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The expression of cytochrome P450 (CYP) enzymes is highly variable and associated with factors, such as age, genotype, sex, and disease states. In this study, quantification of metronidazole metabolizing CYP isoforms (CYP2A6, CYP2E1, CYP3A4, CYP3A5, and CYP3A7) in human liver microsomes from 115 children and 35 adults was performed using a quantitative proteomics method. The data confirmed age‐dependent increase in CYP2A6, CYP2E1, and CYP3A4 abundance, whereas, as expected, CYP3A7 abundance showed postnatal decrease with age. In particular, the fold difference (neonatal to adulthood levels) in the protein abundance of CYP2A6, CYP2E1, and CYP3A4 was 14, 11, and 20, respectively. In contrast, protein abundance of CYP3A7 was > 125‐fold higher in the liver microsomes of neonates than of adults. The abundance of CYP2A6 and CYP3A5 was associated with genotypes, rs4803381 and rs776746, respectively. A proteomics‐informed physiologically based pharmacokinetic (PBPK) model was developed to describe the pharmacokinetics of metronidazole and its primary metabolite, 2‐hydroxymethylmetronidazole. The model revealed an increase in the metabolite‐to‐parent ratio with age and showed a strong correlation between CYP2A6 abundance and metabolite formation (r2 = 0.75). Notably, the estimated contribution of CYP3A7 was ~ 75% in metronidazole clearance in neonates. These data suggest that variability in CYP2A6 and CYP3A7 in younger children poses the risk of variable pharmacokinetics of metronidazole and its active metabolite with a potential impact on drug efficacy and safety. No sex‐dependent difference was observed in the protein abundance of the studied CYPs. The successful integration of hepatic CYP ontogeny data derived from a large liver bank into the pediatric PBPK model of metronidazole can be extended to other drugs metabolized by the studied CYPs. [ABSTRACT FROM AUTHOR]

AimWe sought to discover endogenous urinary biomarkers of human CYP2D6 activity.Patients & methodsHealthy pediatric subjects (n = 189) were phenotyped using dextromethorphan and randomized for candidate biomarker selection and validation. Global urinary metabolomics was performed using liquid chromatography quadrupole time-of-flight mass spectrometry. Candidate biomarkers were tested in adults receiving fluoxetine, a CYP2D6 inhibitor.ResultsA biomarker, M1 (m/z 444.3102) was correlated with CYP2D6 activity in both the pediatric training and validation sets. Poor metabolizers had undetectable levels of M1, whereas it was present in subjects with other phenotypes. In adult subjects, a 9.56-fold decrease in M1 abundance was observed during CYP2D6 inhibition.ConclusionIdentification and validation of M1 may provide a noninvasive means of CYP2D6 phenotyping.

Byline: J. Steven Leeder, PharmD, PhD (Section of Developmental Pharmacology and Experimental Therapeutics, Department of Pediatrics, Children's Mercy Hospitals and Clinics, and University of Missouri-Kansas City, Kansas City, Missouri, sleeder@cmh.edu); [...]

The 13C‐pantoprazole breath test (PAN‐BT) is a safe, noninvasive, in vivo CYP2C19 phenotyping probe for adults. Our objective was to evaluate PAN‐BT performance in children, with a focus on discriminating individuals who, according to guidelines from the Clinical Pharmacology Implementation Consortium (CPIC), would benefit from starting dose escalation versus reduction for proton pump inhibitors (PPIs). Children (n = 65, 6–17 years) genotyped for CYP2C19 variants *2, *3, *4, and *17 received a single oral dose of 13C‐pantoprazole. Plasma concentrations of pantoprazole and its metabolites, and changes in exhaled 13CO2 (termed delta‐over‐baseline or DOB), were measured 10 times over 8 h using high performance liquid chromatography with ultraviolet detection and spectrophotometry, respectively. Pharmacokinetic parameters of interest were generated and DOB features derived using feature engineering for the first 180 min postadministration. DOB features, age, sex, and obesity status were used to run bootstrap analysis at each timepoint (Ti) independently. For each iteration, stratified samples were drawn based on genotype prevalence in the original cohort. A random forest was trained, and predictive performance of PAN‐BT was evaluated. Strong discriminating ability for CYP2C19 intermediate versus normal/rapid metabolizer phenotype was noted at DOBT30 min (mean sensitivity: 0.522, specificity: 0.784), with consistent model outperformance over a random or a stratified classifier approach at each timepoint (p < 0.001). With additional refinement and investigation, the test could become a useful and convenient dosing tool in clinic to help identify children who would benefit most from PPI dose escalation versus dose reduction, in accordance with CPIC guidelines. [ABSTRACT FROM AUTHOR]

