A high-throughput liquid chromatography-tandem mass spectrometry assay for the simultaneous quantification of p-cresol sulfate, p-cresol glucuronide, indoxyl sulfate, and indoxyl glucuronide in HepaRG culture medium and the demonstration of mefenamic acid as a potent and selective detoxifying agent.

BACKGROUND: p-cresol and indole are uremic compounds which undergo sulfonation to generate the highly toxic p-cresol sulfate (pCS) and indoxyl sulfate (IxS). They are also subjected to glucuronidation to produce the less toxic p-cresol glucuronide (pCG) and indoxyl glucuronide (IG). We developed and validated an assay to quantify these metabolites in HepaRG cells. We also tested the effects of mefenamic acid on their in-situ formations in relation to the development of cellular necrosis. RESEARCH DESIGN AND METHODS: HepaRG cells were exposed to p-cresol or indole (0-1 mM) with mefenamic acid (0-3000 nM) for 24 hours to generate uremic metabolites. Cells were also exposed to 0.5 mM p-cresol or indole with/without 30 nM mefenamic acid to characterize lactate dehydrogenase (LDH) release. RESULTS: The assay exhibited high sensitivity and wide calibration ranges covering human concentrations. HepaRG cells also generated physiologically-relevant concentrations of each metabolite. Mefenamic acid inhibited pCS formation in a concentration-dependent manner without affecting pCG, IxS, or IG. Mefenamic acid also reduced LDH release from p-cresol (by 50.12±5.86%) or indole (56.26±3.58%). CONCLUSIONS: This novel assay is capable of quantifying these metabolites in HepaRG cells. Our novel findings suggest that mefenamic acid can be potentially utilized therapeutically to attenuate pCS-associated toxicities.

Women's perceptions and experiences with cannabis use in menopause: a qualitative study.

OBJECTIVES: Since the legalization of recreational cannabis in 2018, the use of cannabis for medical reasons has increased in Canada. The aim of this study was to explore the experiences and perceptions of midlife women using cannabis for medical purposes coinciding with menopause symptom management. METHODS: Semistructured, one-on-one interviews were conducted using a qualitative description method. This was the second phase of a mixed methods study, where interviewees were purposefully selected from a sample of women (ages 35 and over, located in Alberta) surveyed during the first phase of the study. Interviews were by phone or virtual meeting, audio-recorded, and transcribed verbatim. Qualitative content analysis was applied to analyze the data collected. RESULTS: Twelve interviews were conducted between December 2020 and April 2021. Menopause was perceived as a complex experience for women. Cannabis was described as a therapeutic agent, providing symptom relief through the menopause transition. Women reported similarities in their menopause and cannabis use experiences in the lack of information available, limited role of healthcare providers, feelings of stigmatization, and emphasis on self-education. Women self-managed their cannabis use, learning from their own experiences or the anecdotal sharing of others', accessed cannabis from a variety of medical and nonmedical sources, and relied on experimentation, and a range of supports were described. CONCLUSION: Midlife women pursued the use of cannabis medically to manage symptoms that overlap with menopause. Understanding how and why midlife women use cannabis medically can provide insight for future research and the development of educational resources to support women in menopause.

Liquid Chromatography-Mass Spectrometry Analytical Methods for the Quantitation of p-Cresol Sulfate and Indoxyl Sulfate in Human Matrices: Biological Applications and Diagnostic Potentials.

Indoxyl sulfate (IxS) and p-cresyl sulfate (pCS) are toxic uremic compounds with documented pathological outcomes. This review critically and comprehensively analyzes the available liquid chromatography-mass spectrometry methods quantifying IxS and pCS in human matrices and the biological applications of these validated assays. Embase, Medline, PubMed, Scopus, and Web of Science were searched until December 2023 to identify assays with complete analytical and validation data (N = 23). Subsequently, citation analysis with PubMed and Scopus was utilized to identify the biological applications for these assays (N = 45). The extraction methods, mobile phase compositions, chromatography, and ionization methods were evaluated with respect to overall assay performance (e.g., sensitivity, separation, interference). Most of the assays focused on human serum/plasma, utilizing acetonitrile or methanol (with ammonium acetate/formate or formic/acetic acid), liquid-liquid extraction, reverse phase (e.g., C18) chromatography, and gradient elution for analyte separation. Mass spectrometry conditions were also consistent in the identified papers, with negative electrospray ionization, select multiple reaction monitoring transitions and deuterated internal standards being the most common approaches. The validated biological applications indicated IxS and/or pCS were correlated with renal disease progression and cardiovascular outcomes, with limited data on central nervous system disorders. Methods for reducing IxS and/or pCS concentrations were also identified (e.g., drugs, natural products, diet, dialysis, transplantation) where inconsistent findings have been reported. The clinical monitoring of IxS and pCS is gaining significant interest, and this review will serve as a useful compendium for scientists and clinicians.

