II-90 Zdrav Vestn 2007; 76: SUPPL II
REVERSE-HYBRIDIZATION-BASED GENETIC TESTING FOR THE
PREDICTION OF ANTICOAGULANT DOSE REQUIREMENT
H. Puehringer H,1 C. Stoellberger,2 W. Krugluger,3 C. Oberkanins1
1 ViennaLab Diagnostics GmbH, Vienna, Austria
2 Second Medical Department, Rudolfstiftung Hospital, Vienna, Austria
3 Department of Clinical Chemistry, Rudolfstiftung Hospital, Vienna, Austria;
puehringer@viennalab.co.at
Background Coumarin derivatives (as warfarin or phenprocoumon) are the most widespread oral an-
ticoagulant drugs for the prevention and treatment of arterial and venous thromboembol-
ic disorders. However, these vitamin K antagonists have a narrow therapeutic range and a
wide interindividual variability in dose requirement. Despite adjustment for clinical vari-
ables adverse events (delay in achieving a stable maintenance dose or bleeding complica-
tions) are frequently encountered during the initial phase of therapy. Genetic polymor-
phisms in the drug-targeted vitamin K epoxide reductase complex 1 (VKORC1) and in the
drug metabolizing enzyme CYP2C9 have been reported to account for the majority of vari-
ations in the therapeutic response to warfarin.
Results We developed a genetic test (StripAssay) for the detection of -1639G > A and 3730G > A in
the VKORC1 gene, and 430C > T and 1075A > C in the CYP2C9 gene. The test is based on
multiplex PCR, followed by reverse-hybridization of biotin-labeled amplification products
to a parallel array of allele-specific oligonucleotides immobilized on membrane teststrips.
Genotyping for VKORC1 polymorphisms and the functionally defective CYP2C9 variants
*2 and *3 allowed the classification of patients into high, intermediate and low dose re-
sponders to phenprocoumon (Marcumar™), the most commonly used oral anticoagulant
in Central and North European countries. A rapid DNA extraction protocol, ready-to-use
reagents and teststrips, and the potential for automation of the hybridization/detection
step, make the StripAssay convenient and easy to perform within < 6 hours.
Conclusion A simple and reliable diagnostic tool was developed and evaluated for predicting the re-
sponse of patients to phenprocoumon treatment. The results obtained in our study will
assist clinicians to achieve a more individualized anticoagulant therapy.