Minor allele of the factor V K858R variant protects from venous thrombosis only in non-carriers of factor V Leiden mutation.

Factor V serves an important role in the regulation of blood coagulation. The rs6025 (R534Q) and rs4524 (K858R) polymorphisms in the F5 gene, are known to influence the risk of venous thrombosis. While the rare Q534 (factor V Leiden) allele is associated with an increased risk of venous thrombosis, the minor R858 allele is associated with a lower risk of disease. However, no study has deeply examined the cumulative impact of these two variations on venous thrombosis risk. We study the association of these polymorphisms with the risk of venous thrombosis in 4 French case-control populations comprising 3719 patients and 4086 controls. We demonstrate that the Q534 allele has a dominant effect over R858. Besides, we show that in individuals not carrying the Q534 allele, the protective effect of the R858 allele acts in a dominant mode. Thrombin generation-based normalized activated protein C sensitivity ratio was lower in the 858R/R homozygotes than in the 858K/K homozygotes (1.92 ± 1.61 vs 2.81 ± 1.57, p = 0.025). We demonstrate that the R858 allele of the F5 rs4524 variant protects from venous thrombosis only in non-carriers of the Q534 allele of the F5 rs6025. Its protective effect is mediated by reduced factor VIII levels and reduced activated protein C resistance.

Protein S Heerlen mutation heterozygosity is associated with venous thrombosis risk.

Hereditary Protein S (PS) deficiency is a rare coagulation disorder associated with an increased risk of venous thrombosis (VT). The PS Heerlen (PSH) mutation is a rare S501P mutation that was initially considered to be a neutral polymorphism. However, it has been later shown that PSH has a reduced half-life in vivo which may explain the association of PSH heterozygosity with mildly reduced levels of plasma free PS (FPS). Whether the risk of VT is increased in PSH carriers remains unknown. We analyzed the association of PSH (rs121918472 A/G) with VT in 4,173 VT patients and 5,970 healthy individuals from four independent case-control studies. Quantitative determination of FPS levels was performed in a subsample of 1257 VT patients. In the investigated populations, the AG genotype was associated with an increased VT risk of 6.57 [4.06-10.64] (p = 1.73 10-14). In VT patients in whom PS deficiency was excluded, plasma FPS levels were significantly lower in individuals with PSH when compared to those without [72 + 13 vs 91 + 21 UI/dL; p = 1.86 10-6, mean + SD for PSH carriers (n = 21) or controls (n = 1236) respectively]. We provide strong evidence that the rare PSH variant is associated with VT in unselected individuals.

Homocysteine concentration in coronary artery disease: Influence of three common single nucleotide polymorphisms.

BACKGROUND AND AIMS: Whether single nucleotide polymorphisms (SNPs) of homocysteine metabolism enzymes influence the rate of cardiovascular (CV) events in coronary artery disease (CAD) patients remains controversial. METHODS AND RESULTS: In this analysis, 1126 subjects from the AtheroGene study with CAD and 332 control subjects without known CAD were included. The following SNPs were investigated: methylentetrahydrofolate reductase (MTHFR-C667T), methionin synthetase (MS-D919G), and cystathionin beta synthetase (CBS-I278T). The endpoint was the combination of cardiovascular death, stroke, and non-fatal myocardial infarction (N = 286). The median follow-up time was 6.4 years. Kaplan-Meier curve analysis showed an increasing event rate with rising homocysteine levels (p < 0.001) in CAD patients. Further, in Cox-Regression analysis homocysteine was a predictor of the endpoint with a hazard ratio (HR) of 6.5 (95% CI: 2.9-14.6, p < 0.001) in the adjusted model including cardiovascular risk factors. Of the three SNPs, homozygous MTHFR SNP increased homocysteine levels significantly in patients with CAD and individuals without CAD (both p < 0.001). The SNPs in MS and CBS were not related to relevant changes in homocysteine levels in CAD patients or controls. The different SNPs of MTHFR, MS, and CBS were not related to an increased event rate. CONCLUSION: Homocysteine level is a strong predictor of CV events. Subjects with and without CAD and SNPs in the enzyme MTHFR had increased homocysteine levels. This was not observed for MS and CBS SNPs. Although MTHFR SNPs alter homocysteine levels in patients and controls, these polymorphisms had no impact on prognosis in CAD patients.

