Physiologically based pharmacokinetic modeling to predict the impact of liver cirrhosis on glucuronidation via UGT1A4 & UGT2B7/2B4 - a case study with midazolam.

Hepatic impairment, due to liver cirrhosis, decreases the activity of cytochrome P450 enzymes (CYPs). The use of physiologically-based pharmacokinetic (PBPK) modeling to predict this effect for CYP substrates has been well-established, but the effect of cirrhosis on uridine-glucuronosyltransferase (UGT) activities is less studied and few PBPK models have been reported. UGT enzymes are involved in primary N-glucuronidation of midazolam and glucuronidation of 1'-OH-midazolam following CYP3A hydroxylation. In this study Simcyp® was used to establish PBPK models for midazolam, its primary metabolites midazolam-N-glucuronide (UGT1A4) and 1'-OH midazolam (CYP3A4/3A5) and the secondary metabolite 1'-OH-midazolam-O-glucuronide (UGT2B7/2B4), allowing to simulate the impact of liver cirrhosis on the primary and secondary glucuronidation of midazolam. The model was verified in non-cirrhotic subjects before extrapolation to cirrhotic patients of Child-Pugh (CP) classes A, B, and C. Our model successfully predicted the exposures of midazolam and its metabolites in non-cirrhotic and cirrhotic patients, with 86% of observed plasma concentrations within 5th-95th percentiles of predictions and observed geometrical mean of AUCinf and Cmax within 0.7-1.43-fold of predictions. The simulated metabolic ratio (AUCglucuronide/AUCparent, MR), was calculated for midazolam-N-glucuronide to midazolam (indicative of UGT1A4 activity) and decreased by 40% (CP A), 48% (CP B), and 75% (CP C). For 1'-OH-midazolam-O-glucuronide to 1'-OH-midazolam the MR (indicative of UGT2B7/2B4 activity) dropped by 35% (CP A), 51% (CP B), and 64% (CP C). These predicted MRs were corroborated by the observed data. This work thus increases confidence in Simcyp® predictions of the effect of liver cirrhosis on the pharmacokinetics of UGT1A4 and UGT2B7/UGT2B4 substrates. Significance Statement This paper presents a PBPK model for midazolam and its metabolites and verifies the accurate simulation of pharmacokinetic profiles when using the Simcyp® hepatic impairment population models. Exposure changes of midazolam-N-glucuronide and 1'-OH-midazolam-O-glucuronide reflect the impact of decreases in UGT1A4 and UGT2B7/2B4 glucuronidation activity in cirrhotic patients. The approach used in this study may be extended to verify the modeling of other UGT enzymes affected by liver cirrhosis.

Determining the exposure of maternal medicines through breastfeeding: the UmbrelLACT study protocol-a contribution from the ConcePTION project.

INTRODUCTION: Breastfeeding is beneficial for the health of the mother and child. However, at least 50% of postpartum women need pharmacotherapy, and this number is rising due to the increasing prevalence of chronic diseases and pregnancies at a later age. Making informed decisions on medicine use while breastfeeding is often challenging, considering the extensive information gap on medicine exposure and safety during lactation. This can result in the unnecessary cessation of breastfeeding, the avoidance of pharmacotherapy or the off-label use of medicines. The UmbrelLACT study aims to collect data on human milk transfer of maternal medicines, child exposure and general health outcomes. Additionally, the predictive performance of lactation and paediatric physiologically based pharmacokinetic (PBPK) models, a promising tool to predict medicine exposure in special populations, will be evaluated. METHODS AND ANALYSIS: Each year, we expect to recruit 5-15 breastfeeding mothers using pharmacotherapy via the University Hospitals Leuven, the BELpREG project (pregnancy registry in Belgium) or external health facilities. Each request and compound will be evaluated on relevance (ie, added value to available scientific evidence) and feasibility (including access to analytical assays). Participants will be requested to complete at least one questionnaire on maternal and child's general health and collect human milk samples over 24 hours. Optionally, two maternal and one child's blood samples can be collected. The maternal medicine concentration in human milk will be determined along with the estimation of the medicine intake (eg, daily infant dose and relative infant dose) and systemic exposure of the breastfed child. The predictive performance of PBPK models will be assessed by comparing the observed concentrations in human milk and plasma to the PBPK predictions. ETHICS AND DISSEMINATION: This study has been approved by the Ethics Committee Research UZ/KU Leuven (internal study number S67204). Results will be published in peer-reviewed journals and presented at (inter)national scientific meetings. TRIAL REGISTRATION NUMBER: NCT06042803.

