Roles of Individual Human Cytochrome P450 Enzymes in Drug Metabolism.

Our knowledge of the roles of individual cytochrome P450 (P450, CYP) enzymes in drug metabolism has developed considerably in the past 30 years, and this base has been of considerable use in avoiding serious issues with drug interactions and issues due to variations. Some newer approaches are being considered for "phenotyping" of metabolism reactions with new drug candidates. Endogenous biomarkers are being used for non-invasive estimation of levels of individual P450 enzymes. There is also the matter of some remaining "orphan" P450s, which have yet to be assigned reactions. Practical problems that continue in drug development include predicting drug-drug interactions, predicting the effects of polymorphic and other P450 variations, and evaluating inter-species differences in drug metabolism, particularly in the context of "metabolism in safety testing" (MIST) regulatory issues ("disproportionate (human) metabolites"). Significance Statement Cytochrome P450 enzymes are the major catalysts involved in drug metabolism. The characterization of their individual roles has major implications in drug development and clinical practice.

The Pharmacokinetics, Metabolism, and Clearance Mechanisms of Ritlecitinib, a Janus Kinase 3 and Tyrosine-Protein Kinase Family Inhibitor, in Humans.

Ritlecitinib is an oral once-daily irreversible inhibitor of Janus kinase 3 and tyrosine-protein kinase family being developed for the treatment of moderate-to-severe alopecia areata. This study examined the disposition of ritlecitinib in male participants following oral and intravenous administration using accelerator mass spectroscopy methodology to estimate pharmacokinetic parameters and characterize metabolite profiles. The results indicated ritlecitinib had a systemic clearance of 43.7 L/h, a steady state volume of distribution of 73.8 L, extent of absorption of 89%, time to maximum plasma concentration of ~0.5 hour, and absolute oral bioavailability of 64%. An observed long terminal half-life of total radioactivity was primarily attributed to ritlecitinib binding to plasma albumin. Ritlecitinib was the main circulating drug species in plasma (~30%) with one major pharmacologically inactive cysteine conjugated metabolite (M2) at >10%. Oxidative metabolism (fractional clearance 0.47) and glutathione related conjugation (fractional clearance 0.24) were the primary routes of elimination for ritlecitinib with the greatest disposition of radioactivity shown in the urine (~71%). In vitro phenotyping indicated ritlecitinib cytochrome P450 fraction of metabolism assignments of 0.29 for CYP3A, 0.09 for CYP2C8, 0.07 for CYP1A2, and 0.02 for CYP2C9. In vitro phenotyping in recombinant human glutathione S-transferases indicated ritlecitinib was turned over by a number of cytosolic and microsomal enzyme isoforms. Significance Statement This study provides a detailed understanding of the disposition and metabolism of ritlecitinib, a JAK3 and TEC family kinase inhibitor for alopecia areata, in humans, as well as characterization of clearance pathways and PK of ritlecitinib and its metabolites. As an AMS-based ADME study design, we have expanded on reporting the standard ADME endpoints, providing key pharmacokinetic parameters like clearance, volume of distribution and bioavailability allowing for a more comprehensive understanding of drug disposition.

Low molecular weight acids and OATP1B mediated hepatic clearance: In vitro and in vivo evaluation using novel hypoxia-inducible factor prolyl hydroxylase inhibitors (Dustats).