Bioavailability and Metabolism of Bioactive Peptide IRW with Angiotensin-Converting Enzyme 2 (ACE2) Upregulatory Activity in Spontaneously Hypertensive Rats.

Peptide IRW is the first food-derived angiotensin-converting enzyme 2 (ACE2) upregulator. This study aimed to investigate the pharmacokinetic characteristics of IRW and identify the metabolites contributing to its antihypertensive activity in spontaneously hypertensive rats (SHRs). Rats were administered 100 mg of IRW/kg of the body weight via an intragastric or intravenous route. The bioavailability (F %) was determined to be 11.7%, and the half-lives were 7.9 ± 0.5 and 28.5 ± 6.8 min for gavage and injection, respectively. Interestingly, significant blood pressure reduction was not observed until 1.5 h post oral administration, or 2 h post injection, indicating that the peptide's metabolites are likely responsible for the blood pressure-lowering activity. Time-course metabolomics revealed a significant increase in the level of kynurenine, a tryptophan metabolite, in blood after IRW administration. Kynurenine increased the level of ACE2 in cells. Oral administration of tryptophan (W), but not dipeptide IR, lowered the blood pressure and upregulated aortic ACE2 in SHRs. Our study supports the key role of tryptophan and its metabolite, kynurenine, in IRW's blood pressure-lowering effects.

Pharmacokinetics of Long-Acting Methylphenidate: Formulation Differences, Bioequivalence, Interchangeability.

BACKGROUND AND OBJECTIVE: Attention deficit hyperactivity disorder is one of the most common neuropsychiatric conditions in children, and methylphenidate (MPH) is one of the first-line therapies. MPH is available in a variety of extended-release (ER) formulations worldwide, and most formulations are not considered bioequivalent due to differences in pharmacokinetics. It is hypothesized that the current bioequivalence guidelines from the different regulatory bodies may generate inconsistent findings or recommendations when assessing the bioequivalence of ER MPH formulations. This manuscript aims to conduct a comprehensive and narrative critical literature review to analyze pharmacokinetic data pertaining to ER formulations of MPH in order to assess bioequivalence, differences in regulatory guidelines, and additional pharmacokinetic-pharmacodynamic parameters that may help define interchangeability. METHODS: A literature search was conducted in EMBASE, Medline, and Cochrane Library with no time limits. Study characteristics, non-compartmental pharmacokinetic parameters, and bioequivalence data were extracted for analysis. RESULTS: Thirty-three studies were identified with primary pharmacokinetic data after the administration of ER MPH, of which 10 were direct comparative studies (i.e., at least 2 formulations tested within a single setting) and 23 were indirect comparisons (i.e., different experimental settings). Two formulations were consistently reported as bioequivalent across the regulatory bodies using criteria from their guidance documents, although inconsistencies have been observed. However, when additional kinetic criteria (discussed in this manuscript) were imposed, only one study met the more stringent definition of bioequivalence. Various clinical factors also had inconsistent effects on the pharmacokinetics and interchangeability of the different formulations, which were associated with a lack of standardization for assessing covariates across the regulatory agencies. CONCLUSION: Additional pharmacokinetic parameters and consistency in guidelines across the regulatory bodies may improve bioequivalence assessments. Based on our findings, more research is also required to understand whether bioequivalence is an appropriate measure for determining MPH interchangeability. This critical review is suitable for formulation scientists, clinical pharmacologists, and clinicians.

Significant Effects of Renal Function on Mycophenolic Acid Total Clearance in Pediatric Kidney Transplant Recipients with Population Pharmacokinetic Modeling.

