Midazolam microdosing applied in early clinical development for drug-drug interaction assessment.

AIMS: We aimed to incorporate a pharmacologically inactive midazolam microdose into early clinical studies for the assessment of CYP3A drug-drug interaction liability. METHODS: Three early clinical studies were conducted with substances (Compounds A, B and C) which gave positive CYP3A perpetrator signals in vitro. A 75 µg dose of midazolam was administered alone (baseline CYP3A activity) followed by administration with the highest dose groups tested for each compound on Day 1/3 and Day 14 or Day 17. Midazolam exposure (AUC0-∞ , Cmax ) during administration with the test substances was compared to baseline data via an analysis of variance on log-transformed data. Partial AUC2-4 ratios were also compared to AUC0-∞ ratios using linear regression on log-transformed data. RESULTS: Test compound Cmax values exceeded relevant thresholds for drug-drug interaction liability. Midazolam concentrations were quantifiable over the full profiles for all subjects in all studies. Point estimates of the midazolam AUC0-∞ gMean ratios ranged from 108.3 to 127.1% for Compound A, from 93.3 to 114.5% for Compound B, and from 92.0 to 96.7% for the two highest dose groups of Compound C. Cmax gMean ratios were in the same range. Thus, no relevant drug-drug interactions were evident, based on the results of midazolam microdosing. AUC2-4 ratios from these studies were comparable to the AUC0-∞ ratios. CONCLUSION: Midazolam microdosing incorporated into early clinical studies is a feasible tool for reducing dedicated drug-drug interaction studies, meaning reduced subject burden. Limited sampling could further reduce subject burden, costs and needed resources.

Supra-therapeutic doses of roflumilast have no effect on cardiac repolarization in healthy subjects.

OBJECTIVE: This study assesses the potential of the phosphodiesterase 4 inhibitor roflumilast to affect cardiac repolarization. METHODS: In this randomized, placebo-controlled study (n = 80), 40 healthy subjects received moxifloxacin 400 mg p.o. (positive control for prolongation of QT/heart-rate corrected QT [QTc]) and another 40 received placebo. After a 1-day washout, subjects received placebo or ascending doses of roflumilast 500 (therapeutic dose), 750 or 1000 µg/day p.o., for 14, 7 and 14 days, respectively. QT intervals were measured and corrected for heart rate with a Fridericia algorithm (QTc(F)). The primary endpoint was the largest mean time-matched change in QTc(F) from baseline (day 1). Safety and tolerability were monitored. RESULTS: Moxifloxacin gave a maximum time-matched change in QTc(F) versus baseline of 6.79 ms. Repeated administration of roflumilast 500 and 1000 µg resulted in maximum QTc(F) changes from baseline of -3.23 and -4.81 ms, respectively, confirming the absence of any QT/QTc prolongation at therapeutic and supra-therapeutic dosing. There were no changes in other electrocardiographic variables or time intervals, and roflumilast was well tolerated. CONCLUSIONS: Repeated administration of roflumilast at doses ≤ 1000 µg/day had no effect on cardiac repolarization or overall cardiac safety evaluations in healthy subjects. CLINICAL TRIAL REGISTRATION NUMBER: ISRCTN63818313

Effect of steady-state enoxacin on single-dose pharmacokinetics of roflumilast and roflumilast N-oxide.

Roflumilast is an oral phosphodiesterase 4 (PDE4) inhibitor for the treatment of chronic obstructive pulmonary disease (COPD). It is metabolized by CYP1A2 and CYP3A4 to its primary metabolite, roflumilast N-oxide, through which >90% total PDE4 inhibitory activity (tPDE4i) is mediated. Fluoroquinolones, of which enoxacin is the most potent CYP1A2 inhibitor, are used to treat COPD exacerbations. This phase I, open, nonrandomized, fixed-sequence, 2-period study evaluated the effects of steady-state enoxacin on the single-dose pharmacokinetics of roflumilast and roflumilast N-oxide. Twenty healthy participants received roflumilast, 500 µg once daily, on days 1 and 12, and enoxacin, 400 mg twice daily, on days 7 to 18. Pharmacokinetic profiles were obtained for days 1 to 6 and 12 to 19. The safety and tolerability of all treatments were also assessed. In 19 evaluable participants, coadministration led to 56% higher mean systemic exposure, 20% higher mean peak concentrations, and 36% lower mean apparent oral clearance compared with roflumilast alone. For roflumilast N-oxide, 23% higher mean systemic exposure and 14% lower mean peak concentrations were seen after coadministration. Roflumilast was well tolerated both alone and in combination with enoxacin. A weak interaction was shown between roflumilast and enoxacin, as mean tPDE4i increased by 25%, but is unlikely to have clinical relevance.

