Characterization of missense mutations in the signal peptide and propeptide of FIX in hemophilia B by a cell-based assay.

Many mutations in the signal peptide and propeptide of factor IX (FIX) cause hemophilia B. A FIX variants database reports 28 unique missense mutations in these regions that lead to FIX deficiency, but the underlying mechanism is known only for the mutations on R43 that interfere with propeptide cleavage. It remains unclear how other mutations result in FIX deficiency and why patients carrying the same mutation have different bleeding tendencies.

[Development of a cell-based diagnostic system for vitamin K-dependent coagulation factor deficiency 1].

Objective: To develop a cell-based system for the diagnosis of vitamin K-dependent coagulation factor deficiency 1 (VKCFD1).

Methods: In HEK293 cells stably expressing the reporter gene FIX-Gla-PC, the gamma-glutamyl carboxylase (GGCX) gene was knocked out by using CRISPR/Cas9 technology. Enzyme-linked immunosorbent assay (ELISA), DNA sequencing and Western blotting were used to identify the GGCX gene knockout cells.