CYP2C9 SNPs Research Articles

Identification of the effects of pathogenic genetic variations of human CYP2C9 and CYP2D6: an in silico approach

Genetic variations in the human cytochrome P450 family 2 subfamily C member 9 (CYP2C9) and cytochrome P450 family 2 subfamily D member 6 (CYP2D6) genes may affect drug metabolism and lead to alterations in phenotypes. Genetic variations are associated with toxicity, adverse drug reactions, inefficient treatment. Various in silico tools were combined to investigate the deleterious effects of missense non-synonymous single nucleotide polymorphisms (nsSNPs) of the human CYP2C9 and CYP2D6. The structural and functional effects of the high-risk non-synonymous SNPs in the human CYP2C9 and CYP2D6 were predicted by numerous computational mutation analysis methods. Out of 24 pathogenic missense SNPs in the CYP2C9, 22 nsSNPs had a decreasing effect on protein stability and 13 SNPs were showed to be located at conserved positions. Out of 27 high-risk deleterious non-synonymous SNPs in the human CYP2D6, 21 SNPs decreased protein stability and 16 nsSNPs were predicted to be positioned at conserved regions. Our present study suggests that the identified functional SNPs may affect drug metabolism associated with CYP2C9 and CYP2D6 enzymes.

PREDICTION OF THE EFFECT OF SINGLE NUCLEOTIDE POLYMORPHISMS (SNPS) IN THE CYP2C9 ON WARFARIN METABOLISM BY IN SILICO STUDY

Objective: This study aimed to predict the effect of SNPs CYP2C9 s on the metabolic activity of S-warfarin in the body.
 Methods: Molecular modeling was performed to obtain SNPs CYP2C9 and molecular docking was performed to predict the effect of SNPs CYP2C9 on the metabolic activity of S-warfarin.
 Results: The results showed that wild-type CYP2C9 had the strongest binding affinity (∆G: -9.76 kcal/mol), indicating that wild-type CYP2C9 had the best metabolic activity compared to SNPs CYP2C9. There was a decrease in hydrogen bond formation in SNPs CYP2C9 and an increase in the distance between C7 S-warfarin and Fe-Heme in CYP2C9 SNPs when compared to wild-type CYP2C9
 Conclusion: The decrease in binding affinity, decrease in hydrogen bond formation, and an increase in the distance between C7 S-warfarin and Fe-Heme on SNPs CYP2C9 indicated that SNPs CYP2C9 had decreased metabolic activity against S-warfarin which led to an increased risk of bleeding.

Steroid hormone-related polymorphisms associate with the development of bone erosions in rheumatoid arthritis and help to predict disease progression: Results from the REPAIR consortium

Here, we assessed whether 41 SNPs within steroid hormone genes associated with erosive disease. The most relevant finding was the rheumatoid factor (RF)-specific effect of the CYP1B1, CYP2C9, ESR2, FcγR3A, and SHBG SNPs to modulate the risk of bone erosions (P = 0.004, 0.0007, 0.0002, 0.013 and 0.015) that was confirmed through meta-analysis of our data with those from the DREAM registry (P = 0.000081, 0.0022, 0.00074, 0.0067 and 0.0087, respectively). Mechanistically, we also found a gender-specific correlation of the CYP2C9rs1799853T/T genotype with serum vitamin D3 levels (P = 0.00085) and a modest effect on IL1β levels after stimulation of PBMCs or blood with LPS and PHA (P = 0.0057 and P = 0.0058). An overall haplotype analysis also showed an association of 3 ESR1 haplotypes with a reduced risk of erosive arthritis (P = 0.009, P = 0.002, and P = 0.002). Furthermore, we observed that the ESR2, ESR1 and FcγR3A SNPs influenced the immune response after stimulation of PBMCs or macrophages with LPS or Pam3Cys (P = 0.002, 0.0008, 0.0011 and 1.97•10−7). Finally, we found that a model built with steroid hormone-related SNPs significantly improved the prediction of erosive disease in seropositive patients (PRF+ = 2.46•10−8) whereas no prediction was detected in seronegative patients (PRF− = 0.36). Although the predictive ability of the model was substantially lower in the replication population (PRF+ = 0.014), we could confirm that CYP1B1 and CYP2C9 SNPs help to predict erosive disease in seropositive patients. These results are the first to suggest a RF-specific association of steroid hormone-related polymorphisms with erosive disease.