Second generation Al18F-labeled D-amino acid peptide for CXCR4 targeted molecular imaging.

BACKGROUND: The C-X-C chemokine receptor type 4 (CXCR4) is overexpressed in many cancers, e.g. multiple myeloma and acute leukemia, yet solely [68Ga]PentixaFor is used for clinical PET imaging. The aim of this study was to develop and assess a second generation Al18F-labeled D-amino acid peptide based on the viral macrophage inflammatory protein II for CXCR4 targeted molecular imaging. METHODS: We designed a library of monomer and multimer constructs and evaluated their binding affinity for human and mouse CXCR4. Based on these results, we selected the best vector molecule for development of an Al18F-labeled ligand, [18F]AlF-NOTA-2xDV1(c11sc12s), which was further evaluated in a cell-based binding assay to assess its binding properties and specificity for CXCR4. Next, pharmacokinetics and tumor uptake of [18F]AlF-NOTA-2xDV1(c11sc12s) were evaluated in naïve mice and mice with xenografts derived from U87.CXCR4 cells. Finally, we performed an imaging study in a non-human primate to assess the in vivo distribution of this novel radioligand in a species closely related to humans. RESULTS: The lead ligand AlF-NOTA-2xDV1(c11sc12s) showed six-fold higher affinity for human CXCR4 compared to Ga-Pentixafor. The corresponding radiotracer was obtained in a good radiochemical yield of 40.1 ± 13.5 % (n = 4) and apparent molar activity of 20.4 ± 3.3 MBq/nmol (n = 4) after optimization. In U87.CD4.CXCR4 cell binding assays, the total bound fraction of [18F]AlF-NOTA-(2×)DV1(c11sc12s) was 32.4 ± 1.8 %. This fraction could be reduced by 82.5 % in the presence of 75 µM AMD3100. In naïve mice, [18F]AlF-NOTA-2xDV1(c11sc12s) accumulated in organs expressing mouse CXCR4, e.g. the liver (SUVmean (mean standardized uptake value) 75 min p.i. 11.7 ± 0.6), which was blockable by co-injecting AMD3100 (5 mg/kg). In U87.CXCR4 xenografted tumor mice, the tumor uptake of [18F]AlF-NOTA-2xDV1(c11sc12s) remained low (SUVmean 0.5 ± 0.1), but was reduced by co-administration of AMD3100. Surprisingly, [18F]AlF-NOTA-2xDV1(c11sc12s) exhibited a similar biodistribution in a non-human primate as in mice indicating off-target binding of [18F]AlF-NOTA-2xDV1(c11sc12s) in liver tissue. We confirmed that [18F]AlF-NOTA-2xDV1(c11sc12s) is taken up by hepatocytes using in vitro studies and that the uptake can be blocked with AMD3100 and rifampicin, a potent organic anion-transporting-polypeptide (OATP)1B1 and OATP1B3 inhibitor. CONCLUSION: The second generation D-peptide AlF-NOTA-2xDV1(c11sc12s) showed high affinity for human CXCR4 and the corresponding radiotracer was produced in good radiochemical yields. However, [18F]AlF-NOTA-2xDV1(c11sc12s) is not specific for CXCR4 and is also a substrate for OATP1B1 and/or OATP1B3, known to mediate hepatic uptake. Therefore, D-amino acid peptides, based on the viral macrophage inflammatory protein II, are not the prefered vector molecule for the development of CXCR4 targeting molecular imaging tools.

