Interpretation of Pharmacometabolomics Results: Fingerprint of Drug Exposure or Confounder Effects? Insights from a Urinary Metabolomics Study with Voriconazole in Healthy Participants

Interpretation of pharmacometabolomics results, aiming particularly at biomarker (sets) discovery for drug exposure, remains a major challenge. The metabotyping of drug exposure depends on resolution of specific metabolomics techniques and comprises individual metabolic phenotypes (“metabotypes”), disease-, drug- and microbiome-specific patterns, as well as conditional metabolic states (e. g. fasting). In this clinical trial with 16 healthy participants, an exploratory objective was to evaluate the untargeted urinary metabolomics of voriconazole, administered in four single doses, using proton nuclear magnetic resonance (1H-NMR) spectroscopy. Voriconazole is a second-generation triazole and a potent inhibitor of drug-metabolizing enzymes such as cytochrome P450 (CYP) isozymes CYP3A4 and CYP2C19. Therefore, identification of metabolites reflecting acute CYP3A4 inhibition was of particular interest. On two treatment days without and with voriconazole (with background microdosed midazolam and omeprazole administration for CYP3A4 and CYP2C19 phenotyping, respectively), spot urine was collected after overnight fasting (predose) and 4 h later (postdose fasting). In the postdose versus predose fingerprints, most changes at the annotated metabolite level were attributable to fasting metabolomics or potential confounders. 1H-NMR spectroscopy identified neither a short-term voriconazole-specific signature nor patterns or metabolites potentially reflecting acute CYP3A4 inhibition. Our study emphasizes crucial significance of strict standardization of fasting time and minimization of confounder influences by clinical trial design as well as selection of adequate baselines and high-resolution analytical techniques in pharmacometabolomics research, especially for biomarker discovery.