Metabolic Clearance Of Drugs Research Articles

Correction to "Dissecting Parameters Contributing to the Underprediction of Aldehyde Oxidase-Mediated Metabolic Clearance of Drugs".

Correction to "Dissecting Parameters Contributing to the Underprediction of Aldehyde Oxidase-Mediated Metabolic Clearance of Drugs".

Dissecting Parameters Contributing to the Underprediction of Aldehyde Oxidase-Mediated Metabolic Clearance of Drugs.

We investigated the effect of variability and instability in aldehyde oxidase (AO) content and activity on the scaling of in vitro metabolism data. AO content and activity in human liver cytosol (HLC) and five recombinant human AO preparations (rAO) were determined using targeted proteomics and carbazeran oxidation assay, respectively. AO content was highly variable as indicated by the relative expression factor (REF; i.e., HLC to rAO content) ranging from 0.001 to 1.7 across different in vitro systems. The activity of AO in HLC degrades at a 10-fold higher rate in the presence of the substrate as compared with the activity performed after preincubation without substrate. To scale the metabolic activity from rAO to HLC, a protein-normalized activity factor (pnAF) was proposed wherein the activity was corrected by AO content, which revealed up to sixfold higher AO activity in HLC versus rAO systems. A similar value of pnAF was observed for another substrate, ripasudil. Physiologically based pharmacokinetic (PBPK) modeling revealed a significant additional clearance (CL; 66%), which allowed for the successful prediction of in vivo CL of four other substrates, i.e., O-benzyl guanine, BIBX1382, zaleplon, and zoniporide. For carbazeran, the metabolite identification study showed that the direct glucuronidation may be contributing to around 12% elimination. Taken together, this study identified differential protein content, instability of in vitro activity, role of additional AO clearance, and unaccounted metabolic pathways as plausible reasons for the underprediction of AO-mediated drug metabolism. Consideration of these factors and integration of REF and pnAF in PBPK models will allow better prediction of AO metabolism. SIGNIFICANCE STATEMENT: This study elucidated the plausible reasons for the underprediction of aldehyde oxidase (AO)-mediated drug metabolism and provided recommendations to address them. It demonstrated that integrating protein content and activity differences and accounting for the loss of AO activity, as well as consideration of extrahepatic clearance and additional pathways, would improve the in vitro to in vivo extrapolation of AO-mediated drug metabolism using physiologically based pharmacokinetic modeling.

Evaluation of Renal Impairment Influence on Metabolic Drug Clearance using a Modelling Approach.

Chronic kidney disease (CKD) may alter drug renal elimination but is also known for interacting with hepatic metabolism via multiple uremic components. However, few global models, considering the five major cytochromes, have been published, and none specifically address the decrease in cytochrome P450 (CYP450) activity. The aim of our study was to estimate the possibility of quantifying residual cytochrome activity as a function of filtration rate, according to the data available in the literature. For each drug in the DDI-predictor database, we collected available pharmacokinetic data comparing drug exposition in the healthy patient and in various stages of CKD, before building a model capable of predicting the variation of exposure according to the degree of renal damage. We followed an In vivo Mechanistic Static Model (IMSM) approach, previously validated for predicting change in liver clearance. We estimated the remaining fraction parameters at glomerular filtration rate (GFR) = 0 and the alpha value of GFR to 50% impairment for the 5 major cytochromes using a non-linear constrained regression using Matlab software. Thirty-one compounds had usable pharmacokinetic data, with 51 AUC ratios between healthy and renal impaired patients. The remaining CYP3A4 activity was estimated to be 0.4 when CYP2D6, 2C9, 2C19 and 1A2 activity was estimated to be 0.43; 1; 0.73 and 0.7, respectively. The alpha value was estimated to be at 6.62; 25; 9.8; 1.38 and 11.04 for each cytochrome. In comparison with published data, all estimates but one were correctly predicted in the range of 0.5-2. Our approach was able to describe the impact of CKD on metabolic elimination. Modelling this process makes it possible to anticipate changes in clearance and drug exposure in CKD patients, with the advantage of greater simplicity than approaches based on physiologically-based pharmacokinetic modelling. However, a precise estimation of the impact of renal failure is not possible with an IMSM approach due to the large variability of the published data, and thus should rely on specific pharmacokinetic modelling for narrow therapeutic margin drugs.

Hepatic Scaling Factors for In Vitro-In Vivo Extrapolation of Metabolic Drug Clearance in Patients with Colorectal Cancer with Liver Metastasis.

