Uridine Glucuronosyltransferase 2B7 Polymorphism-Based Pharmacogenetic Dosing of Epirubicin in FEC Chemotherapy for Early-Stage Breast Cancer.

BACKGROUND: Epirubicin is metabolized by uridine glucuronosyltransferase 2B7 (UGT2B7). Patients homozygous for the minor allele (CC) in the UGT2B7 -161 promoter polymorphism have lower clearance and significantly higher rates of leukopenia compared to wild-type homozygote (TT) or heterozygote (CT) patients. This study was designed to determine if TT and CT genotype patients could tolerate a higher epirubicin dose compared to CC genotype patients. PATIENTS AND METHODS: We studied women with histologically confirmed non-metastatic, invasive breast cancer who were scheduled to receive at least three cycles of FE100C in the (neo)adjuvant setting. Patients received standard-dose FE100C during the first 21-day cycle. Based on genotype, the epirubicin dose was escalated in the second and third cycles to 115 and 130 mg/m2 or to 120 and 140 mg/m2 for CT and TT genotype patients, respectively. The main outcome measurements were myelosuppression and dose-limiting toxicity. These were analyzed for relationships with the three genotypes. RESULTS: Forty-five patients were enrolled (10 CC, 21 CT, and 14 TT genotypes) and received 100 mg/m2 of epirubicin in the first cycle. Twelve and 10 TT patients were dose escalated at the second and third cycles, respectively; 16 CT patients were dose escalated at the second and third cycles. Leukopenia, but not febrile neutropenia, was genotype and dose dependent and increased in patients with CT and TT genotypes as their dose was increased. However, the third-cycle leukopenia rates were comparable to patients with the CC genotype receiving standard-dose epirubicin. CONCLUSION: Pharmacogenetically guided epirubicin dosing is well tolerated and allowed dose escalation without increased toxicity.

Population Pharmacokinetic Modeling in the Presence of Missing Time-Dependent Covariates: Impact of Body Weight on Pharmacokinetics of Paracetamol in Neonates.

Body weight is the primary covariate in pharmacokinetics of many drugs and dramatically changes during the first weeks of life of neonates. The objective of this study is to determine if missing body weights in preterm and term neonates affect estimates of model parameters and which methods can be used to improve performance of a population pharmacokinetic model of paracetamol. Data for our analysis were obtained from previously published studies on the pharmacokinetics of intravenous paracetamol in neonates. We adopted a population model of body weight change in neonates to implement three previously introduced methods of handling missing covariates based on data imputation, likelihood function modification, and full random effects modeling. All models were implemented in NONMEM 7.4, and population parameters were estimated using the FOCE method. Our major finding was that missing body weights minimally affect population estimates of pharmacokinetic parameters but do affect the covariate relationship parameters, particularly the one describing dependence of clearance on body weight. None of the tested methods changed estimates of between-subject variability nor impacted the predictive performance of the model. Our analysis shows that a modeling approach towards handling missing covariates allows borrowing information gathered in various studies as long as they target the same population. This approach is particularly useful for handling time-dependent missing covariates.

Mathematical modeling of intracrine androgen metabolism in prostate cancer: Methodological aspects.

BACKGROUND: Progression of castration-recurrent/resistant prostate cancer (CRPC) relies in part on dihydrotestosterone derived from intratumoral androgen metabolism. Mathematical modeling provides a valuable tool for studies of androgen metabolism in CRPC. This modeling approach integrates existing knowledge about complex biologic systems and provides a means of interrogating the effects of various interventions. We sought to model a single reaction in the androgen biosynthesis network, namely the oxidation of androsterone (AND) to androstanedione (5α-dione) by four 3α-oxidoreductase enzymes, as an initial effort to establish the feasibility of our modeling approach. METHODS: Models were constructed for two cell culture systems, a non-prostate cancer cell line (CV-1) and a prostate cancer cell line (LAPC-4), using the SimBiology app (version 5.3) in MATLAB (version 8.6). The models included components for substrate (AND), product (5α-dione), each of the four enzymes, and each of the four enzyme-substrate complexes. Each enzymatic reaction consisted of a reversible enzyme-substrate binding step and an irreversible catalysis step. Rates of change for each component were described using ordinary differential equations. RESULTS: Mathematical models were developed with model parameter values derived from literature sources or from existing experimental data, which included gene expression measurements and substrate and product concentrations determined using liquid chromatography-tandem mass spectrometry. The models for both cell lines adequately described substrate and product concentrations observed after 12 h treatment with AND. CONCLUSIONS: This modeling approach represents an adaptable, extensible and mechanistic framework that reflects androgen metabolism. The models can be expanded systematically to describe the complex androgen metabolic pathways important for study of novel therapies for CRPC.