BACKGROUND AND OBJECTIVES: Mycophenolic acid (MPA) is an immunosuppressant commonly prescribed in pediatric kidney transplantation to prevent graft rejection. Large variabilities in MPA plasma exposures have been observed in this population, which could result in severe adverse effects. The majority of the MPA pharmacokinetic data have been reported in adult populations, whereas information in pediatric patients is still very limited. The objective of this study was to establish a novel, nonlinear mixed-effects model for MPA and investigate the clinical variables affecting MPA population pharmacokinetics in pediatric kidney transplant recipients. METHODS: Data were collected retrospectively from pediatric kidney transplant patients (≤ 18 years when MPA concentrations were initially collected; on oral administration of mycophenolate mofetil) in Calgary, Alberta, Canada. Nonlinear mixed-effect modeling was conducted using stochastic approximation expectation-maximization in Monolix 2021R2 (Lixoft SAS, France) to determine population pharmacokinetic estimates, interindividual variabilities, and interoccasional variabilities. Covariate models were constructed using the Model Proposal function in Monolix in conjunction with a systematic stepwise inclusion/elimination protocol. The best model was selected based on objective function values, relative standard errors, goodness-of-fit plots, prediction-corrected visual predictive checks, and numerical predictive checks. RESULTS: A total of 50 pediatric kidney transplant patients (25 female) with 219 MPA plasma concentration-time profiles were included. The average age (± standard deviation) and posttransplant time for the sample population were 12.8 ± 4.8 years and 762 ± 1160 days, respectively. The majority of study subjects (i.e., > 85% based on all occasions) were co-administered tacrolimus. A two-compartment, first-order absorption with lag time and linear elimination structural model with lognormal distributed proportional residual errors best described the MPA concentration-time data. The absorption rate constant (2.52 h-1 or 0.042 min-1), lag time (0.166 h or 9.96 min), volumes of distributions of the central (22.8 L) and peripheral (216 L) compartments, and intercompartment clearance (17.6 L h-1 or 0.293 L min-1) were consistent with literature values; whereas total MPA clearance (0.72 L h-1 or 0.012 L min-1) was relatively reduced, likely due to the general lack of cyclosporine interactions and the stabilized graft functions from significantly longer posttransplant time in our sample population. Of the clinical variables tested, only estimated glomerular filtration rate (eGFR) was identified a significant covariate affecting total MPA clearance with a positive, exponential relationship. The final population pharmacokinetic model was successfully evaluated/validated using a variety of complementary methods. CONCLUSION: We have successfully constructed and validated a novel population pharmacokinetic model of MPA in pediatric kidney transplant patients. A positive, nonlinear relationship between eGFR and total MPA clearance identified in our model is likely attributed to multiple concurrent mechanisms, which warrant further systematic investigations.

Cannabis use for menopause in women aged 35 and over: a cross-sectional survey on usage patterns and perceptions in Alberta, Canada.

OBJECTIVE: Use of cannabis for medical reasons has increased in Canada since legalisation of recreational cannabis in 2018. The objective of this study was to examine the pattern of use and perceptions about cannabis for menopause in women aged 35 and over in Alberta, Canada. DESIGN: Cross-sectional, web-based survey. SETTING: Online (location of participant residence in Alberta, Canada). PARTICIPANTS: Self-selected sample of women recruited through social media (Facebook, Instagram, Twitter) between October and December 2020. Inclusion criteria included: identified as woman, ages 35 and over, living in Alberta, Canada. PRIMARY AND SECONDARY OUTCOMES MEASURES: Self-reported data were collected on demographics, menopause status and symptoms, cannabis usage and how participants perceived cannabis. Descriptive statistics, comparative analysis and logistic regression explored relations in cannabis use and participant characteristics. RESULTS: Of 1761 responses collected, 1485 were included for analysis. Median age was 49 years; 35% were postmenopausal and 33% perimenopausal. Among analysed responses, 499 (34%) women reported currently using cannabis and 978 (66%) indicated ever using cannabis. Of the 499 current cannabis users, over 75% were using cannabis for medical purposes. Most common reasons for current use were sleep (65%), anxiety (45%) and muscle/joint achiness (33%). In current users, 74% indicated that cannabis was helpful for symptoms. Current cannabis users were more likely to report experiencing menopause symptoms compared with non-users. History of smoking and general health status were associated with current cannabis use. CONCLUSIONS: Some women are using cannabis for symptoms related to menopause. Further research is required to assess safety and efficacy of cannabis for managing menopause and develop clinical resources for women on cannabis and menopause.

Clinical Evidence on the Purported Pharmacokinetic Interactions between Corticosteroids and Mycophenolic Acid.

Corticosteroids (steroids) are commonly used concurrently with mycophenolic acid (MPA) as the first-line immunosuppression therapy for the prevention of rejection in solid organ transplantations. Steroids are also commonly administered with MPA in various autoimmune disorders such as systemic lupus erythematosus and idiopathic nephrotic syndrome. Despite various review articles having suggested the presence of pharmacokinetic interactions between MPA and steroids, definitive data have not yet been demonstrated. The aim of this Current Opinion is to critically evaluate the available clinical data and propose the optimal study design for characterising the MPA-steroid pharmacokinetic interactions. The PubMed and Embase databases were searched for relevant clinical articles in English as of September 29, 2022, where a total of 8 papers have been identified as supporting and 22 as non-supporting the purported drug interaction. To objectively evaluate the data, novel assessment criteria to effectively diagnose the interaction based on known MPA pharmacology were formulated, including the availability of independent control groups, prednisolone concentrations, MPA metabolite data, unbound MPA concentrations, and the characterisations of entero-hepatic recirculation and MPA renal clearance. Overall, the majority of the identified corticosteroid data were pertaining to prednisone or prednisolone. Our assessment indicated that no conclusive mechanistic data supporting the interaction are available in the current clinical literature, and further studies are required to quantify the effects/mechanisms of steroid-tapering or withdrawal on MPA pharmacokinetics. This current opinion provides justification for further translational investigations, as this particular drug interaction has the potential to exert significant adverse outcomes in patients prescribed MPA.