rs5758550 has been associated with enhanced transcription and suggested to be a useful marker of CYP2D6 activity. As there are limited and inconsistent data regarding the utility of this distant "enhancer" single nucleotide polymorphism (SNP), our goal was to further assess the impact of rs5758550 on CYP2D6 activity toward two probe substrates, atomoxetine (ATX) and dextromethorphan (DM), using in vivo urinary metabolite (DM; n = 188) and pharmacokinetic (ATX; n = 70) and in vitro metabolite formation (ATX and DM; n = 166) data. All subjects and tissues were extensively genotyped, the "enhancer" SNP phased with established CYP2D6 haplotypes either computationally or experimentally, and the impact on CYP2D6 activity investigated using several linear models of varying complexity to determine the proportion of variability in CYP2D6 activity captured by each model. For all datasets and models, the "enhancer" SNP had no or only a modest impact on CYP2D6 activity prediction. An increased effect, when present, was more pronounced for ATX than DM suggesting potential substate‐dependency. In addition, CYP2D6*2 alleles with the "enhancer" SNP were associated with modestly higher metabolite formation rates in vitro, but not in vivo; no effect was detected for CYP2D6*1 alleles with "enhancer" SNP. In summary, it remains inconclusive whether the small effects detected in this investigation are indeed caused by the "enhancer" SNP or are rather due to its incomplete linkage with other variants within the gene. Taken together, there does not appear to be sufficient evidence to warrant the "enhancer" SNP be included in clinical CYP2D6 pharmacogenetic testing. [ABSTRACT FROM AUTHOR]

The ontogeny of hepatic uridine diphosphate‐glucuronosyltransferases (UGTs) was investigated by determining their protein abundance in human liver microsomes isolated from 136 pediatric (0–18 years) and 35 adult (age >18 years) donors using liquid chromatography / tandem mass spectrometry (LC‐MS/MS) proteomics. Microsomal protein abundances of UGT1A1, UGT1A4, UGT1A6, UGT1A9, UGT2B7, and UGT2B15 increased by ∼8, 55, 35, 33, 8, and 3‐fold from neonates to adults, respectively. The estimated age at which 50% of the adult protein abundance is observed for these UGT isoforms was between 2.6–10.3 years. Measured in vitro activity was generally consistent with the protein data. UGT1A1 protein abundance was associated with multiple single nucleotide polymorphisms exhibiting noticeable ontogeny–genotype interplay. UGT2B15 rs1902023 (*2) was associated with decreased protein activity without any change in protein abundance. Taken together, these data are invaluable to facilitate the prediction of drug disposition in children using physiologically based pharmacokinetic modeling as demonstrated here for zidovudine and morphine. [ABSTRACT FROM AUTHOR]

CYP2B6*6 and CYP2B6*18 are the most clinically important variants causing reduced CYP2B6 protein expression and activity. However, these variants do not account for all variability in CYP2B6 activity. Emerging evidence has shown that genetic variants in the 3′UTR may explain variable drug response by altering microRNA regulation. Five 3′UTR variants were associated with significantly altered efavirenz AUC0‐48 (8‐OH‐EFV/EFV) ratios in healthy human volunteers. The rs70950385 (AG>CA) variant, predicted to create a microRNA binding site for miR‐1275, was associated with a 33% decreased CYP2B6 activity among normal metabolizers (AG/AG vs. CA/CA (P < 0.05)). In vitro luciferase assays were used to confirm that the CA on the variant allele created a microRNA binding site causing an 11.3% decrease in activity compared to the AG allele when treated with miR‐1275 (P = 0.0035). Our results show that a 3′UTR variant contributes to variability in CYP2B6 activity. [ABSTRACT FROM AUTHOR]