Polymorphic Expression of UGT1A9 is Associated with Variable Acetaminophen Glucuronidation in Neonates: A Population Pharmacokinetic and Pharmacogenetic Study.

INTRODUCTION: Acetaminophen (paracetamol, APAP) is widely used as an analgesic and antipyretic drug in children and neonates. A number of enzymes contribute to the metabolism of acetaminophen, and genetic factors might be important to explain variability in acetaminophen metabolism among individuals. METHODS: The current investigation utilized a previously published parent-metabolite population pharmacokinetic model describing acetaminophen glucuronidation, sulfation, and oxidation to examine the potential role of genetic variability on the relevant metabolic pathways. Neonates were administered 30-min intravenous infusions of acetaminophen 15 mg/kg every 12 h (< 28 weeks' gestational age [GA]) or every 8 h (≥ 28 weeks GA) for 48 h. A total of 18 sequence variations (SVs) in UDP-glucuronosyltransferase (UGT), sulfotransferase (SULT), and cytochrome P450 (CYP) genes from 33 neonates (aged 1-26 days) were examined in a stepwise manner for an effect on the metabolic formation clearance of acetaminophen by glucuronidation (UGT), sulfation (SULT), and oxidation (CYP). The stepwise covariate modeling procedure was performed using NONMEM® version 7.3. RESULTS: Incorporation of genotype as a covariate for one SV located in the UGT1A9 gene promoter region (rs3832043, - 118 > insT, T9 > T10) significantly improved model fit (likelihood ratio test, p < 0.001) and reduced between-subject variability in glucuronide formation clearance. Individuals with the UGT1A9 T10 polymorphism, indicating insertion of an additional thymidine nucleotide, had a 42% reduction in clearance to APAP-glucuronide as compared to their wild-type counterparts. CONCLUSION: This study shows a pharmacogenetic effect of an SV in the UGT1A9 promoter region on the metabolism of acetaminophen in neonates.

Neonatal Maturation of Paracetamol (Acetaminophen) Glucuronidation, Sulfation, and Oxidation Based on a Parent-Metabolite Population Pharmacokinetic Model.

OBJECTIVES: This study aimed to model the population pharmacokinetics of intravenous paracetamol and its major metabolites in neonates and to identify influential patient characteristics, especially those affecting the formation clearance (CLformation) of oxidative pathway metabolites. METHODS: Neonates with a clinical indication for intravenous analgesia received five 15-mg/kg doses of paracetamol at 12-h intervals (<28 weeks' gestation) or seven 15-mg/kg doses at 8-h intervals (≥28 weeks' gestation). Plasma and urine were sampled throughout the 72-h study period. Concentration-time data for paracetamol, paracetamol-glucuronide, paracetamol-sulfate, and the combined oxidative pathway metabolites (paracetamol-cysteine and paracetamol-N-acetylcysteine) were simultaneously modeled in NONMEM 7.2. RESULTS: The model incorporated 259 plasma and 350 urine samples from 35 neonates with a mean gestational age of 33.6 weeks (standard deviation 6.6). CLformation for all metabolites increased with weight; CLformation for glucuronidation and oxidation also increased with postnatal age. At the mean weight (2.3 kg) and postnatal age (7.5 days), CLformation estimates (bootstrap 95% confidence interval; between-subject variability) were 0.049 L/h (0.038-0.062; 62 %) for glucuronidation, 0.21 L/h (0.17-0.24; 33 %) for sulfation, and 0.058 L/h (0.044-0.078; 72 %) for oxidation. Expression of individual oxidation CLformation as a fraction of total individual paracetamol clearance showed that, on average, fractional oxidation CLformation increased <15 % when plotted against weight or postnatal age. CONCLUSIONS: The parent-metabolite model successfully characterized the pharmacokinetics of intravenous paracetamol and its metabolites in neonates. Maturational changes in the fraction of paracetamol undergoing oxidation were small relative to between-subject variability.

Population Pharmacokinetics of Intravenous Paracetamol (Acetaminophen) in Preterm and Term Neonates: Model Development and External Evaluation.