Organic anion transporting polypeptide (OATP1B) plays a key role in the hepatic clearance of a majority of high molecular weight (MW) acids and zwitterions. Here, we evaluated the role of OATP1B-mediated uptake in the clearance of novel hypoxia-inducible factor prolyl hydroxylase inhibitors ("Dustats"), which are typically low MW (300-400 daltons) aliphatic carboxylic acids. Five acid dustats, namely daprodustat, desidustat, enarodustat, roxadustat and vadadustat, showed specific transport by OATP1B1/1B3 in transporter-transfected HEK293 cells. Neutral compound, molidustat, was not a substrate to OATP1B1/1B3. None of the dustats showed transport by other hepatic uptake transporters, including NTCP, OAT2 and OAT7. In the primary human hepatocytes, uptake of all acids was significantly reduced by rifampin (OATP1B inhibitor); with an estimated fraction transported by OATP1B (ft ,OATP1B) of up to >80% (daprodustat). Molidustat uptake was minimally inhibited by rifampin; and low permeability acids (desidustat and enarodustat) also showed biliary efflux in sandwich culture human hepatocytes. In vivo, intravenous pharmacokinetics of all 5 acids was significantly altered by a single-dose rifampin (30 mg/kg) in Cynomolgus monkey. Hepatic clearance (non-renal) was about 4-fold (vadadustat) to >11-fod (daprodustat and roxadustat) higher in control group compared to rifampin-treated subjects. In vivo ft ,OATP1B was estimated to be ~70-90%. In the case of molidustat, rifampin had a minimal effect on overall clearance. Rifampin also considerably reduced volume of distribution of daprodustat and roxadustat. Overall, OATP1B significantly contribute to the hepatic clearance and pharmacokinetics of several dustats, which are low MW carboxylic acids. OATP1B activity should therefore by evaluated in this property space. Significance Statement Our in vitro and in vivo results suggest that OATP1B-mediated hepatic uptake play a significant role in the pharmacokinetics of low MW acidic dustats, which are being developed or approved for the treatment of anemia in chronic kidney disease. Significant active uptake mechanisms are not apparent for the neutral compound, molidustat. Characterization of uptake mechanisms is therefore important in predicting human pharmacokinetics and evaluating drug-drug interactions for low MW acids.

Real-world application of PBPK in drug discovery.

The utility of PBPK models in support of drug development has been well documented. During the discovery stage, PBPK has increasingly been applied for early risk assessment, prediction of human dose, toxicokinetic dose projection and early formulation assessment. Previous review articles have proposed model building and application strategies for PBPK-based first in human predictions with comprehensive descriptions of the individual components of PBPK models. This includes the generation of decision trees, based on comprehensive literature reviews, to guide the application of PBPK in the discovery setting. The goal of this mini review is to provide additional guidance on the real-world application of PBPK, in support of the discovery stage of drug development. In this mini review, our goal is to provide guidance on the typical steps involved in the development and application of a PBPK model during drug discovery to assist in decision making. We have illustrated our recommended approach through description of case examples, where PBPK has been successfully applied to aid in human PK projection, candidate selection and prediction of drug interaction liability for parent and metabolite. Through these case studies, we have highlighted fundamental issues, including pre-verification in preclinical species, the application of empirical scalars in the prediction of in vivo clearance from in vitro systems, in silico prediction of permeability and the exploration of aqueous and biorelevant solubility data to predict dissolution. In addition, current knowledge gaps have been highlighted and future directions proposed. Significance Statement Through description of three case studies, we have highlighted the fundamental principles of PBPK application during drug discovery. These include pre-verification of the model in preclinical species, application of empirical scalars where necessary in the prediction of clearance, in silico prediction of permeability, and the exploration of aqueous and biorelevant solubility data to predict dissolution. In addition, current knowledge gaps have been highlighted and future directions proposed.

Application of empirical scalars to enable early prediction of human hepatic clearance using IVIVE in drug discovery: an evaluation of 173 drugs.

