The FVII R402X nonsense mutation, associated with an asymptomatic phenotype, is responsible for small amounts of circulating protein with improved coagulant activity

Rationale: Factor VII (FVII) deficiency is associated to variable bleeding tendency, only partially explained by FVII gene mutations. Nonsense changes are often associated with a severe bleeding tendency. We investigated an asymptomatic girl, homozygous for the R402X nonsense change, which affects the carboxy-terminus of the FVII molecule (wild-type stop codon at position 407). Results: FVII antigen and coagulant levels in the proposita’s plasma were 0.6–0.8% and 3–5% of pooled normal plasma (PNP), respectively, thus suggesting the presence of small amounts of truncated molecules (FVII-402X) with improved coagulant activity. Functional FXa (1.8% ± 0.5% of PNP) and Thrombin generation assays (1.5% ± 0.1%) corroborated these observations. Since the carboxyl-terminal region is crucial for secretion of the highly homologous Protein C and Factor IX, we investigated this issue by expression of a series of truncated, recombinant FVII (rFVII) variants. An inverse relationship between the deletion extent and the secreted protein levels was detected for the rFVII-406X (50– 60% of rFVII-wt), rFVII-405X (15–20%) and rFVII-404X (9–12%) variants, which displayed a normal specific activity in FXa assays. The rFVII-403X and rFVII-402X were poorly secreted (< 1%). Intriguingly, upon concentration of conditioned media, the specific activity of the rFVII-402X appeared to be two to threefold higher than that of rFVII-wt. Conclusions: The R402X nonsense mutation is responsible for small amounts of secreted FVII, characterized by supra-normal coagulant activity. Our data highlight the role of the FVII carboxyl-terminal residues for protein secretion and function, and provide the rationale for the asymptomatic phenotype in the homozygous proposita.