Interdisciplinary Collaboration on Real World Data to Close the Knowledge Gap: A Reflection on "De Sutter et al. Predicting Volume of Distribution in Neonates: Performance of Physiologically Based Pharmacokinetic Modelling".

This commentary further reflects on the paper of De Sutter et al. on predicting volume of distribution in neonates, and the performance of physiologically based pharmacokinetic models We hereby stressed the add on value to collaborate on real world data to further close this knowledge gap. We illustrated this by weight distribution characteristics in breastfed (physiology) and in asphyxiated (pathophysiology), with additional reflection on their kidney and liver function.

A Population Model of Time-Dependent Changes in Serum Creatinine in (Near)term Neonates with Hypoxic-Ischemic Encephalopathy During and After Therapeutic Hypothermia.

The objective was to apply a population model to describe the time course and variability of serum creatinine (sCr) in (near)term neonates with moderate to severe encephalopathy during and after therapeutic hypothermia (TH). The data consisted of sCr observations up to 10 days of postnatal age in neonates who underwent TH during the first 3 days after birth. Available covariates were birth weight (BWT), gestational age (GA), survival, and acute kidney injury (AKI). A previously published population model of sCr kinetics in neonates served as the base model. This model predicted not only sCr but also the glomerular filtration rate normalized by its value at birth (GFR/GFR0). The model was used to compare the TH neonates with a reference full term non-asphyxiated population of neonates. The estimates of the model parameters had good precision and showed high between subject variability. AKI influenced most of the estimated parameters denoting a strong impact on sCr kinetics and GFR. BWT and GA were not significant covariates. TH transiently increased [Formula: see text] in TH neonates over the first days compared to the reference group. Asphyxia impacted not only GFR, but also the [Formula: see text] synthesis rate. We also observed that AKI neonates exhibit a delayed onset of postnatal GFR increase and have a higher [Formula: see text] synthesis rate compared to no-AKI patients. Our findings show that the use of [Formula: see text] as marker of renal function in asphyxiated neonates treated with TH to guide dose selection for renally cleared drugs is challenging, while we captured the postnatal sCr patterns in this specific population.

Challenges Related to Acquisition of Physiological Data for Physiologically Based Pharmacokinetic (PBPK) Models in Postpartum, Lactating Women and Breastfed Infants-A Contribution from the ConcePTION Project.

Physiologically based pharmacokinetic (PBPK) modelling is a bottom-up approach to predict pharmacokinetics in specific populations based on population-specific and medicine-specific data. Using an illustrative approach, this review aims to highlight the challenges of incorporating physiological data to develop postpartum, lactating women and breastfed infant PBPK models. For instance, most women retain pregnancy weight during the postpartum period, especially after excessive gestational weight gain, while breastfeeding might be associated with lower postpartum weight retention and long-term weight control. Based on a structured search, an equation for human milk intake reported the maximum intake of 153 mL/kg/day in exclusively breastfed infants at 20 days, which correlates with a high risk for medicine reactions at 2-4 weeks in breastfed infants. Furthermore, the changing composition of human milk and its enzymatic activities could affect pharmacokinetics in breastfed infants. Growth in breastfed infants is slower and gastric emptying faster than in formula-fed infants, while a slower maturation of specific metabolizing enzymes in breastfed infants has been described. The currently available PBPK models for these populations lack structured systematic acquisition of population-specific data. Future directions include systematic searches to fully identify physiological data. Following data integration as mathematical equations, this holds the promise to improve postpartum, lactation and infant PBPK models.

PBPK Modelling for Drugs Cleared by Non-CYP Enzymes: State-of-the-Art and Future Perspectives.

