ALBERT

Beschreibung: Abstract 150 Background: While bivariate interactions between Factor V Leiden, prothrombin G20201A and hereditary deficiency of antithrombin, protein C and protein S compound VTE risk, whether other gene-gene interactions are associated with VTE is largely unknown. Objective: To test gene-gene interactions for an association with VTE. Methods: Cases (n=1004) were Mayo Clinic European-American patients of non-Hispanic ancestry with objectively-diagnosed VTE in the absence of active cancer, venous catheter or antiphospholipid antibodies. Controls (n=1066) were Mayo Clinic outpatients without VTE who were frequency-matched on case age, gender, race, MI/stroke status and state of residence. We selected candidate genes relevant to the anticoagulant, procoagulant, fibrinolytic and innate immunity pathways, focusing on platelet, monocyte, neutrophil and endothelial cell agonists, receptors, ligands, signal transduction and adhesion molecules, granule contents and effectors; plasma proteases and inhibitors; matrix metalloproteases; inflammatory cytokines and receptors; estrogen, progesterone and androgen receptors; co-regulators and enzymes related to estrogen metabolism; important enzymes for catechol, homocysteine, thromboxane A2 and prostacyclin biosynthesis and metabolism; and HMG CoA reductase. For these genes (n=780), we selected all non-synonymous coding single nucleotide polymorphisms (SNPs) with minor allele frequency ≥0.5%; the remaining SNPs were selected using an LD tagging algorithm (Carlson et al. AJHG 2004); 558 ancestry-informative markers were also included (total n=13,237 SNPs). Leukocyte genomic DNA was genotyped using a custom Illumina Infinium iSelect chemistry and platform, including appropriate controls. For these analyses, all pairwise SNP-SNP interactions for 12,313 SNPs were tested using logistic regression. In addition, we tested all Factor V Leiden -, prothrombin G20210A - and ABO non-O blood group - SNP interactions. Results: The mean ± SD case and control ages were 55.3 ± 16.4 and 56.5 ± 15.9 years, respectively, and 51% were female. Analysis of ancestry-informative markers gave no evidence of population stratification. Among almost 76 million pairwise interactions tested, 504, 44 and 8 SNP-SNP interactions reached p-values ≤ 1E-5, 1E-6, and 1E-7 respectively; none reached the Bonferroni statistical significance threshold (≤ 6.6E-10). The gene-gene pairwise interactions with p-values ≤ 1E-7 included: Leptin receptor - Estrogen receptor 1, α1 Antichymotrypsin - β2 glycoprotein 1, CRADD(caspace) - B1 Bradykinin receptor, ILF10 - Prolactin receptor and GFRA2(TGFβ neurotrophic factor receptor) - Bactericidal permeability-increasing protein. Gene-gene pairwise interactions with the lowest p-value for Factor V Leiden -, prothrombin G20210A - and ABO non-group O - were Protein kinase C beta (2.1E-5), von Willebrand Factor (7.6E-4) and Phospholipase C gamma-2 (6.7E-4), respectively. Conclusion: Pairwise gene-gene interaction analyses suggest potential associations between VTE and novel genes and gene pathways. These potential associations require confirmation in future replication studies. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Abstract 709 Background: A recent analysis of merged genome-wide and candidate gene genotypes in VTE cases and controls identified multiple tag SNPs that were strongly associated with VTE. Objective: To identify rare and/or novel functional variants by sequencing the implicated genes. Methods: Cases (n=1488) were Mayo Clinic European-American patients of non-Hispanic ancestry with objectively-diagnosed VTE in the absence of active cancer, venous catheter or antiphospholipid antibodies. Controls (n=1439) were Mayo Clinic outpatients without VTE who were frequency-matched on case age, gender, race, MI/stroke status and state of residence. For this analysis, we selected a subset of these cases and controls for sequencing to take advantage of the joint configuration of two ABO SNPs of primary interest, rs8176719 (ABO exon 6 deletion determining type O blood group) and rs2519093 (ABO intron 1 tag SNP), which were previously shown to be strongly associated with VTE (p=5.7E-12 and