Thirteen novel VKORC1 mutations associated with oral anticoagulant resistance: insights into improved patient diagnosis and treatment

Abstract

Summary. Background: Vitamin K 2,3-epoxide reductase complex subunit 1 (VKORC1) is the molecular target of oral anticoagulants. Mutations in VKORC1 cause partial or total coumarin resistance. Objectives: To identify new VKORC1 oral anticoagulant (OAC) resistance (OACR) mutations and compare the severity of patient phenotypes across different mutations and prescribed OAC drugs. Patients/Methods: Six hundred and twenty-six individuals exhibiting partial or complete coumarin resistance were analyzed by VKORC1 gene sequencing and CYP2C9 haplotyping. Results: We identified 13 patients, each with a different, novel human VKORC1 heterozygous mutation associated with an OACR phenotype. These mutations result in amino acid substitutions: Ala26 fi Thr, His28 fi Gln, Asp36 fi Gly, Ser52 fi Trp, Ser56 fi Phe, Trp59 fi Leu, Trp59 fi Cys, Val66 fi Gly, Gly71 fi Ala, Asn77 fi Ser, Asn77 fi Tyr, Ile123 fi Asn, and Tyr139 fi His. Ten additional patients each had one of three previously reported VKORC1 mutations (Val29 fi Leu, Asp36 fi Tyr, and Val66 fi Met). Genotyping of frequent VKORC1 and CYP2C9 polymorphisms in these patients revealed a predominant association with combined non-VKORC1*2 and wild-type CYP2C9 haplotypes. Additionally, data for OAC dosage and the associated measured International Normalized Ratio (INR) demonstrate that OAC therapy is often discontinued by physicians, although stable therapeutic INR levels may be reached at higher OAC dosages. Bioinformatic analysis of VKORC1 homologous protein sequences indicated that most mutations cluster into protein sequence segments predicted to be localized in the lumenal loop or at the endoplasmic reticulum membrane-lumen interface. Conclusions: OACR mutations of VKORC1 predispose afflicted patients to high OAC dosage requirements, for which stable, therapeutic INRs can sometimes be attained.

Adaptative evolution of the Vkorc1 gene in Mus musculus domesticus is influenced by the selective pressure of anticoagulant rodenticides

“…This variability being partly due to genetic polymorphisms in the Vkorc1 gene, numerous studies have been performed to detect new Vkorc1 mutations in patients resistant to AVK. These studies allowed to detect in humans 27 mutations in the coding sequence of the Vkorc1 gene (Bodin, Horellou, Flaujac, Loriot, & Samama, 2005; Bodin, Perdu, Diry, Horellou, & Loriot, 2008; D'Andrea et al., 2005; Harrington, Siddiq, Allford, Shearer, & Mumford, 2011; Harrington et al., 2008; Loebstein et al., 2007; Osman, Enström, Arbring, Söderkvist, & Lindahl, 2006; Peoc'h, Pruvot, Gourmel, Dit Sollier, & Drouet, 2009; Rieder et al., 2005; Rishavy, Usubalieva, Hallgren, & Berkner, 2011; Rost et al., 2004; Schmeits et al., 2010; Watzka et al., 2011; Wilms, Touw, Conemans, Veldkamp, & Hermans, 2008). In rats, 15 missense mutations in the Vkorc1 gene of Rattus norvegicus have been described in Europe (Grandemange, Lasseur, Longin‐Sauvageon, Benoit, & Berny, 2010; Haniza et al., 2015; Pelz et al., 2005, 2012; Rost et al., 2004).…”

The Arg98Trp mutation in human VKORC1 causing VKCFD2 disrupts a di-arginine–based ER retention motif

“…10,11 Currently, the membrane topology of hVKORC1 is not fully resolved, and 3TM or 4TM a-helices have been proposed for hVKORC1. 10,[21][22][23][24][25] However, for both topology models, the putative di-arginine ER retention motif Arg98_Arg100 is predicted to be exposed to the cytoplasmic face of the ER membrane. In the 3TM model, this di-arginine motif is part of the end of the big cytoplasmatic loop close to the second transmembrane a-helix.…”

Genetic resistance to warfarin therapy masked by amiodarone in a 2-year-old girl with mitral valve replacement

“…To investigate this partial pharmacodynamic resistance to warfarin [3], we analyzed the gene encoding vitamin K epoxide reductase complex subunit 1 (VKORC1), which is the pharmacologic target of vitamin K antagonists (VKAs). Exonic regions, intron-exon boundaries and fragments covering 2000 bp of the 5′-flanking VKORC1 region were sequenced as previously described [4,5].…”

“…These excessive VKA dose requirements cannot be explained entirely by non-genetic factors and common VKORC1/CYP2C9 SNPs. Patients requiring such excessive doses should benefit from detailed VKORC1 genetic analysis [3]; rare VKORC1 mutations associated with VKA resistance have been reported in adults, but not in pediatric patients. According to our previously published dose prediction algorithm incorporating height, target INR, and classic VKORC1/ CYP2C9 genotypes [2], the expected dose for our 2.2-year-old patient was calculated to be 25 mg weekly, i.e.…”

“…We identified three VKORC1 mutations: c.106G>T, leading to p.Asp36Tyr; and, in the 5′-flanking region, g.À1185G>A and g.À679A>G. The p.Asp36Tyr mutation has been reported in adults as a factor predisposing to warfarin resistance and a marker for high dose requirements that overrides the dose-reducing effect of CYP2C9*2, CYP2C9*3 and VKORC1 À1639A alleles [3,4,7]. Adults heterozygous for the p.Asp36Tyr mutation have been reported to require a mean weekly warfarin dose of~70 mg in the largest published cohort, i.e.…”