The utilization of in vitro data to predict drug pharmacokinetics (PK) in vivo has been a consistent practice in early drug discovery for decades. However, its success is hampered by mispredictions attributed to uncharacterized biological phenomena/experimental artifacts. Predicted drug clearance (CL) from experimental data (i.e. hepatocyte intrinsic clearance: CLint, fraction unbound in plasma: fu,p) is often systematically underpredicted using the well-stirred model (WSM). The objective of this study was to evaluate using empirical scalars in the WSM to correct for CL mispredictions. Drugs (N=28) were used to generate numerical scalars on CLint (α), and fu,p (ß) to minimize the error (AAFE) for CL predictions. These scalars were validated using an additional dataset (N=28 drugs) and applied to a non-redundant AstraZeneca (AZ) dataset available in the literature (N=117 drugs) for a total of 173 compounds. CL predictions using the WSM were improved for most compounds using an α value of 3.66 (~64%<2-fold) compared to no scaling (~46%<2-fold). Similarly, using a ß value of 0.55 or combination of α and ß scalars (values of 1.74 and 0.66, respectively) resulted in a similar improvement in predictions (~64%<2-fold and ~65%<2-fold, respectively). For highly bound compounds (fu,p{less than or equal to}0.01), AAFE was substantially reduced across all scaling methods. Using the ß scalar alone or a combination of α and ß appeared optimal; and produce larger magnitude corrections for highly-bound compounds. Some drugs are still disproportionally mispredicted, however the improvements in prediction error and simplicity of applying these scalars suggests its utility for early-stage CL predictions. Significance Statement In early drug discovery, prediction of human clearance using in vitro experimental data plays an essential role in triaging compounds prior to in vivo studies. These predictions have been systematically underestimated. Here we introduce empirical scalars calibrated on the extent of plasma protein binding that appear to improve clearance prediction across multiple datasets. This approach can be used in early phases of drug discovery prior to the availability of pre-clinical data for early quantitative predictions of human clearance.

Influence of Proteome Profiles and Intracellular Drug Exposure on Differences in CYP Activity in Donor-Matched Human Liver Microsomes and Hepatocytes.

Human liver microsomes (HLM) and human hepatocytes (HH) are important in vitro systems for studies of intrinsic drug clearance (CLint) in the liver. However, the CLint values are often in disagreement for these two systems. Here, we investigated these differences in a side-by-side comparison of drug metabolism in HLM and HH prepared from 15 matched donors. Protein expression and intracellular unbound drug concentration (Kpuu) effects on the CLint were investigated for five prototypical probe substrates (bupropion-CYP2B6, diclofenac-CYP2C9, omeprazole-CYP2C19, bufuralol-CYP2D6, and midazolam-CYP3A4). The samples were donor-matched to compensate for inter-individual variability but still showed systematic differences in CLint. Global proteomics analysis outlined differences in HLM from HH and homogenates of human liver (HL), indicating variable enrichment of ER-localized cytochrome P450 (CYP) enzymes in the HLM preparation. This suggests that the HLM may not equally and accurately capture metabolic capacity for all CYPs. Scaling CLint with CYP amounts and Kpuu could only partly explain the discordance in absolute values of CLint for the five substrates. Nevertheless, scaling with CYP amounts improved the agreement in rank order for the majority of the substrates. Other factors, such as contribution of additional enzymes and variability in the proportions of active and inactive CYP enzymes in HLM and HH, may have to be considered to avoid the use of empirical scaling factors for prediction of drug metabolism.

Augmented renal clearance in pediatric intensive care: are we undertreating our sickest patients?

Many critically ill patients display a supraphysiological renal function with enhanced renal perfusion and glomerular hyperfiltration. This phenomenon described as augmented renal clearance (ARC) may result in enhanced drug elimination through renal excretion mechanisms. Augmented renal clearance seems to be triggered by systemic inflammation and therapeutic interventions in intensive care. There is growing evidence that ARC is not restricted to the adult intensive care population, but is also prevalent in critically ill children. Augmented renal clearance is often overlooked due to the lack of reliable methods to assess renal function in critically ill children. Standard equations to calculate glomerular filtration rate (GFR) are developed for patients who have a steady-state creatinine production and a stable renal function. Those formulas are not reliable in critically ill patients with acutely changing GFR and tend to underestimate true GFR in patients with ARC. Tools for real-time, continuous, and non-invasive measurement of fluctuating GFR are most needed to identify changes in kidney function during critical illness and therapeutic interventions. Such devices are currently being validated and hold a strong potential to become the standard of practice. In the meantime, urinary creatinine clearance is considered the most reliable method to detect ARC in critically ill patients. Augmented renal clearance is clearly associated with subtherapeutic antimicrobial concentrations and subsequent therapeutic failure. This warrants the need for adjusted dosing regimens to optimize pharmacokinetic and pharmacodynamic target attainment. This review aims to summarize current knowledge on ARC in critically ill children, to give insight into its possible pathophysiological mechanism, to evaluate screening methods for ARC in the pediatric intensive care population, and to illustrate the effect of ARC on drug exposure, therapeutic efficacy, and clinical outcome.