Significant Correlations between p-Cresol Sulfate and Mycophenolic Acid Plasma Concentrations in Adult Kidney Transplant Recipients.

BACKGROUND AND OBJECTIVES: Mycophenolic acid (MPA) is a commonly prescribed life-long immunosuppressant for kidney transplant recipients. The frequently observed large variations in MPA plasma exposure may lead to severe adverse outcomes; therefore, characterizations of contributing factors can potentially improve the precision dosing of MPA. Our group recently reported the potent inhibitory effects of p-cresol (a protein-bound uremic toxin that can be accumulated in kidney transplant patients) on the hepatic metabolism of MPA in human in vitro models. Based on these data, the hypothesis for this clinical investigation was that a direct correlation between p-cresol and MPA plasma exposure should be evident in adult kidney transplant recipients. METHODS: Using a prospective and observational approach, adult kidney transplant recipients within the first year after transplant on oral mycophenolate mofetil (with tacrolimus ± prednisone) were screened for recruitment. The exclusion criteria were cold ischemia time > 30 h, malignancy, pregnancy, severe renal dysfunction (i.e., estimated glomerular filtration rate, eGFR, < 10 mL/min/1.73 m2), active graft rejection, or MPA intolerance. Patients' demographic and biochemistry data were collected. Total and free plasma concentrations of MPA, MPA glucuronide (MPAG), and total p-cresol sulfate (the predominant, quantifiable form of p-cresol in the plasma) were quantified using validated assays. Correlational and categorical analyses were performed using GraphPad Prism. RESULTS: Forty patients (11 females) were included: donor type (living/deceased: 20/20), induction regimen (basiliximab/thymoglobulin/basiliximab followed by thymoglobulin: 35/3/2), post-transplant time (74 ± 60 days, mean ± standard deviation), age (53.7 ± 12.4 years), bodyweight (79.8 ± 18.5 kg), eGFR (51.9 ± 18.0 mL/min/1.73 m2), serum albumin (3.6 ± 0.5 g/dL), prednisone dose (18.5 ± 13.2 mg, n = 33), and tacrolimus trough concentration (9.4 ± 2.4 µg/L). Based on Spearman analysis, significant control correlations supporting the validity of our dataset were observed between total MPA trough concentration (C0) and total MPAG C0 (correlation coefficient [R] = 0.39), ratio of total MPAG C0-to-total MPA C0 and post-transplant time (R = - 0.56), total MPAG C0 and eGFR (R = - 0.35), and p-cresol sulfate concentration and eGFR (R = - 0.70). Our primary analysis indicated the novel observation that total MPA C0 (R = 0.39), daily dose-normalized total MPA C0 (R = 0.32), and bodyweight-normalized total MPA C0 (R = 0.32) were significantly correlated with plasma p-cresol sulfate concentrations. Consistently, patients categorized with elevated p-cresol sulfate concentrations (i.e., ≥ median of 3.2 µg/mL) also exhibited increased total MPA C0 (by 57 % vs those below median), daily dose-normalized total MPA C0 (by 89 %), and bodyweight-normalized total MPA C0 (by 62 %). Our secondary analyses with MPA metabolites, unbound concentrations, free fractions, and MPA metabolite ratios supported additional potential interacting mechanisms. CONCLUSION: We have identified a novel, positive association between p-cresol sulfate exposure and total MPA C0 in adult kidney transplant recipients, which is supported by published mechanistic in vitro data. Our findings confirm a potential role of p-cresol as a significant clinical variable affecting the pharmacokinetics of MPA. These data also provide the justifications for conducting subsequent full-scale pharmacokinetic-pharmacodynamic studies to further characterize the cause-effect relationships of this interaction, which could also rule out potential confounding variables not adequately controlled in this correlational study.

Effects of p-Cresol on Oxidative Stress, Glutathione Depletion, and Necrosis in HepaRG Cells: Comparisons to Other Uremic Toxins and the Role of p-Cresol Glucuronide Formation.