Genetic risk factors for venous thrombosis in women using combined oral contraceptives: update of the PILGRIM study.

Identifying women at risk of venous thrombosis (VT) under combined oral contraceptives (COC) is a major public health issue. The aim of this study was to investigate in COC users the impact on disease of genetic polymorphisms recently identified to associate with VT risk in the general population. Nine polymorphisms located on KNG1, F11, F5, F2, PROCR, FGG, TSPAN and SLC44A2 genes were genotyped in a sample of 766 patients and 464 controls as part of the PILGRIM (PILl Genetic Risk Monitoring) study. Cases were women who experienced an episode of documented VT during COC use, while controls were women with no history of VT using COC at the time of inclusion. Among the studied polymorphisms, only F11 rs2289252 was significantly associated with VT. The F11 rs2289252-A allele was associated with a 1.6-fold increased risk of VT (p < 0.0001). Besides, the combination of the rs2289252-A allele with non-O blood group, present in 52% of the cohort, was associated with an odds ratio of 4.00 (2.49-6.47; p < 10-4 ). The consideration of this genetic risk factor could help to better assess the risk of VT in COC users.

Single nucleotide polymorphisms in an intergenic chromosome 2q region associated with tissue factor pathway inhibitor plasma levels and venous thromboembolism.

Essentials Tissue factor pathway inhibitor (TFPI) regulates the blood coagulation cascade. We replicated previously reported linkage of TFPI plasma levels to the chromosome 2q region. The putative causal locus, rs62187992, was associated with TFPI plasma levels and thrombosis. rs62187992 was marginally associated with TFPI expression in human aortic endothelial cells. Click to hear Ann Gil's presentation on new insights into thrombin activatable fibrinolysis inhibitor SUMMARY: Background Tissue factor pathway inhibitor (TFPI) regulates fibrin clot formation, and low TFPI plasma levels increase the risk of arterial thromboembolism and venous thromboembolism (VTE). TFPI plasma levels are also heritable, and a previous linkage scan implicated the chromosome 2q region, but no specific genes. Objectives To replicate the finding of the linkage region in an independent sample, and to identify the causal locus. Methods We first performed a linkage analysis of microsatellite markers and TFPI plasma levels in 251 individuals from the F5L Family Study, and replicated the finding of the linkage peak on chromosome 2q (LOD = 3.06). We next defined a follow-up region that included 112 603 single nucleotide polymorphisms (SNPs) under the linkage peak, and meta-analyzed associations between these SNPs and TFPI plasma levels across the F5L Family Study and the Marseille Thrombosis Association (MARTHA) Study, a study of 1033 unrelated VTE patients. SNPs with false discovery rate q-values of < 0.10 were tested for association with TFPI plasma levels in 892 patients with coronary artery disease in the AtheroGene Study. Results and Conclusions One SNP, rs62187992, was associated with TFPI plasma levels in all three samples (ß = + 0.14 and P = 4.23 × 10-6 combined; ß = + 0.16 and P = 0.02 in the F5L Family Study; ß = + 0.13 and P = 6.3 × 10-4 in the MARTHA Study; ß = + 0.17 and P = 0.03 in the AtheroGene Study), and contributed to the linkage peak in the F5L Family Study. rs62187992 was also associated with clinical VTE (odds ratio 0.90, P = 0.03) in the INVENT Consortium of > 7000 cases and their controls, and was marginally associated with TFPI expression (ß = + 0.19, P = 0.08) in human aortic endothelial cells, a primary site of TFPI synthesis. The biological mechanisms underlying these associations remain to be elucidated.

Is there still room for additional common susceptibility alleles for venous thromboembolism?

