Characterisation of previously unknown protein C gene variants
S. Körber1, L. Böff1, F. Bergmann2, A. Czwalinna2, B. Bomke3, C. Heller1, M. Krause4, B. Luxembourg1, K. Kurnik5, W. Miesbach1, C. Geisen1 (1Frankfurt am Main, Germany, 2Hannover, Germany, 3Düsseldorf, Germany, 4Wiesbaden, Germany, 5Munich, Germany)
Time: 17:15 - 18:15
Objective: Hereditary deficiencies of natural anticoagulant proteins including protein C are known causes of thrombophilia. Up to now more than 350 different mutations in the Protein C gene (PROC) have been described. Our aim was to analyse the molecular defect underlying Protein C deficiency in a cohort of 314 patients and to predict the potential effect of previously unknown variants detected in this cohort.
Methods: DNA of 314 patients was analysed by Sanger Sequencing of the coding region of PROC including the exon intron boundaries. In cases without mutation MLPA was performed to detect large deletions or duplications. The characterisation of the variants was performed with in silico evaluation tools, including Polyhen-2, SIFT, multiple sequence alignment, splicing prediction and molecular graphic imaging.
Results: 101 different Mutations were detected in 226 patients, correlating with an overall mutation detection rate (MDR) for the PROC gene of 72,0 %. Altogether, 36 (35,6%) unknown and 65 (63,4%) previously described variants of PROC were identified. Of the unknown variants 23 were potentially missense changes and two were located in the intron, near to the exon intron boundary (potentially splice site changes). Of the previously unknown missense changes, one was classified as variant of unknown significance and 22 as potential missense mutations with pathogenic effect. The two variants located near the exon intron boundary were predicted as potential splice site mutations by in silico methods.
Conclusion: In silico methods and molecular graphic imaging are useful tools to predict the effect of genetic variants on the protein function or structure and by this to predict the pathogenic effect of certain variants. Nevertheless these methods cannot replace in vitro analysis by laboratory experiments, phenotypic studies of patients and cosegregation analysis within families.