The toxicological effects of p-cresol have primarily been attributed to its metabolism products; however, very little human data are available in the key organ (i.e., liver) responsible for the generation of these metabolites. Experiments were conducted in HepaRG cells utilizing the following markers of cellular toxicity: 2'-7'-dichlorofluorescein (DCF; oxidative stress) formation, total cellular glutathione (GSH) concentration, and lactate dehydrogenase (LDH; cellular necrosis) release. Concentrations of p-cresol, p-cresol sulfate, and p-cresol glucuronide were determined using validated assays. p-Cresol exposure resulted in concentration- and time-dependent changes in DCF (EC50 = 0.64 ± 0.37 mM at 24 h of exposure) formation, GSH (EC50 = 1.00 ± 0.07 mM) concentration, and LDH (EC50 = 0.85 ± 0.14 mM) release at toxicologically relevant conditions. p-Cresol was also relatively more toxic than 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid, indole-3-acetic acid, indoxyl sulfate, kynurenic acid, and hippuric acid on all markers. Although the exogenous administration of p-cresol sulfate and p-cresol glucuronide generated high intracellular concentrations of these metabolites, both metabolites were less toxic compared to p-cresol at equal-molar conditions. Moreover, p-cresol glucuronide was the predominant metabolite generated in situ from p-cresol exposure. Selective attenuation of glucuronidation (without affecting p-cresol sulfate formation, while increasing p-cresol accumulation) using independent chemical inhibitors (i.e., 0.75 mM l-borneol, 75 µM amentoflavone, or 100 µM diclofenac) consistently resulted in further increases in LDH release associated with p-cresol exposure (by 28.3 ± 5.3%, 30.0 ± 8.2% or 27.3 ± 6.8%, respectively, compared to p-cresol treatment). These novel data indicated that p-cresol was a relatively potent toxicant, and that glucuronidation was unlikely to be associated with the manifestation of its toxic effects in HepaRG cells.

Characterization of human sulfotransferases catalyzing the formation of p-cresol sulfate and identification of mefenamic acid as a potent metabolism inhibitor and potential therapeutic agent for detoxification.

p-Cresol sulfate, the primary metabolite of p-cresol, is a uremic toxin that has been associated with toxicities and mortalities. The study objectives were to i) characterize the contributions of human sulfotransferases (SULT) catalyzing p-cresol sulfate formation using multiple recombinant SULT enzymes (including the polymorphic variant SULT1A1*2), pooled human liver cytosols, and pooled human kidney cytosols; and ii) determine the potencies and mechanisms of therapeutic inhibitors capable of attenuating the production of p-cresol sulfate. Human recombinant SULT1A1 was the primary enzyme responsible for the formation of p-cresol sulfate (Km = 0.19 ±â€¯0.02 µM [with atypical kinetic behavior at lower substrate concentrations; see text discussion], Vmax = 789.5 ±â€¯101.7 nmol/mg/min, Ksi = 2458.0 ±â€¯332.8 µM, mean ±â€¯standard deviation, n = 3), while SULT1A3, SULT1B1, SULT1E1, and SULT2A1 contributed negligible or minor roles at toxic p-cresol concentrations. Moreover, human recombinant SULT1A1*2 exhibited reduced enzyme activities (Km = 81.5 ±â€¯31.4 µM, Vmax = 230.6 ±â€¯17.7 nmol/mg/min, Ksi = 986.0 ±â€¯434.4 µM) compared to the wild type. The sulfonation of p-cresol was characterized by Michaelis-Menten kinetics in liver cytosols (Km = 14.8 ±â€¯3.4 µM, Vmax = 1.5 ±â€¯0.2 nmol/mg/min) and substrate inhibition in kidney cytosols (Km = 0.29 ±â€¯0.02 µM, Vmax = 0.19 ±â€¯0.05 nmol/mg/min, Ksi = 911.7 ±â€¯278.4 µM). Of the 14 investigated therapeutic inhibitors, mefenamic acid (Ki = 2.4 ±â€¯0.1 nM [liver], Ki = 1.2 ±â€¯0.3 nM [kidney]) was the most potent in reducing the formation of p-cresol sulfate, exhibiting noncompetitive inhibition in human liver cytosols and recombinant SULT1A1, and mixed inhibition in human kidney cytosols. Our novel findings indicated that SULT1A1 contributed an important role in p-cresol sulfonation (hence it can be considered a probe reaction) in liver and kidneys, and mefenamic acid may be utilized as a potential therapeutic agent to attenuate the generation of p-cresol sulfate as an approach to detoxification.

Recent lessons learned from population pharmacokinetic studies of mycophenolic acid: physiological, genomic, and drug interactions leading to the prediction of drug effects.

