[Validation of a diagnostic kit for factor V Leiden mutation]. / Validation d'un kit de diagnostic de la mutation Leiden du facteur V.

The Stagen factor V Leiden mutation is a new kit allowing to perform all steps necessary to identify the Leiden mutation of Factor V, from DNA extraction from patient's blood sample to electrophoresis of amplification products. The method is based on an allele specific amplification, which allows patient's genotype to be established in a single step. Analytical properties of the kit have been tested first, and revealed the robustness of the kit. In a second step, a prospective study of a cohort of 300 thrombophilic patients demonstrated the specificity and the sensitivity of the method. In additions, several controls and methodological ingenious processes allow to minimize the risk of human or method errors, making the reliability of the kit. Finally, the Stagen factor V Leiden mutation is easy and rapid to use in hospital or private laboratories, and is well-suited for small series of patients of the latter.

Homozygosity for the R1268Q mutation in MRP6, the pseudoxanthoma elasticum gene, is not disease-causing.

Pseudoxanthoma elasticum (PXE) is an inherited systemic disorder of connective tissue, characterized by progressive calcification of the elastic fibers in the eye, the skin, and the cardiovascular system, resulting in decreased vision, skin lesions, and life-threatening vascular disease, with highly variable phenotypic expression. The PXE locus has been mapped to chromosome 16p13.1, and was recently further refined to a 500 kb-region, containing two pseudogenes and four candidate genes. In a comprehensive mutational screening, we were able to exclude the responsibility of pM5, UNK, and MRP1 genes, candidate on the basis of their genetic localization. Conversely, we have found pathogenetic mutations in the MRP6 gene, in patients affected with PXE, indicating that human MRP6, which encodes a 1503 amino-acids membrane protein, member of the human ATP binding cassette (ABC) transporters superfamily, is the gene responsible for PXE. In one large PXE pedigree for which we had identified a nonsense mutation (R1141X), we came across a G to A transition at position 3803 of the MRP6 cDNA sequence (R1268Q). Astonishingly, this latter variant was found at the homozygous state in the proband's unaffected husband. We investigated the R1268Q mutation, and found the Q1268 allele at a relatively high frequency (0.19) in a Caucasian control population (n = 62 subjects). Genotype frequencies were in Hardy-Weinberg equilibrium, and three healthy volunteers were homozygous for the Q1268 allele. These data indicate that the R1268Q variant in the MRP6 gene does not cause PXE per se. Further studies will elucidate if it may play a role when found in compound heterozygotes.