UNLABELLED: Essentials Genetic architecture of venous thromboembolism (VTE) remains to be fully disentangled. 11 newly discovered candidate polymorphisms were genotyped in 3019 VTE cases and 2605 controls. None of the 11 polymorphisms were significantly associated with VTE risk. Additional major efforts are needed to identify VTE-associated genetic variants. SUMMARY: Background Through a meta-analysis of 12 genome-wide association studies, the International Network against VENous Thrombosis (INVENT) consortium identified two novel susceptibility loci for venous thromboembolism (VTE). This project has also generated other candidates that need to be confirmed. Objectives To assess the association with VTE of common single-nucleotide polymorphisms (SNPs) that demonstrated strong statistical, but not genome-wide, significance in the INVENT cohorts. Patients/methods Eleven SNPs were genotyped and tested for association with VTE in three case-control studies totaling 3019 patients and 2605 healthy individuals. Results and conclusions None of the tested SNPs showed evidence for association with VTE. Different strategies are needed to decipher the whole spectrum of common and rare genetic variations associated with VTE risk.

Genetic determinants of tissue factor pathway inhibitor plasma levels.

Tissue factor pathway inhibitor (TFPI) impedes early stages of the blood coagulation response, and low TFPI plasma levels increase the risk of thrombosis. TFPI plasma levels are heritable, but specific genetic determinants are unclear. We conducted a comprehensive review of genetic risk factors for TFPI plasma levels and identified 26 studies. We included 16 studies, as well as results from two unpublished genome-wide studies, in random effects meta-analyses of four commonly reported genetic variants in TFPI and its promoter (rs5940, rs7586970/rs8176592, rs10931292, and rs10153820) and 10 studies were summarised narratively. rs5940 was associated with all measures of TFPI (free, total, and activity), and rs7586970 was associated with total TFPI. Neither rs10931292 nor rs10153820 showed evidence of association. The narrative summary included 6 genes and genetic variants (P151L mutation in TFPI, PROS1, F5, APOE, GLA, and V617F mutation in JAK2) as well as a genome-wide linkage study, and suggested future research directions. A limitation of the systematic review was the heterogeneous measurement of TFPI. Nonetheless, our review found robust evidence that rs5940 and rs7586970 moderate TFPI plasma levels and are candidate risk factors for thrombosis, and that the regulation of TFPI plasma levels involves genetic factors beyond the TFPI gene.

Risk assessment of venous thrombosis in families with known hereditary thrombophilia: the MARseilles-NImes prediction model.

BACKGROUND: Although predicting the risk of venous thrombosis (VT) in an individual from a family with inherited thrombophilia is of major importance, it is often not feasible. OBJECTIVES: To develop a simple risk assessment model that improves prediction of the risk of VT for individuals of families with inherited thrombophilia. PATIENTS/METHODS: 1201 relatives from 430 families with inherited thrombophilia (deficiencies of antithrombin, protein C or protein S, and the factor V Leiden and F2 20210A mutations) were recruited at the referral center for thrombophilia in Marseilles, France, from 1986 to 2008. One hundred and twenty-two individuals had a personal history of VT. Sixteen preselected clinical and laboratory variables were used to derive the VT risk score. RESULTS: The scores based on the 16 variables and on the five most strongly associated variables performed similarly (areas under receiver operating characteristic curves of 0.85 and 0.83, respectively). For the five-variable score, named the MARNI score, derived from family history score of VT, von Willebrand factor antigen levels, age, severity of thrombophilia, and FGG rs2066865, the risk of VT ranged from 0.2% for individuals with a score of 0 (n = 186) to > 70% for individuals with a score of ≥ 7 (n = 27). The model was validated with an internal bootstrap method. CONCLUSIONS: With the use of a simple scoring system, assessment of the risk of VT in subjects from families with inherited thrombophilia can be greatly improved. External validation is now needed to replicate these findings.

Risk assessment of venous thrombosis in families with known hereditary thrombophilia: the MARseilles-NImes prediction model.