Ex vivo Rezafungin Adsorption and Clearance During Continuous Renal Replacement Therapy.

BACKGROUND/AIMS: To determine adsorption and transmembrane clearances (CLTM) of rezafungin, a novel long-acting echinocandin, in continuous venovenous hemofiltration (CVVH). METHODS: A validated ex vivo bovine blood CVVH model using polysulfone and AN69 hemodiafilters was used to evaluate urea and rezafungin CLTM at 3 different ultrafiltrate flow rates. Rezafungin adsorption to the CRRT apparatus was determined for each hemodiafilter. RESULTS: The sieving coefficient (SC) from CVVH with 3 different ultrafiltrate flow rates was 0 for both HF1400 and Multiflow-150 hemodiafilters, while urea SC was approximately 1 at all flow rates. Hemodiafilter type and ultrafiltrate flow rate did not influence CLTM. Rezafungin adsorption to the CVVH apparatus was not observed for either hemodiafilter. CONCLUSION: Rezafungin is not removed by CVVH by membrane adsorption or via CLTM. Ultrafiltrate flow rates and hemodiafilter types are unlikely to influence rezafungin CLTM. No dosage adjustment of rezafungin is likely required for critically ill patients receiving CVVH.

Treatment of Gabapentin Toxicity With Peritoneal Dialysis: Assessment of Gabapentin Clearance.

Gabapentin is almost exclusively cleared by the kidney and thus presents challenges in patients with kidney failure. Gabapentin is known to be effectively cleared by hemodialysis, but the efficiency of clearance by peritoneal dialysis (PD) has not been previously described. We report a case of gabapentin toxicity in a patient on long-term PD who was treated with continuous automated cycling PD. We find that continuous PD provides significant clearance of gabapentin. With 2-L exchanges every 2 hours, we document an apparent elimination half-life of 41.33 hours, which is substantially shorter than the reported elimination half-life of 132 hours in the absence of kidney function. Further, our patient's symptoms of gabapentin toxicity gradually improved and had fully resolved after about 36 hours of dialysis. Gabapentin clearance by PD was estimated at 94% of urea clearance. We conclude that intensive PD provides gabapentin clearance that approximates that of urea and is an effective but slow method to treat gabapentin overdose and toxicity.

Treatment of Metformin Intoxication Complicated by Lactic Acidosis and Acute Kidney Injury: The Role of Prolonged Intermittent Hemodialysis.

Metformin intoxication with lactic acidosis, a potentially lethal condition, may develop in diabetic patients when the drug dose is inappropriate and/or its clearance is reduced. Diagnosis and therapy may be delayed due to nonspecific symptoms at presentation, with severe anion gap metabolic acidosis and elevated serum creatinine values being the most prominent laboratory findings. Confirmation requires measurement of serum metformin by high-performance liquid chromatography-tandem mass spectrometry, but this technique is available only at specialized institutions and cannot be relied on as a guide to immediate treatment. Thus, based on strong clinical suspicion, renal replacement therapy must be started promptly to achieve efficient drug clearance and correct the metabolic acidosis. However, because metformin accumulates in the intracellular compartment with prolonged treatment, a rebound in serum concentrations due to redistribution is expected at the end of dialysis. We report a case of metformin intoxication, severe lactic acidosis, and acute kidney injury in a diabetic patient with pre-existing chronic kidney disease stage 3, treated effectively with sustained low-efficiency dialysis. We discuss the pathophysiology, differential diagnosis, and treatment options and highlight specific pharmacokinetic issues that should be considered in selecting the appropriate modality of renal replacement therapy.