INTRODUCTION: Mycophenolic acid (MPA) is a widely used immunosuppressant in transplantation and autoimmune disease. Highly variable pharmacokinetics have been observed with MPA, but the exact mechanisms remain largely unknown. AREAS COVERED: The current review provided a critical, comprehensive update of recently published population pharmacokinetic/dynamic models of MPA (n = 16 papers identified from PubMed and Embase, inclusive from January 2017 to August 2021), with specific emphases on the intrinsic and extrinsic factors influencing the pharmacology of MPA. The significance of the identified covariates, potential mechanisms, and comparisons to historical literature have been provided. EXPERT OPINION: While select covariates affecting the population pharmacokinetics of MPA are consistently observed and mechanistically supported (e.g. cyclosporine and post-transplant time on MPA clearance), some variables have not been regularly reported and/or lacked mechanistic explanation (e.g. diarrhea and several genetic polymorphisms). Very few pharmacodynamic models were available, pointing to the need to extrapolate pharmacokinetic findings. Ideal models of MPA should consist of: i) utilizing optimal sampling points to allow the characterizations of absorption, re-absorption, and elimination phases; ii) characterizing unbound/total MPA, MPA metabolites, plasma/urinary concentrations, and genetic polymorphisms to facilitate mechanistic interpretations; and iii) incorporating actual outcomes (e.g. rejection, leukopenia, infections) and pharmacodynamic data (e.g. inosine-5'-monophosphate dehydrogenase activities) to establish clinical relevance. We anticipate the field will continue to expand in the next 5 to 10 years.

Population pharmacokinetics of mycophenolic acid in paediatric patients.

Mycophenolic acid (MPA) is widely used in paediatric kidney transplant patients and sometimes prescribed for additional indications. Population pharmacokinetic or pharmacodynamic modelling has been frequently used to characterize the fixed, random and covariate effects of MPA in adult patients. However, MPA population pharmacokinetic data in the paediatric population have not been systematically summarized. The objective of this narrative review was to provide an up-to-date critique of currently available paediatric MPA population pharmacokinetic models, with emphases on modelling techniques, pharmacological findings and clinical relevance. PubMed and EMBASE were searched from inception of database to May 2020, where a total of 11 studies have been identified representing kidney transplant (n = 4), liver transplant (n = 1), haematopoietic stem cell transplant (n = 1), idiopathic nephrotic syndrome (n = 2), systemic lupus erythematosus (n = 2), and a combined population consisted of kidney, liver and haematopoietic stem cell transplant patients (n = 1). Critical analyses were provided in the context of MPA absorption, distribution, metabolism, excretion and bioavailability in this paediatric database. Comparisons to adult patients were also provided. With respect to clinical utility, Bayesian estimation models (n = 6) with acceptable accuracy and precision for MPA exposure determination have also been identified and systematically evaluated. Overall, our analyses have identified unique features of MPA clinical pharmacology in the paediatric population, while recognizing several gaps that still warrant further investigations. This review can be used by pharmacologists and clinicians for improving MPA pharmacokinetic-pharmacodynamic modelling and patient care.

Clinical Pharmacokinetics and Pharmacodynamics of Anti-Tubercular Drugs in Pregnancy.

The objectives of this qualitative review were to critically evaluate and summarize the currently available data on the use of anti-tuberculosis (TB) drugs during pregnancy, with a focus on treatment outcomes, safety, and pharmacokinetics. This qualitative, narrative review was based on literature searches in Medline, Pubmed, Embase, and Google Scholar (from their inception to 13 August 2020). Our search identified 22 papers related to treatment outcomes and 14 papers related to pharmacokinetic exposures and fetal distributions. While it is challenging to study this patient population, current evidence supports treatment of drug-susceptible TB, multidrug-resistant TB and latent TB infections. However, decisions regarding initiating, continuing, or discontinuing anti-tubercular medications while pregnant should be individualized and discussed with a specialist. Similarly, the pharmacokinetic data of anti-TB agents were mainly derived from small scale, observational studies many of which lacked high quality controls. Based on these data, it does not appear that pregnancy has an extensive impact on the pharmacokinetics of the majority of first-line and second-line agents, although caution (discussed in the review) should be exercised in data interpretation. Fetal drug exposure can also be significant and should be considered when selecting an anti-TB agent for longer term treatment. Overall, it is generally difficult to predict pregnancy-associated pharmacokinetic changes based only on drug's physiochemical characteristics.

Comparisons of Four Protein-Binding Models Characterizing the Pharmacokinetics of Unbound Phenytoin in Adult Patients Using Non-Linear Mixed-Effects Modeling.