Physiologically-based pharmacokinetic (PBPK) modeling has become the established method for predicting human pharmacokinetics (PK) and drug-drug interactions (DDI). The number of drugs cleared by non-CYP enzyme metabolism has increased steadily and to date, there is no consolidated overview of PBPK modeling for drugs cleared by non-CYP enzymes. This review aims to describe the state-of-the-art for PBPK modeling for drugs cleared via non-CYP enzymes, to identify successful strategies, to describe gaps and to provide suggestion to overcome them. To this end, we conducted a detailed literature search and found 58 articles published before the 1st of January 2023 containing 95 examples of clinical PBPK models for 62 non-CYP enzyme substrates. Reviewed articles covered the drug clearance by uridine 5'-diphospho-glucuronosyltransferases (UGTs), aldehyde oxidase (AO), flavin-containing monooxygenases (FMOs), sulfotransferases (SULTs) and carboxylesterases (CES), with UGT2B7, UGT1A9, CES1, FMO3 and AO being the enzymes most frequently involved. In vitro-in vivo extrapolation (IVIVE) of intrinsic clearance and the bottom-up PBPK modeling involving non-CYP enzymes remains challenging. We observed that the middle-out modeling approach was applied in 80% of the cases, with metabolism parameters optimized in 73% of the models. Our review could not identify a standardized approach used for model optimization based on clinical data, with manual optimization employed most frequently. Successful development of models for UGT2B7, UGT1A9, CES1, and FMO3 substrates provides a foundation for other drugs metabolized by these enzymes and guides the way forward in creating PBPK models for other enzymes in these families. Significance Statement Our review charts the rise of PBPK modeling for drugs cleared by non-CYP enzymes. Analyzing 58 articles and 62 non-CYP enzyme substrates, we found that UGTs, AO, FMOs, SULTs, and CES were the main enzyme families involved and that UGT2B7, UGT1A9, CES1, FMO3 and AO are the individual enzymes with the strongest PBPK modeling precedents. Approaches established for these enzymes can now be extended to additional substrates and to drugs metabolized by enzymes that are similarly well characterized.

Automated Generation of hiPSC-Derived Hepatic Progeny by Cost-Efficient Compounds.

Human pluripotent stem cell (hPSC)-derived hepatocyte-like cells (HLCs) hold great promise for liver disease modeling, drug discovery, and drug toxicity screens. Yet, several hurdles still need to be overcome, including among others decrease in the cost of goods to generate HLCs and automation of the differentiation process. We here describe that the use of an automated liquid handling system results in highly reproducible HLC differentiation from hPSCs. This enabled us to screen 92 chemicals to replace expensive growth factors at each step of the differentiation protocol to reduce the cost of goods of the differentiation protocol by approximately 79%. In addition, we also evaluated several recombinant extracellular matrices to replace Matrigel. We demonstrated that differentiation of hPSCs on Laminin-521 using an optimized small molecule combination resulted in HLCs that were transcriptionally identical to HLCs generated using the growth factor combinations. In addition, the HLCs created using the optimized small molecule combination secreted similar amounts of albumin and urea, and relatively low concentrations of alfa-fetoprotein, displayed similar CYP3A4 functionality, and a similar drug toxicity susceptibility as HLCs generated with growth factor cocktails. The broad applicability of the new differentiation protocol was demonstrated for 4 different hPSC lines. This allowed the creation of a scalable, xeno-free, and cost-efficient hPSC-derived HLC culture, suitable for high throughput disease modeling and drug screenings, or even for the creation of HLCs for regenerative therapies.

Pharmacokinetics during therapeutic hypothermia in neonates: from pathophysiology to translational knowledge and physiologically-based pharmacokinetic (PBPK) modeling.

