Elsevier

Thrombosis Research

Volume 136, Issue 2, August 2015, Pages 367-370
Thrombosis Research

Full Length Article
Could MicroRNA polymorphisms influence warfarin dosing? A pharmacogenetics study on mir133 genes

https://doi.org/10.1016/j.thromres.2015.06.026Get rights and content

Highlights

  • MiRNAs play a role in regulation of physiological and pathological processes

  • VKORC1 (the target of warfarin treatment) is possibly regulated by mir-133.

  • We identified 4 SNPs in MIR133A2 gene and 1 SNP in MIR133B gene

  • RS45547937 in MIR133A2 gene seems to be involved in warfarin dosing variability

Abstract

MicroRNAs are small single stranded molecules that play a crucial role in regulation of physiological and pathological processes. Recent studies showed that VKORC1 gene contains an highly evolutionary conserved binding site for mir-133. Moreover, in human hepatocytes mir-133 is constitutively co-expressed with VKORC1. Since VKORC1 protein is the target of warfarin treatment, the aim of this study was to verify if genetic variations in MIR133A1, MIR133A2 and MIR133B could contribute to warfarin dose variability. By direct sequencing, we identified 4 SNPs in MIR133A2 gene and 1 SNP in MIR133B gene. Three SNPs in MIR133A2 were in complete linkage disequilibrium and correlated with warfarin dose: indeed, for each SNP, patients carrying the GA or AA genotype required a MWWD significantly higher than the wildtype genotype (P = 0.019). We also inferred the haplotypes in MIR133A2 gene. The GC haplotype required a MWWD significantly lower than AT haplotype (P = 0.012). The multiple linear regression analysis confirmed that rs45547937 (as tag SNP) in MIR133A2 could be involved in warfarin dosing variability, (P = 0.016). These results seem to suggest that also polymorphisms in miRNA precursors may potentially affects drug response variability.

Introduction

Recent advances in genomic technologies and bioinformatics approaches have had a significant impact on microRNA research. MicroRNAs (miRNAs or miRs) are a recently discovered class of small (19–24 nucleotides) endogenous non-coding oligonucleotides that regulate gene expression at the post-transcriptional level [1], [2], [3]. They play a crucial role in regulation of physiological and pathological processes and some miRNAs have been described as associated with distinct clinical characteristics, such as diagnosis or disease activity [4], [5], [6], [7], [8], [9]. For these reasons, miRNAs are attractive as potential biomarkers for the diagnosis, prognosis, disease activity and severity of various diseases.

By using bioinformatic tools, Shomron [10] identified two highly evolutionary conserved binding sites for mir-133 and mir-137 on VKORC1 gene, suggesting that VKORC1 is possibly regulated by these miRNAs. VKORC1 is the molecular target of vitamin k anticoagulants, such as warfarin. Polymorphisms in VKORC1 gene had been described associated with warfarin dose requirement [11], [12], [13], [14]. Recently, Perez-Andreu et al. [15] confirmed the potential binding sites for mir-133 and mir-137, but showed that only mir-133 is constitutively co-expressed with VKORC1 in human hepatocytes. They also showed that an over-expression of mir-133 decrease the VKORC1 mRNA expression in a dose dependent-manner. Moreover, this regulation of VKORC1 mRNA expression had also been observed in subjects carrying the G allele (rs9923231 in the promoter region), that is associated with a higher transcription rate of VKORC1 mRNA. Therefore, it is possible to speculate that variations in mir-133 genes, leading to an aberrant regulation of VKORC1 gene, could alter the efficacy of warfarin treatment.

The aim of our study was to detect the genetic variability in genes coding for mir-133 and to evaluate a possible correlation of such variability with warfarin dosing requirement. At this purpose we performed a full sequencing of MIR133A1, MIR133A2 and MIR133B genes in a sample of patients treated with warfarin and we evaluated whether genetic variants in these genes could influence the required dose.

Section snippets

Patients

Patients were consecutively recruited at the Centre of Haemostasis and Thrombosis of the PTV (Policlinico Tor Vergata, Rome, Italy). After obtaining written informed consent from all individuals, a blood sample from 205 patients was collected for genetic analysis.

Stable anticoagulation was achieved once the patients INR (International Normalized Ratio) values were within the therapeutic range (tINR: 2–3 for atrial fibrillation and deep vein thrombosis, 2.5-3.5 for aortic valve prosthesis, 3–4

Results

Three genes, MIR133A1, MIR133A2 and MIR133B, coding for the mature mir-133 were studied by direct sequencing. We identified 4 SNPs in MIR133A2 gene and 1 SNP in MIR133B gene. All the identified variations had already been described and located in the upstream or downstream region of the pre-miR. MIR133A1 was found monomorphic in our sample. Location and allele frequencies for all identified variants are listed in Table 1. The rs45547937, rs13040566 and rs13040413 SNPs in MIR133A2 were found in

Discussion

MiRNAs are a class of small RNA that play an important regulatory role in a plethora of pathways and that can be implicated in diseases development and progression. It is evident that miRNAs could control also the expression of genes that are involved in drug response and therefore they could also play a role in Pharmacogenomics. Several studies have described the role of miRNA in drug response especially in cancer treatments in terms of drug resistance [18], [19], [20] but also in inflammatory

Authorship

CC designed the study, performed the statistical analysis and wrote the manuscript, SR and CP performed the genetic analysis, GN revised the manuscript, VF provided the sample collection, PB supervised the study and wrote the manuscript.

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