From Genotype to Phenotype: Cytochrome P450 2D6-Mediated Drug Clearance in Humans.

How genotypic variation results in phenotypic differences is still a challenge for biology. In the field of drug metabolism, the means by which specific cytochrome P4502D6 (CYP2D6) genotypes yield different phenotypes at various levels (molecular, cellular, and organismal) is an important question, as differences in CYP2D6 activity can contribute to adverse drug reactions. Herein, the genotype of CYP2D6 was determined along with the absolute content of CYP2D6 and microsomal protein per gram of liver in human liver microsomes, the molecular, cellular (microsomal, tissue, organ), and organismal phenotype of CYP2D6 determined; the effect of genotype on each phenotype of CYP2D6-mediated dextromethorphan clearance (CL) was delineated, and the overall genotype-phenotype relationship for CYP2D6 was charted. We demonstrate that changes in the cellular and organismal CL phenotypes are markedly greater than changes seen at the molecular level. With individuals carrying the 1661CC polymorphism, for example, the most noticeable change took place in organ CL phenotype (4.17-fold), followed by tissue (3.75-fold), organism (3.69-fold), microsomal (3.09-fold), and molecular (1.66-fold) phenotypes. In addition, the biggest intragenotype individual coefficient of variation in organismal phenotype was observed in the 1661GG individuals, which reached 104.5%, followed by that of 100TT, 100CT, 1661GC, 100CC, and 1661CC polymorphisms (102.7%, 62.4%, 53.5%, 49.7%, and 44.8%, respectively). Our study has allowed us to chart the genotype-phenotype relationship for CYP2D6 from the molecular to the organismal level as well as allowed us to determine intragenotype individual variation in phenotype with each genotype.

In vitro effects of an extracorporeal membrane oxygenation circuit on the sequestration of ϵ-aminocaproic acid.

OBJECTIVE: To assess the in vitro effects of drug sequestration in extracorporeal membrane oxygenation (ECMO) on ϵ-aminocaproic acid (EACA) concentrations. METHODS AND DESIGN: This in vitro study will determine changes in EACA concentration over time in ECMO circuits. A pediatric dose of 2,500 mg was administered to whole expired blood in the simulated pediatric ECMO circuit. Blood samples were collected at 0, 30, 60, 360 and 1440-minute intervals after initial administration equilibration from three different sites of the circuit: pre-oxygenator (PRE), post-oxygenator (POST) and PVC tubing (PVC) to determine the predominant site of drug loss. The circuit was maintained for two consecutive days with a re-dose at 24 hours to establish a comparison between unsaturated (New) and saturated (Old) oxygenator membranes. Comparisons between sample sites, sample times and New versus Old membranes were statistically analyzed by a linear mixed-effects model with significance defined as a p-value <0.05. RESULTS: There were no significant differences in EACA concentration with respect to sample site, with PRE and POST samples demonstrating respective mean differences of 0.30 mg/ml and 0.34 mg/ml as compared to PVC, resulting in non-significant p-values of 0.373 [95% CI (-0.37, 0.98)] and 0.324 [95% CI (-0.34, 1.01)], respectively. The comparison of New vs. Old ECMO circuits resulted in non-significant changes from baseline, with a mean difference of 0.50 mg/ml, 95% CI (-0.65, 1.65), p=0.315. CONCLUSION: The findings of this study did not show any significant changes in drug concentration that can be attributed to sequestration within the ECMO circuit. Mean concentrations between ECMO circuit sample sites did not differ significantly. Comparison between New and Old circuits also did not differ significantly in the change from baseline concentration over time. Sequestration within ECMO circuits appears not to be a considerable factor for EACA administration.

Drug-induced liver injury: Interactions between drug properties and host factors.