Chronopharmacology of roflumilast: a comparative pharmacokinetic study of morning versus evening administration in healthy adults.

The human circadian system is known to affect the pharmacokinetics and pharmacodynamics of several classes of respiratory disease medications. The current study involving 16 healthy adults investigated if the time-of-day of dosing of roflumilast, a novel phosphodiesterase-4 inhibitor, affects its pharmacokinetics. The rate of drug absorption (t(max): 1.50 versus 2.00 h) and peak concentration at t(max) (C(max): 3.79 versus 3.06 µg/L) was slightly greater with morning than evening administration, but without clinical significance. The extent of drug absorption (AUC) and drug elimination (t(1/2)) did not differ between the two dosing times. The pharmacokinetics of the active main metabolite, roflumilast N-oxide, also was not affected by the time of drug administration. Finally, the safety and tolerability of roflumilast did not differ between the two different times of administration.

Effects of rifampicin on the pharmacokinetics of roflumilast and roflumilast N-oxide in healthy subjects.

AIMS: To evaluate the effect of co-administration of rifampicin, an inducer of cytochrome P450 (CYP)3A4, on the pharmacokinetics of roflumilast and roflumilast N-oxide. Roflumilast is an oral, once-daily phosphodiesterase 4 (PDE4) inhibitor, being developed for the treatment of chronic obstructive pulmonary disease. Roflumilast is metabolized by CYP3A4 and CYP1A2, with further involvement of CYP2C19 and extrahepatic CYP1A1. In vivo, roflumilast N-oxide contributes >90% to the total PDE4 inhibitory activity. METHODS: Sixteen healthy male subjects were enrolled in an open-label, three-period, fixed-sequence study. They received a single oral dose of roflumilast 500 microg on days 1 and 12 and repeated oral doses of rifampicin 600 mg once daily on days 5-15. Plasma concentrations of roflumilast and roflumilast N-oxide were measured for up to 96 h. Test/Reference ratios and 90% confidence intervals (CIs) of geometric means for AUC and C(max) of roflumilast and roflumilast N-oxide and for oral apparent clearance (CL/F) of roflumilast were estimated. RESULTS: During the steady-state of rifampicin, the AUC(0-infinity) of roflumilast decreased by 80% (point estimate 0.21; 90% CI 0.16, 0.27); C(max) by 68% (0.32; CI 0.26, 0.39); for roflumilast N-oxide, the AUC(0-infinity) decreased by 56% (0.44; CI 0.36, 0.55); C(max) increased by 30% (1.30; 1.15, 1.48); total PDE4 inhibitory activity decreased by 58% (0.42; 0.38, 0.48). CONCLUSIONS: Co-administration of rifampicin and roflumilast led to a reduction in total PDE4 inhibitory activity of roflumilast by about 58%. The use of potent cytochrome P450 inducers may reduce the therapeutic effect of roflumilast.

Effect of single and repeated doses of ketoconazole on the pharmacokinetics of roflumilast and roflumilast N-oxide.

Effects of single and multiple doses of oral ketoconazole on roflumilast and its active metabolite, roflumilast N-oxide, were investigated in healthy subjects. In study 1, subjects (n = 26) received oral roflumilast 500 microg once daily for 11 days and a concomitant 200-mg single dose of ketoconazole on day 11. In study 2, subjects (n = 16) received oral roflumilast 500 microg on days 1 and 11 and a repeated dose of ketoconazole 200 mg twice daily from days 8 to 20. Coadministration of single-dose ketoconazole with steady-state roflumilast increased the AUC of roflumilast by 34%; C(max) was unchanged. For roflumilast N-oxide, AUC and C(max) decreased by 12% and 20%, respectively. Repeated doses of ketoconazole increased the AUC and C(max) of roflumilast by 99% and 23%, respectively; for roflumilast N-oxide, AUC was unchanged, and C(max) decreased by 38%. No clinically relevant adverse events were observed. Coadministration of ketoconazole and roflumilast does not require dose adjustment of roflumilast.