BACKGROUND AND OBJECTIVE: Phenytoin is extensively protein bound with a narrow therapeutic range. The unbound phenytoin is pharmacologically active, but total concentrations are routinely measured in clinical practice. The relationship between free and total phenytoin has been described by various binding models with inconsistent findings. Systematic comparison of these binding models in a single experimental setting is warranted to determine the optimal binding behaviors. METHODS: Non-linear mixed-effects modeling was conducted on retrospectively collected data (n = 37 adults receiving oral or intravenous phenytoin) using a stochastic approximation expectation-maximization algorithm in MonolixSuite-2019R2. The optimal base structural model was initially developed and utilized to compare four binding models: Winter-Tozer, linear binding, non-linear single-binding site, and non-linear multiple-binding site. Each binding model was subjected to error and covariate modeling. The final model was evaluated using relative standard errors (RSEs), goodness-of-fit plots, visual predictive check, and bootstrapping. RESULTS: A one-compartment, first-order absorption, Michaelis-Menten elimination, and linear protein-binding model best described the population pharmacokinetics of free phenytoin at typical clinical concentrations. The non-linear single-binding-site model also adequately described phenytoin binding but generated larger RSEs. The non-linear multiple-binding-site model performed the worst, with no identified covariates. The optimal linear binding model suggested a relatively high binding capacity using a single albumin site. Covariate modeling indicated a positive relationship between albumin concentration and the binding proportionality constant. CONCLUSIONS: The linear binding model best described the population pharmacokinetics of unbound phenytoin in adult subjects and may be used to improve the prediction of free phenytoin concentrations.

Characterizations of Human UDP-Glucuronosyltransferase Enzymes in the Conjugation of p-Cresol.

p-Cresol is a uremic toxin that is formed by intestinal microbiota and extensively conjugated by first-pass metabolism. p-Cresol glucuronide exerts various forms of cellular toxicity in vitro and is accumulated in the plasma of subjects with kidney disease, where associations with adverse cardiovascular and renal outcomes are evident. The objective of this study was to determine the contributions of human UDP-glucuronosyltransferase (UGT) enzymes in the formation of p-cresol glucuronide. Utilizing commonly expressed hepatic or renal human recombinant UGTs (ie, hrUGT1A1, 1A3, 1A4, 1A6, 1A7, 1A8, 1A9, 1A10, 2B4, 2B7, 2B10, 2B15, and 2B17), hrUGT1A6 and hrUGT1A9 exhibited the highest catalytic activities in the generation of p-cresol glucuronide. The kinetics of p-cresol glucuronide formation in hrUGT1A6 and pooled human liver microsomes were best described by the Hill equation and in hrUGT1A9 and pooled human kidney microsomes by substrate inhibition. Using inhibitory and selective UGT inhibitors (ie, acetaminophen or amentoflavone for UGT1A6 and niflumic acid for UGT1A9), UGT1A6 was identified the predominant enzyme responsible for p-cresol glucuronide production in pooled human liver (78.4%-81.3% contribution) and kidney (54.3%-62.9%) microsomes, whereas UGT1A9 provided minor contributions (2.8% and 35.5%, respectively). The relative contributions of UGT1A6 (72.6 ± 11.3%, mean ± SD) and UGT1A9 (5.7 ± 4.1%) in individual human liver microsomes from 12 adult donors were highly variable, where an inverse association (R = -.784, p = .003) between UGT1A6 contribution and UGT1A9 probe substrate activity (ie, mycophenolic acid) was evident. Our novel findings provide valuable tools for conducting further mechanistic studies and for designing clinical interventions to mitigate the toxicities associated with p-cresol glucuronide.

Mechanisms of Metabolism Interaction Between p-Cresol and Mycophenolic Acid.

Mycophenolic acid (MPA) is commonly prescribed for preventing graft rejection after kidney transplantation. The primary metabolic pathways of MPA are hepatic glucuronidation through UDP-glucuronosyltransferase (UGT) enzymes in the formation of MPA-glucuronide (MPAG, major pathway) and MPA-acyl glucuronide (AcMPAG). p-Cresol, a potent uremic toxin known to accumulate in patients with renal dysfunction, can potentially interact with MPA via the inhibition of glucuronidation. We hypothesized that the interaction between MPA and p-cresol is clinically relevant and that the estimated exposure changes in the clinic are of toxicological significance. Using in vitro approaches (ie, human liver microsomes and recombinant enzymes), the potency and mechanisms of inhibition by p-cresol towards MPA glucuronidation were characterized. Inter-individual variabilities, effects of clinical co-variates, in vitro-in vivo prediction of likely changes in MPA exposure, and comparison to other toxins were determined for clinical relevance. p-Cresol inhibited MPAG formation in a potent and competitive manner (Ki=5.2 µM in pooled human liver microsomes) and the interaction was primarily mediated by UGT1A9. This interaction was estimated to increase plasma MPA exposure in patients by approximately 1.8-fold, which may result in MPA toxicity. The mechanism of inhibition for AcMPAG formation was noncompetitive (Ki=127.5 µM) and less likely to be clinically significant. p-Cresol was the most potent inhibitor of MPA-glucuronidation compared with other commonly studied uremic toxins (eg, indole-3-acetic acid, indoxyl sulfate, hippuric acid, kynurenic acid, and 3-carboxy-4-methyl-5-propyl-2-furanpropionic acid) and its metabolites (ie, p-cresol sulfate and p-cresol glucuronide). Our findings indicate that the interaction between p-cresol and MPA is of toxicological significance and warrants clinical investigation.

