A patient with type 2N von Willebrand disease is heterozygous for a new mutation: Gly22Glu. Demonstration of a defective expression of the second allele by the use of monoclonal antibodies.

We report the case of a Chinese patient who has subnormal von Willebrand factor (vWF) level and normal vWF multimeric pattern, but a lack of vWF capacity to bind factor VIII (FVIII). Exons 18 to 20 of the patient's vWF gene were analyzed by DGGE and a G2354 --> A substitution which changes the encoded amino acid sequence from Gly22 to Glu was identified. The patient is heterozygous for this substitution, creating a unique Sac I restriction site. Recombinant vWF (rvWF) containing the candidate mutation was transiently expressed in COS-7 cells. It was processed and secreted normally but failed to bind FVIII. FVIII binding ability of hybrid rvWF, obtained by cotransfection of normal and mutated expression vectors and corresponding to a heterozygous genotype, was moderately decreased. To explain this functional discrepancy between patient's plasma vWF and hybrid rvWF, we used anti-vWF monoclonal antibodies (MoAbs) as capture in an enzyme-linked immunosorbent assay test. MoAb 32B12 recognized both wild-type and mutated rvWFs whereas MoAb 418 did not recognize mutated rvWF. Because MoAb 418 also failed to capture the plasma vWF from propositus, it means that his second nonmutated allele is not expressed or expressed at a very low level.