INTRODUCTION: Perinatal asphyxia (PA) still causes significant morbidity and mortality. Therapeutic hypothermia (TH) is the only effective therapy for neonates with moderate to severe hypoxic-ischemic encephalopathy after PA. These neonates need additional pharmacotherapy, and both PA and TH may impact physiology and, consequently, pharmacokinetics (PK) and pharmacodynamics (PD). AREAS COVERED: This review provides an overview of the available knowledge in PubMed (until November 2022) on the pathophysiology of neonates with PA/TH. In vivo pig models for this setting enable distinguishing the effect of PA versus TH on PK and translating this effect to human neonates. Available asphyxia pig models and methodological considerations are described. A summary of human neonatal PK of supportive pharmacotherapy to improve neurodevelopmental outcomes is provided. EXPERT OPINION: To support drug development for this population, knowledge from clinical observations (PK data, real-world data on physiology), preclinical (in vitro and in vivo (minipig)) data, and molecular and cellular biology insights can be integrated into a predictive physiologically-based PK (PBPK) framework, as illustrated by the I-PREDICT project (Innovative physiology-based pharmacokinetic model to predict drug exposure in neonates undergoing cooling therapy). Current knowledge, challenges, and expert opinion on the future directions of this research topic are provided.

Safety of medicines during breastfeeding - from case report to modeling: a contribution from the ConcePTION project.

INTRODUCTION: Despite many research efforts, current data on the safety of medicines during breastfeeding are either fragmented or lacking, resulting in restrictive labeling of most medicines. In the absence of pharmacoepidemiologic safety studies, risk estimation for breastfed infants is mainly derived from pharmacokinetic (PK) information on medicine. This manuscript provides a description and a comparison of the different methodological approaches that can yield reliable information on medicine transfer into human milk and the resulting infant exposure. AREA COVERED: Currently, most information on medicine transfer in human milk relies on case reports or traditional PK studies, which generate data that can hardly be generalized to the population. Some methodological approaches, such as population PK (popPK) and physiologically based PK (PBPK) modeling, can be used to provide a more complete characterization of infant medicine exposure through human milk and simulate the most extreme situations while decreasing the burden of sampling in breastfeeding women. EXPERT OPINION: PBPK and popPK modeling are promising approaches to fill the gap in knowledge of medicine safety in breastfeeding, as illustrated with our escitalopram example.

Generic Workflow to Predict Medicine Concentrations in Human Milk Using Physiologically-Based Pharmacokinetic (PBPK) Modelling-A Contribution from the ConcePTION Project.

Women commonly take medication during lactation. Currently, there is little information about the exposure-related safety of maternal medicines for breastfed infants. The aim was to explore the performance of a generic physiologically-based pharmacokinetic (PBPK) model to predict concentrations in human milk for ten physiochemically diverse medicines. First, PBPK models were developed for "non-lactating" adult individuals in PK-Sim/MoBi v9.1 (Open Systems Pharmacology). The PBPK models predicted the area-under-the-curve (AUC) and maximum concentrations (Cmax) in plasma within a two-fold error. Next, the PBPK models were extended to include lactation physiology. Plasma and human milk concentrations were simulated for a three-months postpartum population, and the corresponding AUC-based milk-to-plasma (M/P) ratios and relative infant doses were calculated. The lactation PBPK models resulted in reasonable predictions for eight medicines, while an overprediction of human milk concentrations and M/P ratios (>2-fold) was observed for two medicines. From a safety perspective, none of the models resulted in underpredictions of observed human milk concentrations. The present effort resulted in a generic workflow to predict medicine concentrations in human milk. This generic PBPK model represents an important step towards an evidence-based safety assessment of maternal medication during lactation, applicable in an early drug development stage.

Unravelling the Hepatic Elimination Mechanisms of Colistin.