In vitro-in vivo extrapolation (IVIVE) linked with physiologically based pharmacokinetics (PBPK) modeling is used to predict the fates of drugs in patients. Ideally, the IVIVE-PBPK models should incorporate systems information accounting for characteristics of the specific target population. There is a paucity of such scaling factors in cancer, particularly microsomal protein per gram of liver (MPPGL) and cytosolic protein per gram of liver (CPPGL). In this study, cancerous and histologically normal liver tissue from 16 patients with colorectal liver metastasis were fractionated to microsomes and cytosol. Protein content was measured in homogenates, microsomes, and cytosol. The loss of microsomal protein during fractionation was accounted for using corrections based on NADPH cytochrome P450 reductase activity in different matrices. MPPGL was significantly lower in cancerous tissue (24.8 ± 9.8 mg/g) than histologically normal tissue (39.0 ± 13.8 mg/g). CPPGL in cancerous tissue was 42.1 ± 12.9 mg/g compared with 56.2 ± 16.9 mg/g in normal tissue. No correlations between demographics (sex, age, and body mass index) and MPPGL or CPPGL were apparent in the data. The generated scaling factors together with assumptions regarding the relative volumes of cancerous versus noncancerous tissue were used to simulate plasma exposure of drugs with different extraction ratios. The PBPK simulations revealed a substantial difference in drug exposure (area under the curve), up to 3.3-fold, when using typical scaling factors (healthy population) instead of disease-related parameters in cancer population. These indicate the importance of using population-specific scalars in IVIVE-PBPK for different disease states. SIGNIFICANCE STATEMENT: Accuracy in predicting the fate of drugs from in vitro data using IVIVE-PBPK depends on using correct scaling factors. The values for two of such scalars, namely microsomal and cytosolic protein per gram of liver, is not known in patients with cancer. This study presents, for the first time, scaling factors from cancerous and matched histologically normal livers. PBPK simulations of various metabolically cleared drugs demonstrate the necessity of population-specific scaling for model-informed precision dosing in oncology.

Effective Renal Clearance and Photothermal Therapy of a Cyclodextrin-Modified Quaterrylene Derivative.

Metabolic clearance of drugs from the body is a key element for future clinical applications. Compared with the liver-bile metabolic route, the renal-urinary route is the most desirable and efficient clearance pathway for practical use in clinical trials and can effectively decrease the long-term risk of unpredictable intracellular catabolism and the potential toxicity to normal organs. However, renal clearable photothermal therapy (PTT) agents were limited. In this paper, a permethyl-β-cyclodextrin-modified quaterrylene bisimide derivative (QDI-CD) was developed. QDI-CD as a biocompatible molecule could effectively be cleared with the renal clearance efficiency of (67.00 ± 2.37)% at 24 h by the renal-urinary route when administered by tail vein injection. Importantly, the structure of QDI-CD in urine after metabolism does not change. Furthermore, QDI-CD, as a potent photoacoustic and photothermal agent, could effectively be enriched in tumor tissue after 4 h of injection and showed the effective PTT in mice. This work developed a potent organic PTT agent with good renal clearance from the body and with promise for future clinical applications.

Highly Variable Expression of CYP1A1 in Human Liver and Impact on Pharmacokinetics of Riociguat and Granisetron in Humans.

Cytochrome P450s (CYPs), a superfamily of enzymes, are involved in the biotransformation of endogenous and xenobiotic chemicals and mainly responsible for the metabolic clearance of widely prescribed drugs. Out of the 57 human isoforms, only a few, most notably CYP3A4, are considered to be important in this process. CYP1A1, one of the three isoforms of the CYP1 family, is widely believed to play an important role in the metabolism and activation of numerous procarcinogens, e.g., polyaromatic hydrocarbons (PAHs) or aromatic amines. It is also known that CYP1A1 is highly inducible by endogenous and exogenous factors, e.g., PAHs. However, CYP1A1 has not been considered to play a significant role in the metabolic clearance of drugs, since this isoform has been detected only in extrahepatic tissues in small amounts. In contrast to conventional wisdom, we herein demonstrate the expression of CYP1A1 protein in human liver microsomal preparations. The expression levels of CYP1A1 were quantified by Western blot and LC/MS analyses and corresponded well with enzymatic activities of highly selective CYP1A1 reactions. In a panel of 29 individual liver microsomal preparations, highly variable and substantial expression levels (up to ∼10 pmol/mg) were measured. Together with the high selectivity and especially the high metabolic efficiency of CYP1A1 shown for granisetron and riociguat, it is demonstrated that CYP1A1 plays an important role in the metabolic clearance of these drugs and is responsible for the clinically observed interindividual variability in their pharmacokinetics. Therefore, the importance of CYP1A1 in drug discovery and development needs to be reconsidered.

Recovery of OATP1B Activity after Living Kidney Transplantation in Patients with End-Stage Renal Disease.

Recently, several studies have shown that renal failure decreases the metabolic clearance of drugs and the transportation capability of some drug transporters. However, whether organic anion transporting polypeptide (OATP)1B activities decrease in renal failure remains unknown. In this study, we measured plasma concentrations of coproporphyrin-I (CP-I), a specific endogenous OATP1B probe, in patients with end stage renal disease before and after living kidney transplantation and evaluated the effect of renal function on OATP1B activity. This prospective study recruited 13 patients with end-stage renal disease. Plasma CP-I concentrations were measured before and 7, 14, 30 and 90days after living kidney transplantation. Plasma CP-I concentrations decreased over time after living kidney transplantation and showed significant difference on day 90 compared with before living kidney transplantation [1.12 ± 0.59 vs 0.65 ± 0.27ng/mL, p < 0.05 (95% CI of difference - 0.927, -0.013)]. A significant negative correlation was observed between estimated glomerular filtration rate and plasma CP-I concentration (r = -0.30, p < 0.05), suggesting recovery of OATP1B activity with improvement in renal function. OATP1B activity may decrease in renal failure and dose adjustment of OATP1B substrates may be needed in patients with renal failure.