p=3.0E-16, respectively). We randomly sampled 82 cases and 14 controls within 3 of the 9 potential allele frequency cells (Figure). The rs8176719 alleles are -−/−- (double deletion is the common allele), –/G, and G/G (the rare allele). The rs2519093 alleles are GG (G is the common allele), AG, and AA (A is the rare allele). For each SNP, the genotypes are represented as 0, 1, or 2 copies of the minor allele. We represented the joint allelic configuration of the two SNPs with the number of copies of the rs8176719 given first as 0/0 (both with 0 copies of the minor allele), 0/1, 0/2 (0 copies of the rs8176719 SNP), 1/0 (1 copy of the rare rs8176719 SNP), 1/1/, and 1/2, and 2/0 (2 copies of the rare allele for the rs8176719 SNP and 0 copies of the rare allele of the rs2519093 SNP), 2/1, and 2/2. From the Figure one observes discrepancies between cases and controls at the 0/0, 1/1 and 2/2 combinations. We randomly sampled from these three combinations, taking one third of the case series. For each SNP, we had 28 cases with 0/0 copies of the rare allele, 27 cases with 1/1 copies of the rare allele; and 27 cases with the combination of 2/2 copies of the rare allele. We compared these 82 cases with 14 controls that do not have any of these combinations. Sixteen genes were selected for deep sequencing, including 5 genes harboring SNPs significantly associated with VTE (F5, SLC19A2, ABO, NME7, ATP1B1), 10 genes with SNPs marginally associated with VTE (C1orf114, KLKB1, SELP, F11, SCUBE1, PRKCB1, CD44, ITPR1, GFRA1, BLZF1), and CYP4V2 which reportedly confounds F11 and KLKB1. Agilent SureSelect probes were designed to capture and enrich the ∼2 Mb genomic regions of these 16 genes. Samples were multiplexed (12-plex) and sequenced using Illumina HiSeq 2000. The sequence reads were aligned to the human genome build 36 using Burrows-Wheeler Aligner, and the single nucleotide variants (SNVs) and small INDELs were called using SNVMix and GATK, respectively. For this analysis, novel ABO SNVs were tested for an association with VTE using age-, sex-adjusted logistic regression and Fisher's Exact Test. Results: 98% of the targeted regions were sequenced with 〉 20X coverage. On average, ∼2500 SNVs and ∼200 INDELs were detected in each sample. Fifteen novel SNVs in intron 6 and 3' of the ABO gene were associated with VTE (p

Beschreibung: Abstract 480 Background: While interaction between Factor V Leiden and other VTE risk exposures (i.e., oral contraceptives, hormone therapy, pregnancy, cancer, minor trauma) compound VTE risk, whether other gene-environment interactions are associated with VTE is largely unknown. Objective: To test gene-environment interactions for an association with VTE. Methods: Cases (n=1488) were Mayo Clinic European-American patients of non-Hispanic ancestry with objectively-diagnosed VTE in the absence of active cancer, venous catheter or antiphospholipid antibodies. Controls (n=1439) were Mayo Clinic outpatients without VTE who were frequency-matched on case age, gender, race, MI/stroke status and state of residence. We selected candidate genes relevant to the anticoagulant, procoagulant, fibrinolytic and innate immunity pathways, focusing on platelet, monocyte, neutrophil and endothelial cell agonists, receptors, ligands, signal transduction and adhesion molecules, granule contents and effectors; plasma proteases and inhibitors; matrix metalloproteases; inflammatory cytokines and receptors; estrogen, progesterone and androgen receptors; co-regulators and enzymes related to estrogen metabolism; important enzymes for catechol, homocysteine, thromboxane A2 and prostacyclin biosynthesis and metabolism; and HMG CoA reductase. For these genes (n=750), we selected all non-synonymous coding single nucleotide polymorphisms (SNPs) with minor allele frequency ≥0.5%; the remaining SNPs were selected using an LD tagging algorithm (Carlson et al. AJHG 2004); 479 ancestry-informative markers were also included (total n=13,241 SNPs). Leukocyte genomic DNA was genotyped using a custom Illumina Infinium iSelect platform, including appropriate controls. We tested all pairwise interactions between 12,483 SNPs (12296 autosomal, 187 chromosome X) that passed quality control and each