Idiosyncratic drug-induced liver injury (DILI) is a common cause for drug withdrawal from the market and although infrequent, DILI can result in serious clinical outcomes including acute liver failure and the need for liver transplantation. Eliminating the iatrogenic "harm" caused by a therapeutic intent is a priority in patient care. However, identifying culprit drugs and individuals at risk for DILI remains challenging. Apart from genetic factors predisposing individuals at risk, the role of the drugs' physicochemical and toxicological properties and their interactions with host and environmental factors need to be considered. The influence of these factors on mechanisms involved in DILI is multi-layered. In this review, we summarize current knowledge on 1) drug properties associated with hepatotoxicity, 2) host factors considered to modify an individuals' risk for DILI and clinical phenotypes, and 3) drug-host interactions. We aim at clarifying knowledge gaps needed to be filled in as to improve risk stratification in patient care. We therefore broadly discuss relevant areas of future research. Emerging insight will stimulate new investigational approaches to facilitate the discovery of clinical DILI risk modifiers in the context of disease complexity and associated interactions with drug properties, and hence will be able to move towards safety personalized medicine.

Effect of CKD and dialysis modality on exposure to drugs cleared by nonrenal mechanisms.

BACKGROUND: Patients with kidney disease frequently experience adverse effects from medication exposure, even when drugs are cleared by nonrenal pathways. Although many studies suggest that nonrenal drug clearance is decreased in chronic kidney disease (CKD), there remains a paucity of in vivo studies in patients with varying degrees of decreased kidney function and those comparing the impact of dialysis modality (eg, hemodialysis [HD] and peritoneal dialysis [PD]). STUDY DESIGN: We performed in vivo clinical pharmacokinetic studies of midazolam, a nonrenally cleared specific probe for CYP3A4, and fexofenadine, a nonspecific probe for hepatic and intestinal transporters. SETTING & PARTICIPANTS: Healthy controls (n=8), patients with non-dialysis-dependent (NDD)-CKD (n=8), and patients receiving HD (n=10) or PD (n=8). OUTCOMES: Exposure to midazolam and fexofenadine were quantified using area under the curve (AUC). Comprehensive pharmacokinetic parameters also were calculated for both probes. RESULTS: Midazolam AUC was significantly higher in the HD group (382.8 h·ng/mL) than in the healthy-control (63.0 h·ng/mL; P<0.001), NDD-CKD (84.5 h·ng/mL; P=0.002), and PD (47.4 h·ng/mL; P<0.001) groups. Fexofenadine AUC was significantly higher in each of the NDD-CKD (2,950 h·ng/mL; P=0.003), HD (2,327 h·ng/mL; P=0.01), and PD (2,095 h·ng/mL; P=0.04) groups compared with healthy controls (1,008 h·ng/mL). LIMITATIONS: Small study groups had different proportions of diabetic patients, early stages of CKD not available. CONCLUSIONS: Our data suggest that selection of dialysis modality is a major determinant of exposure to the CYP3A4 probe midazolam. Exposure to the intestinal and hepatic transporter probe fexofenadine is altered in patients with NDD-CKD and PD and HD patients. Thus, drug development and licensing of nonrenally cleared drugs should include evaluation in these 3 patient groups, with these results included in approved product information labeling. This reinforces the critical need for more in vivo studies of humans that evaluate the exposure to drugs cleared by these pathways.

Extended duration of prehydration does not prevent nephrotoxicity or delayed drug elimination in high-dose methotrexate infusions: a prospectively randomized cross-over study.