BACKGROUND: Although predicting the risk of venous thrombosis (VT) in an individual from a family with inherited thrombophilia is of major importance, it is often not feasible. OBJECTIVES: To develop a simple risk assessment model that improves prediction of the risk of VT for individuals of families with inherited thrombophilia. PATIENTS/METHODS: 1201 relatives from 430 families with inherited thrombophilia (deficiencies of antithrombin, protein C or protein S, and the factor V Leiden and F2 20210A mutations) were recruited at the referral center for thrombophilia in Marseilles, France, from 1986 to 2008. One hundred and twenty-two individuals had a personal history of VT. Sixteen preselected clinical and laboratory variables were used to derive the VT risk score. RESULTS: The scores based on the 16 variables and on the five most strongly associated variables performed similarly (areas under receiver operating characteristic curves of 0.85 and 0.83, respectively). For the five-variable score, named the MARNI score, derived from family history score of VT, von Willebrand factor antigen levels, age, severity of thrombophilia, and FGG rs2066865, the risk of VT ranged from 0.2% for individuals with a score of 0 (n = 186) to > 70% for individuals with a score of ≥ 7 (n = 27). The model was validated with an internal bootstrap method. CONCLUSIONS: With the use of a simple scoring system, assessment of the risk of VT in subjects from families with inherited thrombophilia can be greatly improved. External validation is now needed to replicate these findings.

Familial aggregation of metabolic syndrome indicators in Portuguese families.

BACKGROUND AND AIMS: Family studies are well suited to investigate the genetic architecture underlying the metabolic syndrome (MetS). The purposes of this paper were (i) to estimate heritabilities for each of the MetS indicators, and (ii) to test the significance of familial intratrait and cross-trait correlations in MetS markers. METHODS AND RESULTS: This study included 1,363 individuals from 515 Portuguese families in which five MetS components, including waist circumference (WC), blood pressure (BP), HDL-cholesterol, triglycerides (TG), and glucose (GLU), were measured. Intratrait and cross-trait familial correlations of these five components were estimated using Generalized Estimating Equations. Each MetS component was significantly heritable (h (2) ranged from 0.12 to 0.60) and exhibited strong familial resemblance with correlations between biological relatives of similar magnitude to those observed between spouses. With respect to cross-trait correlations, familial resemblance was very weak except for the HDL-TG pair. CONCLUSIONS: The present findings confirm the idea of familial aggregation in MetS traits. Spousal correlations were, in general, of the same magnitude as the biological relatives' correlations suggesting that most of the phenotypic variance in MetS traits could be explained by shared environment.

Current knowledge on the genetics of incident venous thrombosis.

The genetic burden underlying venous thrombosis (VT) is characterized by a sibling relative risk of 2.5 and a strong heritability whose estimates varied from 35% to 60% according to different studies. However, the genetic factors identified so far only explain about 5% of VT heritability and just 16 genes have been robustly associated with the susceptibility to VT, most of them affecting the coagulation cascade. Eight of these have been identified during the last 5 years, thanks to the development of high-throughput micro-array genotyping technologies, which have radically changed the research landscape in human genetics. The present work is aimed at providing a historical review of the known genetic factors contributing to VT risk, as well as discussing future research strategies to follow to disentangle the whole spectrum of genetic variants associated with VT.

PARVG gene polymorphism and operational renal allograft tolerance.

A unique blood transcriptional profile of 49 genes has been previously highlighted that may be used to distinguish drug-free operationally tolerant kidney recipients (TOL) from other kidney recipients with contrasted clinical situations and healthy volunteers. The aim of this study was to investigate whether the pattern of these 49 genes could be influenced by genetic polymorphisms located in the corresponding genomic sequences and whether some of these single nucleotide polymorphisms (SNPs) could be associated with clinical status of kidney transplant recipients. In this study, 1152 candidate tag SNPs spanning these genes were genotyped using a Golden Gate Illlumina assay in a sample of 164 kidney transplant recipients, including 11 operationally tolerant patients, 134 patients with stable graft function, 19 with proven signs of chronic rejection, and 27 healthy volunteers. The gene expression and clinical status were studied according to the different SNPs. Among the genes demonstrating expression difference between TOL compared with CR&STA patients, PARVG, which is a member of a family of actin-binding proteins associated with focal contacts, stands out with 2 SNPs, (rs139144 and rs5764592) explaining about 20% of the gene expression variability. Linkage disequilibrium analysis of these 2 SNPs showed the rs139144GG genotype was associated with decreased PARVG expression and was numerically more frequent in TOL (60%) than in CR&STA (28%) patients (P = .068). These preliminary results, which should be confirmed in a larger population, open new perspective of regulation pathways and hypothesis in operational tolerance mechanism.

