Comparison of functional testing for resistance to activated protein C and molecular biological testing for factor V R506Q in 370 patients.

OBJECTIVE: To compare functional and molecular biological tests for resistance to activated protein C (APC)/factor V R506Q, the most common cause of familial thrombosis. METHODS: We developed functional and molecular biological tests for resistance to APC/factor V R506Q at our institution and correlated the results for 370 patients studied by both methods. The functional method is based on addition of exogenous APC to an activated partial thromboplastin time-based assay. The molecular biological method is based on polymerase chain reaction followed by endonuclease digestion. RESULTS: Considering the molecular biological test as definitive for detecting the factor V R506Q mutation, the sensitivity of the functional assay was 100%, and the specificity was 74%. The prevalence of the factor V mutation in the population studied was 12% (41 heterozygotes, two homozygotes), and the positive predictive value of the functional assay was 34%. Although a normalized sensitivity ratio (nAPC-SR) less than 0.84 is considered evidence of resistance to APC by functional testing, we found that all patients with factor V R506Q had an nAPC-SR less than or equal to 0.71. When this alternative positive cutoff was used, the specificity of the functional test for factor V R506Q increased to 87%, and the positive predictive value increased to 52%, which constituted a significant improvement. We compared clinical findings from patients with resistance to APC with or without the presence of factor V R506Q, and found that as a group, those with factor V R506Q had a higher incidence of hypercoagulability, but fewer additional risk factors for hypercoagulability. The mechanism of resistance to APC in factor V R506Q-negative individuals is unclear, but may be related to other risk factors for hypercoagulability. CONCLUSIONS: The functional assay for resistance to APC is an excellent screening test for factor V R506Q, but confirmatory molecular biological testing is necessary when the functional test is positive, because of the high false-positive rate.