Abstract 18405: Warfarin Dose Requirements and Polymorphisms in MicroRNA and Nuclear Receptor Genes

Background and Aim: The marked inter-individual variability in warfarin dose requirement is mainly attributed to the variants of the main target genes across the warfarin pharmacological pathway, i.e. cytochrome P-450 (CYP) 2C9, CYP3A4, ABCB1, vitamin K epoxide reductase complex subunit 1 (VKORC1) and epoxide hydrolase 1 (EPHX1). The expression of these genes is primarily regulated by a complex network of nuclear receptors (NR) which are encoded, in turn, by polymorphic genes. Furthermore, microRNAs (miRNA) can reduce the expression of both warfarin target genes and the genes encoding NRs. We investigated whether the variants of genes encoding i) warfarin targets, ii) NRs involved in the regulation of the targets and iii) miRNA involved in the regulation of NRs and the targets, are associated with warfarin dose requirements. Methods and Results: The study was performed using the Quebec Warfarin Cohort consisting of 988 new warfarin users who were recruited over a period of 3 years and were followed-up for 12 months. The candidate genes were selected according to the literature and included 5 warfarin target genes, 9 NRs genes and 29 miRNA genes. SNPs with MAF>0.01 within these genes were selected for the present study from the NCBI database. Genotyping was previously performed using Omni 2.5 array (Illumina, CA) and iPLEX ADME CYP2C9/VKORC1 panel (Agena, CA) and missing SNPs were imputed. We tested the effect of SNPs on the log-transformed values of dose at 3-month post-initiation using multivariate linear regression analysis, and the adjusted significance threshold was set to 2.49 x 10-4. A total of 3970 SNPs, including 702 in target genes, 3162 in NRs and 106 in miRNA genes were studied. For the association with warfarin dose, 137 SNPs in CYP2C9 and VKORC1 were significant, as well as 3 SNPs in nuclear vitamin D receptor (VDR) gene: rs4760658, β = -0,0852, P = 7,32x10-5; rs11168293, β = -0,0809, P = 1,78x10-4; rs4760655, β = -0,0799, P = 2,15x10-4. None of the variants in miRNA genes showed a significant association with warfarin dose. Conclusions: Variants in the gene encoding VDR, a nuclear receptor which has been previously shown to be involved in the regulation of CYP2C9, CYP3A4 and ABCB1, may contribute to the variability in response to warfarin.

A pharmacogenetics-based warfarin maintenance dosing algorithm from Northern Chinese patients.

Inconsistent associations with warfarin dose were observed in genetic variants except VKORC1 haplotype and CYP2C9*3 in Chinese people, and few studies on warfarin dose algorithm was performed in a large Chinese Han population lived in Northern China. Of 787 consenting patients with heart-valve replacements who were receiving long-term warfarin maintenance therapy, 20 related Single nucleotide polymorphisms were genotyped. Only VKORC1 and CYP2C9 SNPs were observed to be significantly associated with warfarin dose. In the derivation cohort (n = 551), warfarin dose variability was influenced, in decreasing order, by VKORC1 rs7294 (27.3%), CYP2C9*3(7.0%), body surface area(4.2%), age(2.7%), target INR(1.4%), CYP4F2 rs2108622 (0.7%), amiodarone use(0.6%), diabetes mellitus(0.6%), and digoxin use(0.5%), which account for 45.1% of the warfarin dose variability. In the validation cohort (n = 236), the actual maintenance dose was significantly correlated with predicted dose (r = 0.609, P<0.001). Our algorithm could improve the personalized management of warfarin use in Northern Chinese patients.

Polymorphism Detection of VKORC1 and CYP2C9 Genes for Warfarin Dose Adjustment by Real-Time PCR

Results: The allelic frequencies for VKORC1 C1173T (CC, CT, TT) were 25%, 51% and 24%, respectively and for VKORC1 G1639A (-GG, GA, AA) were 27%, 51% and 22%, respectively. Despite a significant association between allelic polymorphisms in VKORC1 and warfarin dose (P < 0.001), it was not statistically significant for CYP2C9*2 and CYP2C9*3, most probably due to the low rate of CYP2C9*2 and CYP2C9*3 SNPs observed. Conclusions: SimpleProbe® real-time PCR is a fast and accurate technique qualified to detect VKORC1 and CYP2C9 SNPs. These results encourage taking further steps towards using VKORC1 and CYP2C9 allelic screening to prevent clinical complications due to resistance or sensitivity to warfarin as well as anti-coagulant dose adjustment.

Research Highlights: Novel CYP2C9 Genetic Polymorphisms and Assessment of their Impact On Hydroxylation Capacity

ISSN 1462-2416 10.2217/PGS.13.243 © 2014 Future Medicine Ltd Pharmacogenomics (2014) 15(3), 261–264 261 part of CYP2C9 is a CYP450 that constitutes approximately 20% of the total human liver microsome CYP450 protein con­ tent. CYP2C9 metabolizes approximately 10% of therapeutically important drugs such as oral sulfonylurea hypoglycemics (e.g., glimepiride, gliclazide and tolbuta­ mide), anticoagulants (e.g., warfarin and acenocoumarol), angiotensin II receptor inhibitors (e.g., losartan, irbesartan and candesartan), antiepileptics (e.g., pheny­ toin and phenobarbital) and numerous nonsteroidal anti­inflammatory drugs such as diclofenac [1,2]. The CYP2C9 gene, together with another three CYP2C genes (CYP2C8, CYP2C18 and CYP2C19), is located in a region on chromosome 10q24 [3]. This gene is highly polymorphic, including allelic variants with decreased (e.g., CYP2C9*2 or*3) or null activity (CYP2C9*6 ) [101]. The genotypic features of the CYP2C9 gene have been studied in several popula­ tions showing ethnic variability in allele distribution [2–5]. In Caucasians, the CYP2C9*1 (wild­type), *2 and *3 allelic variants are the most frequent. In Ori­ ental subjects, CYP2C9*4 has also been found, whereas in African subjects other alleles (CYP2C9*5, *6 and *8) have been identified [2–5]. In addition, a large number of SNPs have been described in the regulatory regions of the CYP2C9 gene [101], which appears to be in linkage disequilibrium with known exonic variants [6]. The studies discussed here analyze the presence of novel CYP2C9 SNPs. It appears that these genetic variations are likely to contribute to differences in CYP2C9 activ­ ity on warfarin and acenocoumarol [6,7], losartan [8] and glimepiride [9].