of seven exposures (ever “stress”[defined as ever hospitalized, ever surgery, ever trauma, ever transfemoral procedure or ever leg paresis], ever surgery, ever neurosurgery, ever orthopedic surgery; and among women, ever pregnant, ≥3 pregnancies and ever oral contraceptives/hormone therapy), adjusted for age, gender, MI/stroke status and state of residence, using logistic regression. False discovery rates (q-value) were calculated to estimate the expected fraction of false positive associations due to multiple statistical tests. Results: The mean ± SD case and control ages were 55.3 ± 16.4 and 56.5 ± 15.9 years, respectively, and 51% were female. Analysis of ancestry-informative markers revealed no evidence of population stratification. Among the seven models, a significant interaction (q≤0.05) was found between ever surgery and two genes located on chromosome 1: USF1 (rs2516840, OR=2.1, p=2.0E-06; q=0.02), and F11R (rs790055 & rs790056; OR∼2.29, p∼4E-06; q=0.02); the two F11R SNPs are in complete linkage dysequilibrium (LD), and the USF1 SNP is in moderate LD with the F11R SNPs (r2∼0.6). USF1 encodes for upstream transcription factor 1 which regulates many genes involved in lipid and glucose homeostasis. F11R encodes for the platelet F11 receptor (JAM [junctional adhesion molecule], a cell adhesion molecule important for platelet adhesion to cytokine-stimulated endothelial cells. Conclusion: Interaction between surgery and SNPs within USF1 and F11R are associated with significantly increased risks for VTE. These potential associations require confirmation in future replication studies. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Abstract 1148 Background: Factor V Leiden (F5rs6025) has been suggested as a disease-susceptibility SNP for deep vein thrombosis (DVT) but not PE. Given the significantly reduced survival after PE, identifying individuals at risk for PE is important for targeting prophylaxis and improving survival. Objectives: To identify PE disease-susceptibility genes. Methods: We performed in silicoGWAS analyses of VTE cases using genotype data imputed to ∼2.5 million SNPs from adults with objectively-diagnosed PE ± DVT (n=745), and DVT only (n=758). We tested these SNPs for an association with PE ± DVT after adjusting for age, sex, stroke and/or myocardial infarction, and USA state of residence using unconditional logistic regression. Results: Of all SNPs, three were most strongly associated with PE ± DVT: one at BDH1 on chromosome (chr) 3q29 (rs1309873, OR=1.56, p=4.02E-07) and two at SLC39A10 on chr 2q32.3 (rs7578216, OR=2.55, p=4.57E-07; and rs4589778, OR=2.49, p=7.52E-07). Individually adjusting for F5 rs6025 (chr 1), F2 rs1799963 (prothrombin G20210A; chr 11) and ABO rs8176719 (ABO blood type O allele; chr 9) did not materially affect these associations. None of these three SNPs associated with PE were intragenic. BDH1 encodes for 3-hydroxybutyrate dehydrogenase 1, a key mitochondrial enzyme in fatty acid catabolism, and SLC39A10encodes for solute carrier family 39, member 10, an important protein for zinc transport. Conclusions: Chromosomes 3q29 (BDH1) and 2q32.2 (SLC39A10) may harbor PE disease-susceptibility gene regions (genes). These findings require independent confirmation. If confirmed, additional studies addressing the biologic mechanisms through which these genes and/or gene regions operate to increase PE risk also will be required. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: Background The incidence of VTE in African-Americans (AAs) is similar to or higher than in Americans of European ancestry. However, the carrier frequencies of inherited thrombophilias common in whites (i.e., Factor V Leiden, Prothrombin G20210A) are very low in AAs, suggesting that other inherited thrombophilias may be associated with VTE in AAs. Objective To identify potentially novel non-hypothesis-driven single nucleotide polymorphisms (SNPs) associated with VTE in AAs. Methods We used the resources of the eMERGE Network to perform a GWAS of VTE in AAs. The eMERGE Network (funded by NHGRI) is comprised of nine sites each with DNA biobanks linked to electronic health records (EHRs). Approximately 39,206 unique DNA samples have been