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Alteration of target cleavage patterns and off-target reduction of antisense oligonucleotides incorporating 2-N-carbamoyl- or (2-pyridyl)guanine [Rluc]

ABSTRACT: We performed microarrays to evaluate the gene expression changes after 24 h of antisense oligonucleotide transfection.

ORGANISM(S): Homo sapiens

PROVIDER: GSE228741 | GEO | 2023/06/14

REPOSITORIES: GEO

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Alteration of target cleavage patterns and off-target reduction of antisense oligonucleotides incorporating 2-N-carbamoyl- or (2-pyridyl)guanine [MALAT]

Project description:We performed microarrays to evaluate the gene expression changes after 24 h of antisense oligonucleotide transfection.

2023-06-14 | GSE228740 | GEO

Alteration of target cleavage patterns and off-target reduction of antisense oligonucleotides incorporating 2-N-carbamoyl- or (2-pyridyl)guanine

Project description:This SuperSeries is composed of the SubSeries listed below.

2023-06-14 | GSE228747 | GEO

Reduction of Off-Target Effects of Gapmer Antisense Oligonucleotides by Oligonucleotide Extension.

Project description:AimAntisense oligonucleotide (ASO) has the potential to induce off-target effects by inadvertent binding of ASOs to unintended RNAs that have a sequence similar to the target RNA. In the present study, we focused on the association between oligonucleotide length and off-target effects. Oligonucleotide extension is assumed to have bilateral effects on hybridization-dependent changes in gene expression, i.e., one is the decrease of off-target effects based on the reduced number of off-target candidate genes with perfect matches, and the other is the increase of off-target effects based on the increased binding affinity between the ASO and the complementary RNAs that leads to better tolerability for mismatches.MethodsTo determine the effects of oligonucleotide extension of gapmer ASOs on off-target effects, an extensive microarray analysis was performed using human cells treated with a 14-mer gapmer ASO and the extended 18-mer derivatives with the same core 14-mer region.Results and discussionOur data indicated that change in gene expression in the cells treated with 18-mer ASOs was significantly smaller than those with a 14-mer ASO, showing the decrease of off-target effects by oligonucleotide extension.

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RNA-seq analysis of HeLa cells treated with GapmeR antisense oligonucleotides

Project description:The experiment was carried out to provide insights into biological function of a newly identified long noncoding RNA, PNCTR. HeLa cells were transfected with either a PNCTR-specific GapmeR antisense oligonucleotide or a non-targeting GapmeR control. Total RNAs were extracted at 24 hours post transfection, QC'd and hybridized with oligo(dT) magnetic beads to isolate the poly(A) RNA fraction. Stranded mRNA sequencing libraries were then prepared using the Illumina TruSeq Stranded mRNA Library Kit and sequenced using an Illumina HiSeq 2500 instrument.

2018-10-12 | E-MTAB-6529 | biostudies-arrayexpress

Evaluation of off-target effects of gapmer antisense oligonucleotides using human cells

Project description:Antisense oligonucleotide (ASO) has the potential to induce off-target effects due to complementary binding between the ASO and unintended RNA with a sequence similar to the target RNA. In this study, we evaluated off-target effects of gapmer ASOs, which cleave the target RNA in an RNase H-dependent manner, by introducing the ASO into human cells and performing microarray analysis. Our data indicate that gapmer ASOs induce off-target effects depending on the degree of complementarity between the ASO and off-target candidate genes. Based on our results, we also propose a scheme for assessment of off-target effects of gapmer ASOs.

2021-12-08 | GSE190473 | GEO

Targeting Smyd3 by antisense oligonucleotides attenuates liver tumor growth

Project description:We investigated whether pharamacological inhibition of Smyd3 by antisense oligonucleotides (ASOs) can influence diethylnitrosamine (DEN)-induced liver cancer development. Our phenotypic analyses, among others, included gene expression profiling.

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Antisense oligonucleotides capable of promoting specific target mRNA reduction via competing RNase H1-dependent and independent mechanisms.

Project description:Antisense oligonucleotides (ASOs) are most commonly designed to reduce targeted RNA via RNase H1-dependent degradation. In this paper we demonstrate that cellular proteins can compete for sites targeted by RNase H1-dependent ASOs. We further show that some ASOs designed to mediate RNase H1 cleavage can, in certain instances, promote target reduction both by RNase H1-mediated cleavage and by steric inhibition of binding of splicing factors at a site required for efficient processing of the pre-mRNA. In the latter case, RNase H cleavage was prevented by binding of a second protein, HSPA8, to the ASO/pre-mRNA heteroduplex. In addition, using a precisely controlled minigene system, we directly demonstrated that activity of ASOs targeting sites in introns is strongly influenced by splicing efficiency.

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Antisense inhibition of gene expression in cells by oligonucleotides incorporating locked nucleic acids: effect of mRNA target sequence and chimera design.

Project description:Use of antisense oligonucleotides is a versatile strategy for achieving control of gene expression. Unfortunately, the interpretation of antisense-induced phenotypes is sometimes difficult, and chemical modifications that improve the potency and specificity of antisense action would be useful. The introduction of locked nucleic acid (LNA) bases into oligonucleotides confers exceptional improvement in binding affinity, up to 10 degrees C per substitution, making LNAs an exciting option for the optimization of antisense efficacy. Here we examine the rules governing antisense gene inhibition within cells by oligonucleotides that contain LNA bases. LNA- containing oligomers were transfected into cells using cationic lipid and accumulated in the nucleus. We tested antisense gene inhibition by LNAs and LNA-DNA chimeras complementary to the 5'-untranslated region, the region surrounding the start codon and the coding region of mRNA, and identified effective antisense agents targeted to each of these locations. Our data suggest that LNA bases can be used to develop antisense oligonucleotides and that their use is a versatile approach for efficiently inhibiting gene expression inside cells.

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Expression data from dosing four different antisense oligonucleotides in mice II

Project description:We used microarrays to globally profile the gene expression changes observed in liver after 3 days when dosing antisense oligonucleotides in mice

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

Project description: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.

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