A Novel Complete F8 Tandem Duplication Causing Elevated Factor VIII Activity and Associated with Venous...

Background Coagulation factor VIII (FVIII) is a critical component of the intrinsic coagulation pathway. While elevated FVIII levels are an established risk factor for venous thromboembolism (VTE), genetic variants in the F8 gene directly causing such elevations remain scarce. Here, we report a novel complete F8 tandem duplication identified in a female patient with splanchnic venous thrombosis (SVT). Methods We performed genetic testing using a thrombophilia panel targeting 35 genes involved in thrombosis and haemostasis to detect both point variants and copy number variations (CNVs). Family co-segregation analysis and phenotypic assays for FVIII and von Willebrand factor (VWF) were conducted. The structural basis of the identified F8 copy number gain was elucidated using optical genome mapping (OGM). Full-length F8 mRNA amplification, quantitative PCR, plasma FVIII Western blotting, and X-chromosome inactivation analysis were performed to assess the functional consequences of the duplication. Thrombin generation test (TGT) was employed to assess the hypercoagulable state. Results Genetic testing identified three copies of all 26 exons of the F8 gene in the proband, which was also detected in her mother (CNVs = 3) and son (CNVs = 2). One-stage clotting and chromogenic assays confirmed persistently elevated FVIII activity in the proband and her mother, accompanied by increased FVIII antigen levels. The OGM analysis confirmed a 229 kb tandem duplication including the F8 gene on one of the proband's X chromosomes. The junction regions exhibited high sequence homology and were rich in repetitive sequences, which precluded precise breakpoint mapping. Full-length F8 mRNA amplification revealed no aberrant transcripts, whereas quantitative PCR showed increased F8 mRNA expression in all carriers. Plasma FVIII Western blotting indicated FVIII heavy and light chains of expected molecular weights with increased band intensity in carriers. X-chromosome inactivation analysis in female carriers showed no significant skewing. TGT in two available carriers showed increased thrombin generation compared with a normal control at both low (1 pM) and high (5 pM) tissue factor concentrations. Conclusion We identified a novel complete F8 tandem duplication associated with increased FVIII expression and a hypercoagulable phenotype in a female patient with SVT. These findings support F8 gene dosage gain as a rare gain-of-function mechanism contributing to elevated FVIII levels and thrombophilia, while variation in VWF levels and acquired risk factors may modify thrombotic penetrance.