Exposure-Toxicity Relationships of Mycophenolic Acid in Adult Kidney Transplant Patients.

Mycophenolic acid is commonly prescribed in adult kidney transplant recipients for preventing graft rejection. A therapeutic target for total mycophenolic acid area under the concentration-time curve (30-60 mg h/L) has been established in adult kidney transplant recipients and widely referenced today. However, this specific target range does not adequately characterize mycophenolic acid-associated adverse effects. The primary objective of this qualitative and critical review was to characterize the exposure-toxicity relationships of mycophenolic acid in an attempt to determine whether exposure thresholds can be identified. The secondary objective was to determine the associations of clinical variables with specific adverse effects. The inclusion criteria consisted of all peer-reviewed papers in adult kidney transplant subjects (average study age > 18 years) with both exposure (area under the concentration-time curve) and toxicity data. The exclusion criteria were papers involving the pediatric population, studies lacking either area under the concentration-time curve or toxicity data, or studies with no apparent reported variations in area under the concentration-time curves. Of the 28 papers identified, inconsistent findings have been reported for the most frequently characterized adverse events of mycophenolic acid (gastrointestinal, infectious, and hematological), while promising exposure thresholds (i.e., > 40-60 mg h/L for total mycophenolic acid) have been suggested by a few studies. The roles of free mycophenolic acid exposure, mycophenolic acid metabolites, or clinical factors influencing the manifestation of the toxicities also remain to be clarified. Although it is not yet possible to define toxicity threshold(s) for the purpose of mycophenolic acid therapeutic drug monitoring, the information obtained and the limitations identified in this comprehensive literature body have provided a good foundation for future investigations.

Population Pharmacokinetics of Mycophenolic Acid Co-Administered with Tacrolimus in Corticosteroid-Free Adult Kidney Transplant Patients.

BACKGROUND AND OBJECTIVE: Mycophenolic acid is commonly prescribed to adult kidney transplant recipients. Mycophenolic acid is extensively metabolized to mycophenolic acid-glucuronide (major metabolite) and mycophenolic acid-acyl-glucuronide (minor metabolite). We hypothesized that (1) adult kidney transplant patients on corticosteroid-free regimens exhibit unique mycophenolic acid population pharmacokinetics compared with patients receiving corticosteroid-based therapy, and (2) mycophenolic acid clearance is directly dependent on glucuronide metabolite formation. METHODS: Non-linear mixed-effects modeling was conducted with MonolixSuite-2018R1 (n = 27). Optimal pharmacokinetic models were selected based on objective function values, standard errors, and biological plausibility. RESULTS: Clinical demographic data were sex (female, 16), age (47 ± 13 years, mean ± standard deviation), weight (70 ± 16 kg), height (165 ± 9 cm), albumin (43 ± 4 g/L), serum creatinine (102 ± 27 µmol/L), estimated glomerular filtration rate (61 ± 16 mL/min/1.73 m2), mycophenolic acid dosage (1.4 ± 0.5 g/day, as mycophenolate mofetil), and tacrolimus dosage (5 ± 3 mg/day, immediate release). The population pharmacokinetics of mycophenolic acid can be described by a two-compartment first-order absorption with lag time, and a linear elimination structural model. The apparent oral clearance estimate in the final model (population mean, relative standard error) was 2.87 L/h, 42.3%, which is lower than that reported for similar patients on corticosteroid-based regimens (11.9-26.3 L/h). Other pharmacokinetic parameters were comparable to historical data obtained in corticosteroid-based patients. Both mycophenolic acid-acyl-glucuronide trough concentration and the area under the concentration-time curve ratio were significant covariates that reduced mycophenolic acid apparent oral clearance from 16.5 (base model) to 2.87 L/h. The model was evaluated based on bootstrapping, visual predictive checks, and diagnostic plots. CONCLUSIONS: Our novel findings suggest the potential need to reduce mycophenolic acid dosage in subjects on corticosteroid-free regimens. Corticosteroid-free subjects may also be more sensitive to drug/gene interactions.