PURPOSE: Colistin is an antibiotic which is increasingly used as a last-resort therapy in critically-ill patients with multidrug resistant Gram-negative infections. The purpose of this study was to evaluate the mechanisms underlying colistin's pharmacokinetic (PK) behavior and to characterize its hepatic metabolism. METHODS: In vitro incubations were performed using colistin sulfate with rat liver microsomes (RLM) and with rat and human hepatocytes (RH and HH) in suspension. The uptake of colistin in RH/HH and thefraction of unbound colistin in HH (fu,hep) was determined. In vitro to in vivo extrapolation (IVIVE) was employed to predict the hepatic clearance (CLh) of colistin. RESULTS: Slow metabolism was detected in RH/HH, with intrinsic clearance (CLint) values of 9.34± 0.50 and 3.25 ± 0.27 mL/min/kg, respectively. Assuming the well-stirred model for hepatic drug elimination, the predicted rat CLh was 3.64± 0.22 mL/min/kg which could explain almost 70% of the reported non-renal in vivo clearance. The predicted human CLh was 91.5 ± 8.83 mL/min, which was within two-fold of the reported plasma clearance in healthy volunteers. When colistin was incubated together with the multidrug resistance-associated protein (MRP/Mrp) inhibitor benzbromarone, the intracellular accumulation of colistin in RH/HH increased significantly. CONCLUSION: These findings indicate the major role of hepatic metabolism in the non-renal clearance of colistin, while MRP/Mrp-mediated efflux is involved in the hepatic disposition of colistin. Our data provide detailed quantitative insights into the hereto unknown mechanisms responsible for non-renal elimination of colistin.

Development of a neonatal Göttingen Minipig model for dose precision in perinatal asphyxia: technical opportunities, challenges, and potential further steps.

Animal models provide useful information on mechanisms in human disease conditions, but also on exploring (patho)physiological factors affecting pharmacokinetics, safety, and efficacy of drugs in development. Also, in pediatric patients, nonclinical data can be critical for better understanding the disease conditions and developing new drug therapies in this age category. For perinatal asphyxia (PA), a condition defined by oxygen deprivation in the perinatal period and possibly resulting in hypoxic ischemic encephalopathy (HIE) or even death, therapeutic hypothermia (TH) together with symptomatic drug therapy, is the standard approach to reduce death and permanent brain damage in these patients. The impact of the systemic hypoxia during PA and/or TH on drug disposition is largely unknown and an animal model can provide useful information on these covariates that cannot be assessed separately in patients. The conventional pig is proven to be a good translational model for PA, but pharmaceutical companies do not use it to develop new drug therapies. As the Göttingen Minipig is the commonly used pig strain in nonclinical drug development, the aim of this project was to develop this animal model for dose precision in PA. This experiment consisted of the instrumentation of 24 healthy male Göttingen Minipigs, within 24 h of partus, weighing approximately 600 g, to allow the mechanical ventilation and the multiple vascular catheters inserted for maintenance infusion, drug administration and blood sampling. After premedication and induction of anesthesia, an experimental protocol of hypoxia was performed, by decreasing the inspiratory oxygen fraction (FiO2) at 15%, using nitrogen gas. Blood gas analysis was used as an essential tool to evaluate oxygenation and to determine the duration of the systemic hypoxic insult to approximately 1 h. The human clinical situation was mimicked for the first 24 h after birth in case of PA, by administering four compounds (midazolam, phenobarbital, topiramate and fentanyl), frequently used in a neonatal intensive care unit (NICU). This project aimed to develop the first neonatal Göttingen Minipig model for dose precision in PA, allowing to separately study the effect of systemic hypoxia versus TH on drug disposition. Furthermore, this study showed that several techniques that were thought to be challenging or even impossible in these very small animals, such as endotracheal intubation and catheterization of several veins, are feasible by trained personnel. This is relevant information for laboratories using the neonatal Göttingen Minipig for other disease conditions or drug safety testing.

Physiologically based pharmacokinetic modelling to inform combination dosing regimens of ceftaroline and daptomycin in special populations.

