Genome-Wide Association Transethnic Meta-Analyses Identifies Novel Associations Regulating Coagulation Factor VIII and von Willebrand Factor Plasma Levels

Abstract

Background: Factor VIII (FVIII) and its carrier protein von Willebrand factor (VWF) are associated with risk of arterial and venous thrombosis and with hemorrhagic disorders. We aimed to identify and functionally test novel genetic associations regulating plasma FVIII and VWF.

Methods: We meta-analyzed genome-wide association results from 46 354 individuals of European, African, East Asian, and Hispanic ancestry. All studies performed linear regression analysis using an additive genetic model and associated ≈35 million imputed variants with natural log-transformed phenotype levels. In vitro gene silencing in cultured endothelial cells was performed for candidate genes to provide additional evidence on association and function. Two-sample Mendelian randomization analyses were applied to test the causal role of FVIII and VWF plasma levels on the risk of arterial and venous thrombotic events.

Results: We identified 13 novel genome-wide significant ( P≤2.5×10-8) associations, 7 with FVIII levels ( FCHO2/TMEM171/TNPO1, HLA, SOX17/RP1, LINC00583/NFIB, RAB5C-KAT2A, RPL3/TAB1/SYNGR1, and ARSA) and 11 with VWF levels ( PDHB/PXK/KCTD6, SLC39A8, FCHO2/TMEM171/TNPO1, HLA, GIMAP7/GIMAP4, OR13C5/NIPSNAP, DAB2IP, C2CD4B, RAB5C-KAT2A, TAB1/SYNGR1, and ARSA), beyond 10 previously reported associations with these phenotypes. Functional validation provided further evidence of association for all loci on VWF except ARSA and DAB2IP. Mendelian randomization suggested causal effects of plasma FVIII activity levels on venous thrombosis and coronary artery disease risk and plasma VWF levels on ischemic stroke risk.

Conclusions: The meta-analysis identified 13 novel genetic loci regulating FVIII and VWF plasma levels, 10 of which we validated functionally. We provide some evidence for a causal role of these proteins in thrombotic events.

Keywords: cardiovascular diseases; factor VIII; genetics; genome-wide association studies; risk factors; von Willebrand factor.

Figures

Figure 1.. Manhattan plot for the transethnic analyses factor VIII (FVIII) and von Willebrand factor (VWF).

Representation of genome-wide results for FVIII (A) and VWF (B). Loci named by closest gene. Black indicates novel associations.

Figure 2.. Silencing candidate genes changes basal release and stimulated release of von Willebrand factor (VWF).

Human umbilical vein endothelial cells were transfected with short interfering RNA (siRNA) against selected genes for 4 days; the media was changed; and the cells were cultured for 30 minutes for basal release (A) or were stimulated with histamine 10 μmol/L for 30 minutes (B). VWF was then measured in the supernatant by an ELISA. n=4±SD. All results are relative to VWF release after transfection with a scrambled control siRNA, which is set as reference (100%). *P<0.05. **P<0.01. ***P<0.001.

Figure 3.. Mendelian randomization results.

Results show odds ratio (OR) (95% CI) per every higher SD change in FVIII (A) and von Willebrand factor (VWF; B). CAD indicates coronary artery disease; CVD, cardiovascular disease; FVIII, factor VIII; IVW, inverse-variance-weighted method; IVW.adjusted, IVW factor VIII adjusted for the effects of VWF; and VTE, venous thromboembolism.