Lessons from genome-wide association studies in venous thrombosis.

From the first genome wide association studies (GWAS) conducted on age-related macular degeneration back in 2005 until now, hundreds of studies have applied this strategy to identify novel genetic loci associated with hundreds of human diseases and related quantitative risk factors. While the GWAS revolution has just started to shift towards the next generation sequencing's burst, it is important to illustrate how the genetics research in venous thrombosis has benefit from the GWAS paradigm.

A multi-stage multi-design strategy provides strong evidence that the BAI3 locus is associated with early-onset venous thromboembolism.

BACKGROUND: Factor VIII (FVIII) and von Willebrand factor (VWF) are two known quantitative risk factors for venous thromboembolism (VTE). OBJECTIVES: To identify new loci that could contribute to VTE susceptibility and to modulating FVIII and/or VWF levels. PATIENTS/METHODS: A pedigree linkage analysis was first performed in five extended French-Canadian families, including 253 individuals, to identify genomic regions linked to FVIII or VWF levels. Identified regions were further explored using 'in silico' genome-wide association studies (GWAS) data on VTE (419 patients and 1228 controls), and two independent case-control studies (MARTHA and FARIVE) for VTE, gathering 1166 early-onset patients and 1408 healthy individuals. Single nucleotide polymorphisms (SNPs) associated with VTE risk were further investigated in relation to plasma levels of FVIII and VWF in a cohort of 108 healthy nuclear families. RESULTS: Four main linkage regions were identified, among which the well-characterized ABO locus, the recently identified STAB 2 gene, and a third one, on chromosome 6q13-14, harbouring four non-redundant SNPs, associated with VTE at P < 10(-4) in the GWAS dataset. The association of one of these SNPs, rs9363864, with VTE was further replicated in the MARTHA and FARIVE studies. The rs9363864-AA genotype was associated with a lower risk for VTE (OR = 0.58 [0.42-0.80], P = 0.0005) but mainly in non-carriers of the FV Leiden mutation. This genotype was further found to be associated with the lowest levels of FVIII (P = 0.006) and VWF (P = 0.001). CONCLUSIONS: The BAI3 locus where the rs9363864 maps is a new candidate for VTE risk.

Activated thrombin activatable fibrinolysis inhibitor levels are associated with the risk of cardiovascular death in patients with coronary artery disease: the AtheroGene study.

BACKGROUND: Thrombin activatable fibrinolysis inhibitor (TAFI) attenuates fibrinolysis. Results on the association between TAFI levels and the risk of coronary artery disease (CAD) are inconsistent. OBJECTIVES: We investigated the association between TAFI levels and the risk of cardiovascular events in CAD. PATIENTS/METHODS: 1668 individuals with angiographically proven CAD at baseline were followed for a median of 2.3 years, as part of the prospective AtheroGene cohort. Fifty-six deaths from cardiovascular (CV) causes and 35 non-fatal CV events were observed. RESULTS: At baseline, three TAFI measurements were available: one evaluating the total amount of TAFI (t-TAFI), one measuring the TAFIa/TAFIai amount, and the last the released activated peptide (TAFI-AP). TAFIa/TAFIai levels were associated with increased risk of CV death [hazard ratio (HR) for one tertile increase, 2.38 (1.56-3.63); P < 10(-4)]. This association remained significant after adjustment for conventional risk factors, CRP levels, white blood count and markers of thrombin generation and fibrinolysis [HR = 1.69 (1.07-2.67); P = 0.01]. In addition, CPB2 gene polymorphisms explained 12%, 6%, and 3% of t-TAFI, TAFIa/TAFIai and TAFI-AP levels, respectively, but none was associated with CV events. CONCLUSIONS: The amount of activated TAFI, measured by TAFIa/TAFIai ELISA, but not of the t-TAFI is independently associated with the risk of CV death.

