Exome Sequencing in Venous Thromboembolic Disease Identifies Excess Mutation Burden in PROS1, PROC, SERPINC1 and STAB2

Abstract

Deep vein thrombosis and pulmonary embolism, collectively referred to as venous thromboembolism (VTE), are the third leading cause of cardiovascular death in the United States. Genetic factors account for 50-60% of VTE risk and a recent meta-analysis of genome-wide association studies confirmed that common variants in F5, ABO, and seven other loci are associated with VTE. Rare mutations in the anticoagulant genes PROC, PROS1 and SERPINC1 have been linked to VTE in family studies. In order to identify new genetic variants altering the risk for VTE, we performed whole exome sequencing (WES) in 373 unrelated individuals of European ancestry with unprovoked VTE and compared results to a previously sequenced control cohort of 5784 unrelated Europeans. To avoid variant calling bias, only SNVs from exons with >10X coverage and less than 5% difference in coverage between cases and controls were included, removing 11,813 of 188,689 intervals. We used an emerging framework for a "collapsing" analysis on genes, defining qualifying variants on the basis of annotation and minor allele frequency <0.05%, and assumed a dominant model of inheritance. Tests were performed via a Fisher's exact test for a total of 11,585 CCDS genes. Strikingly, ranked by p-value, the top 4 genes were PROS1 (P= 2.01E-09, OR 11.8), STAB2 (P=2.70E-7, OR 3.37), PROC (P=3.24E-05, OR 11.0) and SERPINC1 (P=1.10E-04, OR 8.5). We detected 29 qualifying variants in 373 cases and 106 qualifying variants in 5784 controls in STAB2. This gene encodes Stabilin-2, which is a transmembrane glycoprotein scavenger receptor. Common variants at ABO are associated with VTE and are also known to regulate von Willebrand Factor (VWF) and coagulation Factor VIII (F8) plasma levels. Common variants in STAB2 are also associated with VWF/F8 levels in a large GWAS and with VTE risk in a smaller candidate gene study, suggesting that haploinsufficiency for Stabilin-2 may increase VTE risk through elevated levels of VWF/F8. Although replication and functional testing of these findings is warranted, this study demonstrates the utility of collapsing analyses using WES data to identify multiple loci harboring an excess of rare variants in individuals with a common complex disease trait. Disclosures Ginsburg: Shire: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties: recombinant VWF and recombinant ADAMTS13; Portola Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

Genetic regulation of plasma von Willebrand factor levels in health and disease

“…Pathogenic variants in STAB2 have been found to be associated with large elevations in plasma VWF:Ag levels (> 33%) in both association and exome sequencing studies , and modify the ability of stabilin‐2‐expressing cells to bind and internalize VWF in vitro . Pathogenic STAB2 variants have also been shown to be associated with the incidence of VTE ; however, given the scavenger function of stabilin‐2, altered clearance of non‐VWF plasma coagulation factors or inflammatory regulators may also contribute to the pathogenic basis of this observation.…”

What is currently known about the genetics of venous thromboembolism at the dawn of next generation sequencing technologies

“…Several other polymorphisms/genes have been proposed as risk factors for VTE but these lack compelling evidence, including robust statistical replication in independent studies, to be definitively claimed as susceptibility variants/genes for the disease. These include the rs2232710 of SERPINA10 , which encodes the Protein Z‐Dependent Protease Inhibitor of coagulation factors Xa and XIa (Folsom et al , ; Gorski et al , ); SERPINE1 rs2227631, influencing circulating plasminogen activator inhibitor‐1 (PAI‐1) levels (Huang et al , ); C4BPB/C4BPA rs3813948, affecting PS associated phenotypes (Buil et al , ); the rs7080536 of the HABP2 gene, encoding factor VII (FVII) activating protease (Ahmad‐Nejad et al , ); HIVEP1 rs169713 with, as yet, no associated intermediate phenotype (Morange et al , ); F13A1 rs5985 (Val34Leu) (Wells et al , ), influencing FXIII activity (Kohler et al , ; Wartiovaara et al , ); APOH rs8178847, associated with thrombin generation (Tang et al , ); genetic variations at the STAB 2 , STX2 and TC2N loci with suggestive impact on VTE risk (Germain et al , ; Morange et al , ; Smith et al , ; Desch et al , ) via their influence on VWF levels (Smith et al , ); and the rs4602861 (Germain et al , ; Klarin et al , ) at the ZFPM2 locus, whose suspected association with VTE could involve platelet count (Gieger et al , ) and vascular endothelial growth factor (Debette et al , ). Some of these associations, if true, could be restricted to some specific ethnic populations.…”

The scavenger receptor SCARA5 is an endocytic receptor for von Willebrand factor expressed by littoral cells in the human spleen

“…Although the associations between VWF and the receptors SR‐A1, asialoglycoprotein receptor, and LRP‐1 were characterized in candidate gene rationalized studies, the influence of stabilin‐2 on VWF clearance was first suggested by the Cohorts for Heart and Aging Research in Genome Epidemiology (CHARGE) genome‐wide association study (GWAS) meta‐analysis, which associated common variants with plasma levels of VWF or its binding partner the coagulation cofactor factor VIII (FVIII) . In follow‐up studies, rare and common STAB2 gene variants have been associated with VWF plasma levels in normal individuals and patients with type 1 VWD or low VWF, or an increased risk for venous thromboembolism . In addition to the STAB2 locus, the CHARGE GWAS identified variants in the genes encoding two other cell surface receptors, the endocytic lectin receptor CLEC4M and the scavenger receptor SCARA5 (Scavenger Receptor Class A Member 5, or SR‐A5), as being associated with plasma VWF and/or FVIII …”