OBJECTIVES: The aims of this study were to develop a population pharmacokinetic model for intravenous paracetamol in preterm and term neonates and to assess the generalizability of the model by testing its predictive performance in an external dataset. METHODS: Nonlinear mixed-effects models were constructed from paracetamol concentration-time data in NONMEM 7.2. Potential covariates included body weight, gestational age, postnatal age, postmenstrual age, sex, race, total bilirubin, and estimated glomerular filtration rate. An external dataset was used to test the predictive performance of the model through calculation of bias, precision, and normalized prediction distribution errors. RESULTS: The model-building dataset included 260 observations from 35 neonates with a mean gestational age of 33.6 weeks [standard deviation (SD) 6.6]. Data were well-described by a one-compartment model with first-order elimination. Weight predicted paracetamol clearance and volume of distribution, which were estimated as 0.348 L/h (5.5 % relative standard error; 30.8 % coefficient of variation) and 2.46 L (3.5 % relative standard error; 14.3 % coefficient of variation), respectively, at the mean subject weight of 2.30 kg. An external evaluation was performed on an independent dataset that included 436 observations from 60 neonates with a mean gestational age of 35.6 weeks (SD 4.3). The median prediction error was 10.1 % [95 % confidence interval (CI) 6.1-14.3] and the median absolute prediction error was 25.3 % (95 % CI 23.1-28.1). CONCLUSIONS: Weight predicted intravenous paracetamol pharmacokinetics in neonates ranging from extreme preterm to full-term gestational status. External evaluation suggested that these findings should be generalizable to other similar patient populations.

Disease Progression Modeling: Key Concepts and Recent Developments.

Disease modeling involves the use of mathematical functions to describe quantitatively the time course of disease progression. In order to characterize the natural progression of disease, these models generally incorporate longitudinal data for some biomarker(s) of disease severity or can incorporate more direct measures of disease severity. Disease models are also often linked to pharmacokinetic-pharmacodynamic models so that the influence of drug treatment on disease progression can be quantified and evaluated. Regulatory agencies have embraced disease progression models as powerful tools that can be used to improve drug development productivity. This article provides a brief overview of key concepts in disease progression modeling followed by illustrative examples from models for Alzheimer's disease. Finally, recent novel applications in which disease progression models have been linked to cost-effectiveness analysis and genomic analysis are described.

Simultaneous quantification of acetaminophen and five acetaminophen metabolites in human plasma and urine by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry: Method validation and application to a neonatal pharmacokinetic study.

Drug metabolism plays a key role in acetaminophen (paracetamol)-induced hepatotoxicity, and quantification of acetaminophen metabolites provides critical information about factors influencing susceptibility to acetaminophen-induced hepatotoxicity in clinical and experimental settings. The aims of this study were to develop, validate, and apply high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) methods for simultaneous quantification of acetaminophen, acetaminophen-glucuronide, acetaminophen-sulfate, acetaminophen-glutathione, acetaminophen-cysteine, and acetaminophen-N-acetylcysteine in small volumes of human plasma and urine. In the reported procedures, acetaminophen-d4 and acetaminophen-d3-sulfate were utilized as internal standards (IS). Analytes and IS were recovered from human plasma (10µL) by protein precipitation with acetonitrile. Human urine (10µL) was prepared by fortification with IS followed only by sample dilution. Calibration concentration ranges were tailored to literature values for each analyte in each biological matrix. Prepared samples from plasma and urine were analyzed under the same HPLC-ESI-MS/MS conditions, and chromatographic separation was achieved through use of an Agilent Poroshell 120 EC-C18 column with a 20-min run time per injected sample. The analytes could be accurately and precisely quantified over 2.0-3.5 orders of magnitude. Across both matrices, mean intra- and inter-assay accuracies ranged from 85% to 112%, and intra- and inter-assay imprecision did not exceed 15%. Validation experiments included tests for specificity, recovery and ionization efficiency, inter-individual variability in matrix effects, stock solution stability, and sample stability under a variety of storage and handling conditions (room temperature, freezer, freeze-thaw, and post-preparative). The utility and suitability of the reported procedures were illustrated by analysis of pharmacokinetic samples collected from neonates receiving intravenous acetaminophen.

A Population Pharmacokinetic Analysis of Dextroamphetamine in the Plasma and Hair of Healthy Adults.