AIMS: The combination of daptomycin and ceftaroline used as salvage therapy is associated with higher survival and decreased clinical failure in complicated methicillin-resistant Staphylococcus aureus (MRSA) infections that are resistant to standard MRSA treatment. This study aimed to evaluate dosing regimens for coadministration of daptomycin and ceftaroline in special populations including paediatrics, renally impaired (RI), obese and geriatrics that generate sufficient coverage against daptomycin-resistant MRSA. METHODS: Physiologically based pharmacokinetic models were developed from pharmacokinetic studies of healthy adults, geriatric, paediatric, obese and RI patients. The predicted profiles were used to evaluate joint probability of target attainment (PTA), as well as tissue-to-plasma ratios. RESULTS: The adult dosing regimens of 6 mg/kg every (q)24h or q48h daptomycin and 300-600 mg q12h ceftaroline fosamil by RI categories achieved ≥90% joint PTA when the minimum inhibitory concentrations in the combination are at or below 1 and 4 µg/mL against MRSA. In paediatrics, wherein there is no recommended daptomycin dosing regimen for S. aureus bacteraemia, ≥90% joint PTA is achieved when the minimum inhibitory concentrations in the combination are up to 0.5 and 2 µg/mL for standard paediatric dosing regimens of 7 mg/kg q24h daptomycin and 12 mg/kg q8h ceftaroline fosamil. Model predicted tissue-to-plasma ratios of 0.3 and 0.7 in the skin and lung, respectively, for ceftaroline and 0.8 in the skin for daptomycin. CONCLUSION: Our work illustrates how physiologically based pharmacokinetic modelling can inform appropriate dosing of adult and paediatric patients and thereby enable prediction of target attainment in the patients during multitherapies.

Dose Adjustment in Patients with Liver Cirrhosis - Comparison of Two Different Modeling Approaches.

Failure to perform adequate dose adjustment in patients with liver cirrhosis may be associated with increased toxicity. We compared the prediction of area under the curve (AUC) and clearance for the six compounds of the Basel phenotyping cocktail (caffeine, efavirenz, flurbiprofen, omeprazole, metoprolol, and midazolam) using a well-known physiology-based pharmacokinetic approach (physiologically-based pharmacokinetic [PBPK] approach, Simcyp) and a novel top-down method based on the systemic clearance in healthy volunteers adjusted for markers of liver and renal dysfunction ("top-down approach"). With few exceptions, plasma concentration-time curves were accurately predicted by the PBPK approach. In comparison to the measured AUC and clearance of these drugs in patients with liver cirrhosis and healthy controls, except for efavirenz, the estimates of both approaches were within two standard deviations of the mean for total and free drug concentrations. For both approaches, a correction factor for dose adjustment in patients with liver cirrhosis could be calculated for the drugs administered. AUCs calculated using the adjusted doses were comparable to the AUCs measured in control subjects, with slightly more accurate predictions generated by the PBPK approach. For drugs with a free fraction < 50%, predictions using free drug concentrations were more accurate than with total drug concentrations. In conclusion, both methods provided good qualitative predictions of the changes by liver cirrhosis in the pharmacokinetics of the six compounds investigated. The top-down approach is easier to implement but the PBPK approach predicted changes in drug exposure more accurately than the top-down approach and provided reliable estimates for plasma concentrations.

PBPK-based dose finding for sildenafil in pregnant women for antenatal treatment of congenital diaphragmatic hernia.