Polymorphisms of the tumor necrosis factor-alpha (TNF) and the TNF-alpha converting enzyme (TACE/ADAM17) genes in relation to cardiovascular mortality: the AtheroGene study.

Tumor necrosis factor (TNF) is a major cytokine involved in inflammatory reaction and a mortality predictor in patients with coronary artery disease (CAD). Plasma levels of soluble TNF (sTNF) depend on the rate of its synthesis but also on its shedding from cell surface, a mechanism mainly regulated by the TNF alpha converting enzyme (TACE or ADAM17). We investigated the relationship between ADAM17 and TNF polymorphisms, circulating levels of shed ADAM17 substrates (sTNF, sTNFR1 and sTNFR2), and cardiovascular risk in a prospective cohort of CAD patients. Five tag single-nucleotide polymorphisms (SNPs) of the ADAM17 gene as well as four previously described TNF SNPs were genotyped in the Atherogene Study composed of 1,400 CAD patients among which 136 died from a cardiovascular (CV) cause. sTNF, sTNFR1, and sTNFR2 concentrations were all significantly elevated in patients with future CV death, independently of other clinical/biological variables. While none of the studied TNF SNPs was associated with sTNF, sTNFR1, nor sTNFR2 levels, the ADAM17 -154A allele was found associated with a 14% increase of sTNF levels as compared to the -154C allele (p = 0.0066). Moreover, individuals carrying the 747Leu allele displayed a borderline increased risk of future cardiovascular death [odds ratio, 2.06 (1.05-4.04), p = 0.03]. These results suggest a role of ADAM17 in the regulation of sTNF plasma levels and identifies ADAM17 gene as a candidate for CAD. Tumor necrosis factor (TNF) is a major cytokine involved in inflammatory reaction and a mortality predictor in patients with coronary artery disease (CAD). We have studied the association of ADAM17 and TNF polymorphisms with circulating levels of shed ADAM17 substrates (sTNF, sTNFR1 and sTNFR2) and with cardiovascular risk in a large population of individuals with CAD (Atherogene Study, n = 1,400). Two newly identified polymorphisms, obtained by a systematic sequencing of the ADAM17 gene, C-154A and Ser747leu, slightly influence respectively sTNF plasma levels and the risk of cardiovascular death.

Dissection of familial correlations in hepatitis C virus (HCV) seroprevalence suggests intrafamilial viral transmission and genetic predisposition to infection.

OBJECTIVE: Unsafe injections and transfusions used during treatments are considered to be responsible for many cases of transmission of hepatitis C virus (HCV) in developing countries, but cannot account for a substantial proportion of present infections. The aim of the present work was to investigate familial clustering of HCV infection in a population living in a highly endemic area. DESIGN, SETTING AND PARTICIPANTS: A large seroepidemiological survey was conducted on 3994 subjects (age range, 2-88 years) from 475 familial clusters in an Egyptian rural area. Epidemiological methods appropriate for the analysis of correlated data were used to estimate risk factors and familial dependences for HCV infection. A phylogenetic analysis was conducted to investigate HCV strain similarities within and among families. MAIN OUTCOME MEASURES: HCV familial correlations adjusted for known risk factors, similarities between viral strains. RESULTS: Overall HCV seroprevalence was 12.3%, increasing with age. After adjustment for relevant risk factors, highly significant intrafamilial resemblances in HCV seroprevalence were obtained between father-offspring (odds ratio (OR) = 3.4 (95% confidence interval (CI), 1.8 to 6.2)), mother-offspring (OR = 3.8 (95% CI, 2.5 to 5.8)), and sibling-sibling (OR = 9.3 (95% CI, 4.9 to 17.6)), while a weaker dependence between spouses (OR = 2.2 (95% CI, 1.3 to 3.7)) was observed. Phylogenetic analysis showed greater HCV strain similarity between family members than between unrelated subjects, indicating that correlations can be explained, in part, by familial sources of virus transmission. In addition, refined dissection of correlations between first-degree relatives supported the role of host genes predisposing to HCV infection. CONCLUSIONS: Current HCV infection in endemic countries has a strong familial component explained, at least partly, by specific modes of intrafamilial viral transmission and by genetic predisposition to infection.