BACKGROUND AND OBJECTIVE: Hair is an attractive matrix for amphetamine drug testing; however, little is known about the rate at which amphetamines are deposited into hair. Therefore, the purpose of this study was to determine the pharmacokinetics of oral dextroamphetamine in plasma and quantify the rate of deposition into hair in healthy adults using a linked population pharmacokinetic model. METHODS: Healthy adults >18 years of age received dextroamphetamine 10 mg orally for 7 days. Plasma samples were collected over 48 h following the final dose, and hair was collected 5 weeks following the first dose. NONMEM 7.2 was used to estimate dextroamphetamine oral absorption rate constant, apparent clearance and volume of distribution of the plasma compartment, the plasma to hair incorporation rate constant, and the apparent volume of distribution in the hair compartment. RESULTS: Dextroamphetamine pharmacokinetics were well-described by a one-compartment model with combined additive and proportional error for the plasma compartment, which was linked to a single compartment for the hair. Apparent clearance and volume of distribution in the plasma compartment were scaled by current body weight (centered on the mean). Melanin hair concentration was included as a significant covariate on the hair compartment. Absorption rate constant, clearance, and volume of distribution for the plasma compartment were estimated as 0.527 h(-1) (95% CI 0.467-0.586), 28.7 L/h (95% CI 27.1-30.3), and 377 L (95% CI 326-428), respectively. The incorporation rate constant from plasma to hair was 1.60e(-6) h(-1) (95% CI 1.06e(-6)-2.14e(-6)) and apparent volume of distribution in hair was 17.7 mg (95% CI 12.5-22.8). CONCLUSIONS: A one-compartment plasma model linked to a single compartment for hair successfully described the pharmacokinetics of dextroamphetamine in healthy adults. The volume of distribution and clearance of dextroamphetamine increased with weight, and the volume of distribution of the hair compartment increased with greater melanin concentrations.

Time course of acetaminophen-protein adducts and acetaminophen metabolites in circulation of overdose patients and in HepaRG cells.

1. It has been suggested that acetaminophen (APAP)-protein adducts can be measured in circulation to diagnose APAP-induced liver injury. However, the full-time course of plasma adducts has not been studied specifically in early-presenting overdose patients. In fact, surprisingly little work has been done on the metabolism of APAP after overdose in general. 2. We measured APAP, five APAP metabolites and APAP-protein adducts in plasma samples from early- and late-presenting overdose patients, and APAP-protein adducts in culture medium from HepaRG cells. 3. In contrast to earlier rodents studies, we found that APAP-protein adducts were lower at early time points and peaked around the time of peak liver injury, suggesting that these adduct levels may take longer to become elevated or remain elevated than previously thought. 4. APAP and its major metabolites were elevated in plasma at early time points and rapidly decreased. 5. Although clinical measurement of APAP-protein adducts holds promise as a diagnostic tool, we suggest caution in its interpretation in very early-presenting patients. Our data also support the idea that sulfation is saturated even at low doses but glucuronidation has a much higher capacity, highlighting the importance of glucuronidation in APAP metabolism.

Quantification of a biomarker of acetaminophen protein adducts in human serum by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry: clinical and animal model applications.

The aims of this study were to develop, validate, and apply a high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) method for quantification of protein-derived 3-(cystein-S-yl)-acetaminophen (APAP-Cys) in human serum. Formation of acetaminophen (APAP) protein adducts is thought to be a critical, early event in the development of APAP-induced hepatotoxicity, and quantification of these protein adducts in human serum represents a valuable tool for assessment of APAP exposure, metabolism, and toxicity. In the reported procedure, serum samples were first dialyzed or passed through gel filtration columns to remove APAP-Cys not covalently bound to proteins. Serum eluates were then subjected to enzymatic protease digestion to liberate protein-bound APAP-Cys. Norbuprenorphine-D3 was utilized as an internal standard (IS). APAP-Cys and IS were recovered from digested serum by protein precipitation with acetonitrile, and sample extracts were analyzed by HPLC-ESI-MS/MS. The method was validated by assessment of intra- and inter-assay accuracy and imprecision on two different analytical instrument platforms. APAP-Cys could be accurately quantified from 0.010 to 10µM, and intra- and inter-assay imprecision were <15% on both analytical instruments. APAP-Cys was stable in human serum for three freeze-thaw cycles and for 24h at ambient temperature. Extracted samples were stable when stored in refrigerated autosamplers for the typical duration of analysis or when stored at -20°C for six days. Results from process efficiency and matrix effect experiments indicated adequate recovery from human serum and insignificant ion suppression or enhancement. The utility and sensitivity of the reported procedure were illustrated by analysis of clinical samples collected from subjects taking chronic, therapeutic doses of APAP. Applicability to other biological matrices was also demonstrated by measurement of protein-derived APAP-Cys in plasma collected from APAP-treated mice, a common animal model of APAP-induced hepatotoxicity.