Evaluation of the warfarin-resistance polymorphism, VKORC1 Asp36Tyr, and its effect on dosage algorithms in a genetically heterogeneous anticoagulant clinic

To assess allele frequency and potential predictive value of recurrent polymorphisms affecting warfarin metabolism in an unselected patient cohort attending an anticoagulant clinic (n=186). Genotyping of ten SNPs in five candidate genes (VKORC1, CYP2C9, CALU, EPHX and GGCX) was carried out by ABI PRISM SNaPshot multiplex method. We confirm the association between high-frequency SNPs, VKORC1 c.-1639G>A and CYP2C9 *2/*3 and warfarin sensitivity, and contribute additional evidence that the VKORC1 p.Asp36Tyr variant is recurrent and independently associated with warfarin resistance in our population. Other SNPs made little contribution. Warfarin sensitivity was predicted by known VKORC1 and CYP2C9 SNPs. However, resistance associated with p.Asp36Tyr in VKORC1 would not be predicted by the usual markers. Despite its relatively low frequency (3/186 or 1.6%) clinical considerations may warrant its inclusion in pharmacogenetic screening for initial warfarin dosing in clinic populations with a heterogeneous population base.

Novel CYP2C9 and VKORC1 gene variants associated with warfarin dosage variability in the South African black population.

Warfarin is a widely used therapeutic agent for long-term oral anticoagulation worldwide. Its administration is challenging owing to its narrow therapeutic range and serious adverse effects. Several environmental factors and numerous genes, of which CYP2C9 and VKORC1 are the most important, have been associated with interindividual dosage variability. Many studies have been conducted to understand warfarin dosage variability better, the majority of which have been focused on the Caucasian and African-American populations. Very little information is available regarding genetic influences of warfarin dosage variability in the South African black population. In this study, we genotyped 213 South African black individuals for CYP2C9 and VKORC1 variants and a small subset of environmental factors that may be responsible for warfarin dosage variability. We observed 26 novel SNPs and seven previously described CYP2C9 variants and three previously described but no novel VKORC1 SNPs. Only 11 of the CYP2C9 variants and two of the VKORC1 variants were observed at high enough allele frequencies to assess their impact on warfarin dosage. We demonstrate that CYP2C9*8 and two novel CYP2C9 SNPs (g.16179 and g.46028) are associated with a decrease in warfarin dosage, β-blockers are independently associated with a decrease in warfarin dosage and two known VKORC1 variants (rs7200749 and rs7294) are associated with an increase in warfarin dosage. The CYP2C9 and VKORC1 variants and a small subset of environmental factors used in the study explain approximately 45% of warfarin dosage variability in the South African black population.

Allelic Variants in the CYP2C9 and VKORC1 Loci and Interindividual Variability in the Anticoagulant Dose Effect Of Warfarin in Italians

Warfarin is currently considered to be the anticoagulant of choice in the long-term treatment and prevention of thromboembolic events. However, it presents a narrow therapeutic range and a great interindividual dose variability. We investigated the influence of variants of the VKORC1 and CYP2C9 loci on the mean weekly warfarin dose (MWWD) required to reach stabilized therapeutic international normalized ratio, in order to confirm and to estimate the contribution of common genetic variability of these two genes in an Italian population and to search for novel rare VKORC1 alleles. A total of 148 patients were followed for 6 months and analyzed for VKORC1 and CYP2C9 gene variants. Analysis of variance and multiple linear regression analysis were used to study the contribution of each genetic factor to MWWD requirement. The complete sequencing of the VKORC1 coding region did not reveal the presence of exonic variants, while two common noncoding SNPs were highly associated: the T allele of VKORC1 1173C>T SNP (tag-SNP of H1-H2 haplotypes) is highly associated with low MWWD (p < 0.0001), while the A allele of VKORC1 3730G>A SNP (tag-SNP of H9 haplotype) is associated with high MWWD (p = 0.001). Also, CYP2C9*2 (Arg144Cys) and CYP2C9*3 (Ile359Leu) variant alleles were significantly associated with low MWWD (p = 0.003 and 0.027, respectively). According to a multiple linear regression model including, besides VKORC1 and CYP2C9 SNPs, also age and weight, this percentage reaches 56% (gender is not significant). Our results clearly indicate VKORC1 as the gene with the largest contribution to MWWD. Analyzing only one tag SNP of VKORC1 gene (1173C>T), it is possible to foresee 20% of the total variability. Our results may contribute to give useful indications for clinicians especially in the initiation of therapy so as to avoid the risk of adverse events.