Genotyping cytochrome P450 3A5 using the Light Cycler.
Cytochrome P450 3A5 (CYP3A5) is involved in the biotransformation of many orally administered drugs, some of which are dose optimized using therapeutic drug monitoring. The CYP3A5 gene exhibits variable inter-individual expression, which is related to a single-nucleotide polymorphism. Producers of the enzyme possess at least one CYP3A5*1 allele. Knowledge of patients' CYP3A5 genotype, in conjunction with therapeutic drug monitoring (TDM), may aid patient management. Real-time polymerase chain reaction (PCR) was used to genotype the A6986G mutation of the CYP3A5 gene. Specific primers were employed to generate a DNA product, co-amplified with two internal hybridization probes, using a LightCycler. DNA melt curve analysis readily identified the genotypes CYP3A5*1/*1, CYP3A5*1/*3 and CYP3A5*3/*3. Results were confirmed using DNA sequencing with 100% correlation. Genotypes were determined from 263 individuals and compared with the genotypes of a pseudogene CYP3AP1, which is in disequilibrium with CYP3A5. This is a rapid and reliable method for genotyping the CYP3A5 polymorphism which may be used as an important adjunct to the TDM service offered by laboratories to optimize drug prescription.
Fredericks
Salim
S
Analytical Unit, Cardiac and Vascular Sciences, St George's Hospital Medical School, St George's University of London, Cranmer Terrace, London SW17 0RE, UK. frederic@sgul.ac.uk
Moreton
Michelle
M
MacPhee
Iain A M
IA
Mohamed
Maha
M
Marlowe
Sharon
S
Jorga
AnaMaria
A
Johnston
Atholl
A
Carter
Nicholas D
ND
Holt
David W
DW
D001483:Base Sequence; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D017931:DNA Primers; D016903:Drug Monitoring / Q000379:methods; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D003577:Cytochrome P-450 Enzyme System; C104464:CYP3A protein, human; C510164:CYP3A5 protein, human; D051544:Cytochrome P-450 CYP3A