Effect of fluvoxamine on the pharmacokinetics of roflumilast and roflumilast N-oxide.

OBJECTIVE: To investigate the effects of steady-state dosing of fluvoxamine, an inhibitor of cytochrome P450 (CYP) 1A2 and CYP2C19, on the pharmacokinetics of roflumilast, an oral, once-daily phosphodiesterase 4 (PDE4) inhibitor and its pharmacodynamically active metabolite roflumilast N-oxide. METHODS: In an open-label, non-randomised, one-sequence, two-period, two-treatment crossover study, 14 healthy subjects received a single oral dose of roflumilast 500 microg on study day 1. After a 6-day washout period, repeated doses of fluvoxamine 50 mg once daily were given from days 8 to 21. On day 15, roflumilast 500 microg and fluvoxamine 50 mg were taken concomitantly. Percentage ratios of test/reference (reference: roflumilast alone; test: roflumilast plus steady-state fluvoxamine) of geometric means and their 90% confidence intervals for area under the plasma concentration-time curve, maximum plasma concentration (roflumilast and roflumilast N-oxide) and plasma clearance of roflumilast were calculated. RESULTS: Upon co-administration with steady-state fluvoxamine, the exposure to roflumilast as well as roflumilast N-oxide increased by a factor of 2.6 and 1.5, respectively. Roflumilast plasma clearance decreased by a factor of 2.6, from 9.06 L/h (reference) to 3.53 L/h (test). The combined effect of fluvoxamine co-administration on roflumilast and roflumilast N-oxide exposures resulted in a moderate (i.e. 59%) increase in total PDE4 inhibitory activity. CONCLUSION: Co-administration of roflumilast and fluvoxamine affects the disposition of roflumilast and its active metabolite roflumilast N-oxide most likely via a potent dual pathway inhibition of CYP1A2 and CYP2C19 by fluvoxamine. The exposure increases observed for roflumilast N-oxide are suggested to be attributable to CYP2C19 co-inhibition by fluvoxamine and thus, are not to be expected to occur when roflumilast is co-administered with more selective CYP1A2 inhibitors.

Identification and characterization of CYP3A4*20, a novel rare CYP3A4 allele without functional activity.

BACKGROUND: The major drug-metabolizing enzyme cytochrome P450 (CYP) 3A4 is genetically conserved. One outlier of Brazilian descent was found in a clinical pharmacokinetic trial exhibiting a 6-fold higher exposure than expected to an investigational drug, shown to be a CYP3A4 substrate. We aimed to investigate the genetic background of this finding. METHODS: The allelic variant of the CYP3A4 gene present in the outlier was sequenced, and the corresponding complementary deoxyribonucleic acid was expressed in yeast and human embryonic kidney cells. The outlier was phenotyped by use of intravenous administration of 1 mg midazolam. Analysis of phenotype and genotype correlation was carried out. The prevalence of the new allele was screened for in a white population. RESULTS: We identified a subject who heterozygously carried a novel CYP3A4 allele, named CYP3A4*20, with a premature stop codon yielding a truncated protein. Heterologous expression revealed that the CYP3A4.20 enzyme does not incorporate heme and thus is devoid of catalytic activity. CYP3A phenotyping in vivo showed that CYP3A4*20 exhibits a clear genotype-phenotype correlation, demonstrated by the subject's low systemic midazolam clearance (2.99 mL x min(-1) x kg(-1)). Genotyping of a white German population (n = 428) and relatives of the subject, as well as a review of published CYP3A4 sequencing data, suggests that CYP3A4*20 is a rare variant allele (<0.06% in white subjects). CONCLUSIONS: CYP3A4*20 represents the first CYP3A4 allele to be identified that has been shown to be devoid of functional activity. It causes an intermediate CYP3A4 metabolizer phenotype in a heterozygous carrier. Subjects of this genotype might be susceptible to side effects during drug therapy with substrates or inhibitors of CYP3A4.