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RNA-sequencing-based off-target analysis of allele-specific siRNA in keratitis-ichthyosis-deafness syndrome patient-derived keratinocytes

ABSTRACT: Purpose: RNA-sequencing (RNA-Seq)-based transcriptomic analysis has facilitated identification of off-target effects of drug of interest. The goals of this study are to investigate off-target effects of the therapeutic, allele-specific siRNA, S7. Methods: mRNA profiles of KID syndrome patient-derived keratinocytes (with c.148G>A mutation in GJB2) with or without S7 treatment were generated by RNA-Seq, in triplicate, using an Illumina NextSeq 500 instrument. The sequence reads that passed quality filters were processed using RNA-STAR (v2.5b) and BioConductor SARTools package (v1.3.2). Real-time qRT–PCR validation was performed using SYBR Green. Results: We mapped about 15 million sequence reads per sample to the human genome (build hg38) and identified expression of 19,667 genes in the samples. Only 6 genes (MMP10, MMP1, NSA2, AFAP1L1, GLB1L2, GPR137) showed differential expression between the treated and untreated cells, with |log2 fold change| ≥ 1 and adjusted p-value (padj) <0.05. Altered expression of the top 10 genes (TMEM109, MMP9, ANGPTL4, CXCL5, in addition to the 6 genes above) was confirmed using real-time qRT–PCR. Conclusions: Our RNA-Seq findings suggest low-level off-target effects of S7, the allele-specific siRNA targeted to the most common GJB2 mutant associated with KID syndrome.

ORGANISM(S): Homo sapiens

PROVIDER: GSE131709 | GEO | 2022/05/23

REPOSITORIES: GEO

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