Sildenafil is a potent vasodilator and phosphodiesterase type five inhibitor, commercially known as Revatio® and approved for the treatment of pulmonary arterial hypertension. Maternal administration of sildenafil during pregnancy is being evaluated for antenatal treatment of several conditions, including the prevention of pulmonary hypertension in fetuses with congenital diaphragmatic hernia. However, determination of a safe and effective maternal dose to achieve adequate fetal exposure to sildenafil remains challenging, as pregnancy almost always is an exclusion criterion in clinical studies. Physiologically-based pharmacokinetic (PBPK) modelling offers an attractive approach for dose finding in this specific population. The aim of this study is to exploit physiologically-based pharmacokinetic modelling to predict the required maternal dose to achieve therapeutic fetal exposure for the treatment congenital diaphragmatic hernia. A full-PBPK model was developed for sildenafil and N-desmethyl-sildenafil using the Simcyp simulator V21 platform, and verified in adult reference individuals, as well as in pregnant women, taking into account maternal and fetal physiology, along with factors known to determine hepatic disposition of sildenafil. Clinical pharmacokinetic data in mother and fetus were previously obtained in the RIDSTRESS study and were used for model verification purposes. Subsequent simulations were performed relying either on measured values for fetal fraction unbound (fu = 0.108) or on values predicted by the simulator (fu = 0.044). Adequate doses were predicted according to the efficacy target of 15 ng/mL (or 38 ng/mL) and safety target of 166 ng/mL (or 409 ng/mL), assuming measured (or predicted) fu values, respectively. Considering simulated median profiles for average steady state sildenafil concentrations, dosing regimens of 130 mg/day or 150 mg/day (administered as t.i.d.), were within the therapeutic window, assuming either measured or predicted fu values, respectively. For safety reasons, dosing should be initiated at 130 mg/day, under therapeutic drug monitoring. Additional experimental measurements should be performed to confirm accurate fetal (and maternal) values for fu. Additional characterization of pharmacodynamics in this specific population is required and may lead to further optimization of the dosing regimen.

Sensitive and rapid method for the quantitation of amoxicillin in minipig plasma and milk by LC-MS/MS: A contribution from the IMI ConcePTION project.

The IMI project ConcePTION was launched to fill the knowledge gap of using medicines during pregnancy and lactation. To achieve this goal, several studies are being conducted, including the bioanalysis of amoxicillin in minipig plasma and milk. A high-throughput, robust and reliable liquid chromatography tandem mass spectrometry method was developed and validated according to FDA and EMA guidelines to determine the concentrations of amoxicillin in a large number of minipig plasma and milk samples. Chromatographic separation was achieved on a Luna® Omega Polar C18, 1.6 µm, 100 × 2.1 mm column, with a mobile phase consisting of 0.1% formic acid in water and acetonitrile. Mass spectrometry used in a positive ionization mode and the transitions m/z 366.1 â†’ 349.2 was selected to monitor amoxicillin, while m/z 370.1 â†’ 114.15 was selected for the stable isotope labelled internal standard. This method features a linear quantification range of 10 ng/mL - 10 µg/mL, recovery of not less than 94.1%, a single sample extraction method for both plasma and milk matrices, and an analysis runtime of 5 min.

Case Report: Bosentan and Sildenafil Exposure in Human Milk - A Contribution From the ConcePTION Project.

Introduction: Quantitative information on disposition of maternal medicines in human milk remains a major knowledge gap. This case report presents the clinical and pharmacokinetic data of a single mother-infant pair exposed to bosentan and sildenafil for the treatment of pulmonary arterial hypertension (PAH) during lactation. Case presentation: A 43-year old mother was treated with sildenafil (20 mg, 3x/day) and bosentan (125 mg, 2x/day) for PAH. Her 21-months old infant received breastfeeding in combination with adequate complementary foods. Milk samples were collected over 24 h, at day 637 and 651 after delivery. The observed average steady-state concentrations of sildenafil (2.84 µg/L) and bosentan (49.0 µg/L) in human milk were low. The Daily Infant Dosage ingested by the nursing infant through human milk was 0.02 µg/kg/day for sildenafil and 0.29 µg/kg/day for bosentan at day 637, and 0.03 µg/kg/day and 0.60 µg/kg/day at day 651. The Relative Infant Dose calculated for an exclusively breastfed infant with an estimated milk intake of 150 ml/kg/day, was 0.06% for sildenafil and 0.24% for bosentan. General health outcome of the infant, reported by the mother, was uneventful until the sampling days. Conclusion: Low medicine concentrations were found in human milk expressed 21 months after delivery after maternal intake of 20 mg sildenafil three times daily and 125 mg bosentan twice daily. General health of the nursing infant until sampling was reported as optimal by the mother.