementarity to the seed region of the siRNA. Similarly, off-targets that were not complementary to the siRNA were reduced at lower doses, including up-regulated genes that are involved in immune response. Importantly, the unintended induction of caspase activity following treatment with a siRNA that targeted hexokinase II was also shown to be a concentration-dependent off-target effect. We conclude that off-targets and their related phenotypic effects can be reduced for certain siRNA that potently silence their intended target at low concentrations.
authors.affiliation: Pfizer Inc, Cambridge, Massachusetts, United States of America. Daniel.Caffrey@umassmed.edu
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Caffrey
authors.forename: Juan
authors.initials: J
authors.lastname: Zhao
authors.forename: Zhili
authors.initials: Z
authors.lastname: Song
authors.forename: Michael E
authors.initials: ME
authors.lastname: Schaffer
authors.forename: Steven A
authors.initials: SA
authors.lastname: Haney
authors.forename: Romesh R
authors.initials: RR
authors.lastname: Subramanian
authors.forename: Albert B
authors.initials: AB
authors.lastname: Seymour
authors.forename: Jason D
authors.initials: JD
authors.lastname: Hughes
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D001483:Base Sequence; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D020869:Gene Expression Profiling; D006593:Hexokinase / Q000235:genetics*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050796:STAT3 Transcription Factor / Q000235:genetics*
chemical_list: D020413:3' Untranslated Regions; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D050796:STAT3 Transcription Factor; D006593:Hexokinase; D053148:Caspase 3; D053179:Caspase 7