In vivo metabolism of the major infertility drug clomiphene strongly depends on CYP2D6 and CYP3A4 activity

B Ganchev1, R Kerb1, M Turpeinen1,2, W Schroth1, GM Böhmer3, S Igel1, M Sonnenberg1, E Schaeffeler1, U Zanger1, H Brauch1, M Schwab1,3, TE Mürdter1. 1Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology and University of Tuebingen, 70376 Stuttgart, Germany, 2University of Oulu, Department of Pharmacology and Toxicology, 90014 Oulu, Finland, 3University Hospital Tuebingen, Department of Clinical Pharmacology, Institute of Experimental and Clinical Pharmacology, 72076 Tuebingen, Germany

Infertility affects 10 to 15% of couples worldwide. Clomiphene has been introduced into clinics in the 1960’s and still is the first line treatment for inducing ovulation in infertile women. Despite dosage escalation in non-responding women, about 15-30% of the patients fail to ovulate. Clomiphene is metabolized mainly by cytochrome P450 (CYP) 2D6 and 3A4 to various metabolites displaying differential activities at the estrogen receptor.

To study the impact of different CYPs in the bioactivation of clomiphene in vivo, we elucidated the influence of well-established inhibitors for CYP2D6 and 3A4 on the single dose pharmacokinetics of clomiphene.

Using in house synthesized reference compounds and stable isotope-labeled internal standards, a UHPLC-MS/MS method was established to quantify both clomiphene isomers comprised in pharmaceutical preparations as well as the respective metabolites. The method was applied in a pharmacokinetic drug-drug interaction study including 12 premenopausal volunteers with different CYP2D6 phenotypes. Plasma levels of clomiphene and its metabolites were determined after single dose of 100 mg clomiphene citrate as well as after co-administration of clarithromycin (CYP3A4 inhibitor) or paroxetine (CYP2D6 inhibitor). The study was approved by the Ethics Committee of the University of Tübingen, Germany and the German Federal Institute for Drugs and Medical Devices (BfArM), Bonn, Germany. All participants gave written informed consent prior to the study.

In volunteers with at least one functional CYP2D6 allele (extensive metabolizers, EM) maximum concentrations (Cmax) of (E)-4-hydroxyclomiphene (E-4-OH-Clom) and (E)-4-hydroxy-N-desethylclomiphene (E-4-OH-DE-Clom) were determined to be 22.8±12.4 and 13.1±3.2 nM, respectively. Participants with two non-functional CYP2D6 alleles (poor metabolizers, PM) showed remarkably lower concentrations of E-4-OH-Clom (Cmax 3.0±1.1 nM) and E-4-OH-DE-Clom (Cmax 1.1±0.1 nM). Co-administration of paroxetine reduced Cmax of E-4-OH-Clom and E-4-OH-DE-Clom in EMs up to 3-fold. Inhibition of CYP3A4 by clarithromycin led to 2 to 6-fold lower Cmax of (E)-N-desethylclomiphene in all participants. Additionally, Cmax of E-4-OH-DE-Clom decreased due to the inhibitory effect of clarithromycin on deethylation of E-4-OH-Clom.

Previous estrogen response element reporter assays have shown strongest antagonistic effects on the estrogen receptor for E-4-OH-Clom and E-4-OH-DE-Clom. Present pharmacokinetic data on clomiphene metabolism demonstrate a clear link between concentrations of the active metabolites and CYP2D6 phenotype. I nhibition of CYP3A4 by clarithromycin results in the reduced formation of the highly active metabolite E-4-OH-DE-Clom. Our data provide first evidence, that CYP2D6 polymorphism and CYP3A4 activity may play a major role in bioactivation of the infertility drug clomiphene.

This study was supported by Robert Bosch Foundation and the BMBF (03IS2061C and 01ZP0502), Germany.