BACKGROUND: Alkalized hydration is used as supportive care to prevent renal toxicity during infusions with high-dose methotrexate (HDMTX). In children with acute lymphoblastic leukemia (ALL), the hydration is commonly initiated 4 hours before start of the methotrexate (MTX) infusion. To test if longer duration of prehydration would prevent MTX-induced renal toxicity, we preformed a randomized cross-over study comparing 12-4 hours of hydration before the infusion of HDMTX. PROCEDURES: Children with ALL and non-Hodgkin lymphoma that were treated with infusions of HDMTX 5 or 8 g/m(2) were randomized to receive intravenous prehydration 12 or 4 hours before the first HDMTX infusion. Patients alternated between 12 and 4 hours of prehydration in the subsequent HDMTX infusions. Renal toxicity was defined as 50% increase in plasma creatinine after the HDMTX infusion. The plasma MTX concentration was measured during and after the HDMTX infusion to determine if the duration of prehydration would influence the systemic MTX clearance. RESULTS: A total of 47 patients (224 HDMTX infusions) with a median age of 4.9 years were included in the study. The duration of prehydration had no effect on MTX induced renal toxicity that occurred in 18.5% of all HDMTX 5 g/m(2) infusions and in 40.0% of all HDMTX 8 g/m(2) infusions. Similar the duration of prehydration had no impact on the systemic clearance of MTX. CONCLUSION: Extending prehydration beyond 4 hours does not reduce the risk of renal toxicity or delayed MTX clearance after infusions with HDMTX 5-8 g/m(2).

The effect of pregnancy-related hormones on hepatic transporters: studies with premenopausal human hepatocytes.

Introduction: Pregnancy results in significant changes in drug pharmacokinetics (PK). While previous studies have elucidated the impact of pregnancy-related hormones (PRH) on mRNA or protein expression and activity of major hepatic metabolizing enzymes, their effect on hepatic drug transporters remains largely unexplored. Therefore, we investigated the effect of a cocktail of PRH on the mRNA expression and activity of hepatic transporters. Methods: Plated human hepatocytes (PHH) from 3 premenopausal donors were incubated, in triplicate, for 72 h, with vehicle (DMSO < 0.01%), rifampin (10 µM; positive control) or a cocktail of PRH consisting of estrone, estradiol, estriol, estetrol, progesterone, cortisol, testosterone, oxytocin, and placental growth hormone. The PRH concentrations replicated 0.1×, 1×, or 10× of the plasma concentrations of these hormones observed during each of the three trimesters of pregnancy. After treatment, mRNA expression (quantified by qPCR) of hepatic influx and efflux transporters as well as the activity of influx transporters was quantified (uptake of a selective substrate ± corresponding transporter inhibitor). The data were expressed relative to that in the control (vehicle) group. Significance was evaluated by ANOVA (followed by Dunn's multiple comparisons) or unpaired t-test when the within-lot data were analyzed, or repeated measures ANOVA (followed by Dunn's multiple comparisons) or paired t-test when data from all 3 lots were analyzed (p < 0.05). Results and Discussion: In general, a) PRH cocktails significantly induced transporter mRNA expression in the following order OAT2 ≈ NTCP ≈ OCT1 > OATP2B1 and repressed mRNA expression in the following order OATP1B3 > OATP1B1; b) these changes translated into significant induction of OAT2 (T1-T3) and NTCP (T2-T3, in only two lots) activity at the 1× PRH concentration. Compared with the influx transporters, the induction of mRNA expression of efflux transporters was modest, with mRNA expression of MRP2 and BSEP being induced the most. Conclusion: Once these data are verified through in vivo probe drug PK studies in pregnancy, they can be populated into physiologically based pharmacokinetic (PBPK) models to predict, for all trimesters of pregnancy, transporter-mediated clearance of any drug that is a substrate of the affected transporters.

Functionalized chitosan for cancer nano drug delivery.

Chitosan is a biotechnological derivative of chitin receiving a widespread pharmaceutical and biomedical applications. It can be used to encapsulate and deliver cancer therapeutics with inherent pH-dependent solubility to confer drug targeting at tumour microenvironment and anti-cancer activity synergizing cancer cytotoxic drug actions. To further reduce the off-target and by-stander adverse effects of drugs, a high targeted drug delivery efficiency at the lowest possible drug doses is clinically required. The chitosan has been functionalized with covalent conjugates or complexes and processed into nanoparticles to encapsulate and control drug release, to avoid premature drug clearance, to deliver drugs passively and actively to cancer site at tissue, cell or subcellular levels, and to promote cancer cell uptake of nanoparticles through membrane permeabilization at higher specificity and scale. Nanomedicine developed using functionalized chitosan translates to significant preclinical improvements. Future challenges related to nanotoxicity, manufacturability, selection precision of conjugates and complexes as a function of cancer omics and their biological responses from administration site to cancer target need critical assessments.