genotyped using either Affymetrix or Illumina genome-wide SNP arrays. Led by the Coordinating Center and eMERGE genomics workgroup, an imputation pipeline was developed for merging genomic data across the different SNP arrays used by the eMERGE sites, to maximize sample size and the power to detect associations. The imputation was performed using the 1000 Genomes Cosmopolitan reference panel which includes 1092 individuals and over 36 million SNPs. From our previously-identified cohort of Olmsted County, MN residents with incident or recurrent VTE, 1996-2005, we derived and validated an EHR-driven phenotype extraction algorithm that leveraged structured data based on ICD-9-CM codes and unstructured data from clinical notes via natural language processing to identify VTE cases and controls with 100% and 94% positive and negative predictive values, respectively. We tested for an association between each SNP and VTE among AAs using unconditional logistic regression, adjusting for age, sex and eMERGE site. Results Among 294 AA VTE cases and 3,661 AA controls (total n=3,955; females, n=2,512), the Factor V Leiden (F5 rs6025) was not analyzed due to a very low minor allele frequency (MAF=0.0036). The prothrombin G20210A (F2 rs1799963) and ABO blood type O (ABO rs8176719) SNPs were not genotyped in any of the arrays and could not be imputed. Among SNPs with an imputation score 〉0.8, the most significant SNPs associated with VTE were ITPR3 (inositol 1,4,5-triphosphate receptor type 3) rs2229637 (OR=1.65; p=3.61E-07; MAF=0.19) and CLEC7A (C-type lectin domain family 7, member A) rs59819090 (OR=2.16; p=1.06E-06; MAF=0.056). ITPR3 SNPs have been associated with coronary artery aneurysm in Kawasaki disease, type 1 diabetes mellitus and other autoimmune disorders. CLEC7A rs59819090 encodes for a serine82 to leucine nonsynonymous amino acid change in dectin-1. Dectin-1 is a transmembrane innate immune pattern recognition receptor on myeloid cells that upon binding it’s agonist, β-glucans, stimulates cytokine production and the respiratory burst, a prerequisite for formation of neutrophil extracelluar traps (NETs). NETs have been associated with VTE (van Montfoort, et al. Ateriosclero Thromb Vasc Biol 2013;33:147-51). Dectin-1 serine82 is not evolutionarily conserved and the serine82 to leucine amino acid change is predicted to be tolerated, with a moderate effect on protein function. Conclusions ITPR3 rs2229637 and CLAC7A rs59819090 may be associated with VTE in African-Americans. These observations require replication and functional studies toward understanding how they may lie on the causal pathway to VTE. Disclosures: No relevant conflicts of interest to declare.

Beschreibung: In this work related to familial aggregation of familial venous thromboembolism, the investigators report genomic and transcriptomic association of 16 novel susceptibility loci for venous thromboembolism.

Beschreibung: Abstract 1242 Background: While bivariate interactions between Factor V Leiden, prothrombin G20201A and hereditary deficiency of antithrombin, protein C and protein S compound VTE risk, whether other gene-gene interactions are associated with VTE is largely unknown. Objective: To test novel gene-gene interactions for an association with VTE. Methods: Genome-wide scan (Illumina 660Q [557,112 SNPs]) and candidate gene (12,551 SNPs in 764 genes within the anticoagulant, procoagulant, fibrinolytic and innate immunity pathways) genotypes from 1270 non-Hispanic adults of European ancestry with objectively-diagnosed VTE (cases; no cancer, venous catheter or antiphospholipid antibodies) and 1302 controls (frequency-matched on case age, gender, race, MI/stroke status) were merged and imputed to about 2.5 million SNPs with MACH using HapMap Phase 2 (60 CEU). We tested all pairwise SNP-SNP interactions using a 2-stage procedure (fast epistasis in the first stage and age, gender, MI/stroke status and state of residence adjusted analysis using logistic regression in the second stage). Results: The mean ± standard deviation case and control ages were 54.4 ± 16.2 and 55.6 ± 15.8 years, respectively and 51% were female. Among 2.299×10^12 pairwise interactions tested, 900 million reached a p-value