Extracellular Vesicles as Surrogates for Drug Metabolism and Clearance: Promise vs. Reality.

Drug-metabolizing enzymes (DMEs) and transporters play a major role in drug efficacy and safety. They are regulated at multiple levels and by multiple factors. Estimating their expression and activity could contribute to predicting drug pharmacokinetics and their regulation by drugs or pathophysiological situations. Determining the expression of these proteins in the liver, intestine, and kidney requires the collection of biopsy specimens. Instead, the isolation of extracellular vesicles (EVs), which are nanovesicles released by most cells and present in biological fluids, could deliver this information in a less invasive way. In this article, we review the use of EVs as surrogates for the expression and activity of DMEs, uptake, and efflux transporters. Preliminary evidence has been provided for a correlation between the expression of some enzymes and transporters in EVs and the tissue of origin. In some cases, data obtained in EVs reflect the induction of phase I-DMEs in the tissues. Further studies are required to elucidate to what extent the regulation of other DMEs and transporters in the tissues reflects in the EV cargo. If an association between tissues and their EVs is firmly established, EVs may represent a significant advancement toward precision therapy based on the biotransformation and excretion capacity of each individual.

Predicting the Effect of Renal Function on Systemic Clearance: Is a Simple Scaling Method Sufficient?

PURPOSE: To employ a simple scaling method to predict systemic or oral clearance for drugs that are primarily renally cleared knowing the fraction eliminated in urine (fe) and a patient's renal function relative to healthy controls (SGFR). METHODS: Observations evaluating drug clearance as a function of creatinine clearance for renally cleared drugs (fe >0.3) were obtained from literature sources. The analysis comprised of 82 unique drugs from 124 studies including 31 drugs with replicate studies. A simple scaler for renal function was employed and compared to the linear regression of available data. For drugs in which replicate studies were available, the ability of the linear regression (Cl vs ClCR) from one pharmacokinetic study was used to predict observations from an assigned replicate and compared to the scaling approach. RESULTS: For patients categorized as severe kidney disease (ClCR fixed at 20 ml/min), the scalar tended to over predict some observations, but 92% of the predictions were within 50-200% of the observed data. For drugs with available replicates, the scalar was as good or better in predicting the influence of ClCR on systemic clearance from a separate study when comparing against the linear regression approach. CONCLUSION: A scaling approach to account for alterations in drug clearance appears to have its advantages and represents a simple and generalizable method for guiding dose adjustments in patients with decreased renal function for drugs that are renally cleared (fe >0.3). In addition to its use in clinical practice, validation of this approach may have implications in facilitating more efficient drug development processes for designing dose-adjusted pharmacokinetic studies in patients with renal disease.

Endogenous markers of kidney function and renal drug clearance processes of filtration, secretion, and reabsorption.

The kidneys are responsible for maintaining physiologic homeostasis. The kidneys clear a variety of drugs and other substances through passive (filtration) and active processes that utilize transport proteins. Renal clearance is comprised of the processes of glomerular filtration, tubular secretion, and tubular reabsorption. Endogenous biomarkers, such as creatinine and cystatin C, are routinely used to estimate renal clearance. Understanding the contributing components of renal function and clearance, through the use of biomarkers, is necessary in elucidating the renal pharmacology of drugs and other substances. While exogenous markers of kidney function have been known for decades, several complexities have limited their usage. Several endogenous markers are being evaluated and hold promise to elucidate the individual components of kidney function that represent filtration, secretion, and reabsorption.