The nested enzyme-within-enterocyte (NEWE) turnover model for predicting dynamic drug and disease effects on the gut wall

Physiologically-based pharmacokinetic (PBPK) models provide a framework for in vitro-in vivo extrapolation of metabolic drug clearance. Many of the concepts in PBPK can have consequential impact on more mechanistic systems pharmacology models. In the gut wall, turnover of enzymes and enterocytes are typically lumped into one rate constant that describes the time dependent enzyme activity. This assumption may influence predictability of any sustained and dynamic effects such as mechanism-based inhibition (MBI), particularly when considering translation from healthy to gut disease.A novel multi-level systems PBPK model was developed. This model comprised a ‘nested enzyme-within enterocyte’ (NEWE) turnover model to describe levels of drug-metabolising enzymes. The ability of the model to predict gut metabolism following MBI and gut disease was investigated and compared to the conventional modelling approach.For MBI, the default NEWE model performed comparably to the conventional model. However, when drug-specific spatial crypt-villous absorption was considered, up to approximately 50% lower impact of MBI was simulated for substrates highly metabolised by cytochrome P450 (CYP) 3A4, interacting with potent inhibitors. Further, the model showed potential in predicting the disease effect of gastrointestinal mucositis and untreated coeliac disease when compared to indirect clinical pharmacokinetic parameters.Considering the added complexity of the NEWE model, it does not provide an attractive solution for improving upon MBI predictions in healthy individuals. However, nesting turnover may enable extrapolation to gut disease-drug interactions. The principle detailed herein may be useful for modelling drug interactions with cellular targets where turnover is significant enough to affect this process.

Application of novel 3D-bioprinted human liver tissue models in drug discovery research

Primary human hepatocytes are widely used to study drug metabolism and enzyme induction. However, primary hepatocytes rapidly lose their hepatic function in conventional 2D cultures. Recently, a microphysiological system that overcomes this drawback has been actively investigated and applied in drug discovery research. Such novel in vitro models are desirable for the evaluation of the metabolic clearance of drugs with low turnover, drug-induced liver injury, and chronic liver diseases like liver fibrosis. This article reviews the characteristics and recent advances in 3D-bioprinted human liver tissue models in drug discovery research.

123I-iomazenil whole-body imaging to detect hepatic carboxylesterase drug-metabolizing enzyme activity.

Drugs are mainly metabolized by hepatic enzymes, the activity of which can differ between individuals. Although it is ideal to measure the hepatic clearance of liver-targeted drugs in individualized medicine, blood enzyme tests typically measure metabolic drug clearance in the entire body, and not just in the liver. We investigated whether I-iomazenil imaging can directly assess and quantify the activity of hepatic drug-metabolizing enzymes. Hepatic enzymes that metabolize I-iomazenil were identified by thin-layer chromatography in mouse liver homogenates with bis(4-nitrophenyl) phosphate (BNPP) inhibitor for carboxylesterase enzymes and nicotinamide adenine dinucleotide phosphate (NADPH) generator for cytochrome P450 enzymes. Whole-body images of mice were acquired using I-iomazenil with and without BNPP, and the distribution was also obtained. The metabolism of I-iomazenil in the blood, liver, gall bladder, and bladder was investigated by thin-layer chromatography. From the in-vitro metabolism of I-iomazenil using BNPP, the enzyme converting I-iomazenil to I-R-COOH was identified as carboxylesterase, and that converting I-iomazenil to M2 was identified as cytochrome P450 in experiments with and without an NADPH generator. The biological distribution and whole-body imaging showed increased accumulation in the liver of mice administered BNPP compared with normal mice, but decreased levels in the gall bladder and small intestine. The main fraction in bile and urine was I-R-COOH, with two unknown metabolites (M1 and M2), I, and I-iomazenil also being present. I-iomazenil whole-body imaging has good possibility of direct measurement of hepatic carboxylesterase activity as accumulation of I-R-COOH in the gall bladder through bile and in the bladder through urine.

Microfluidic Lateral Flow Cytochrome P450 Assay on a Novel Printed Functionalized Calcium Carbonate‐Based Platform for Rapid Screening of Human Xenobiotic Metabolism

AbstractCytochrome P450 (CYP) is a superfamily of enzymes in charge of elimination of the majority of clinically used drugs and other xenobiotics. This study focuses on the development of a rapid microfluidic lateral flow assay to study human phase I metabolism reactions mediated by CYP2A6 isoenzyme, the major detoxification route for many known carcinogens and drugs, with coumarin 7‐hydroxylation, as the prototype model reaction. Assay fabrication utilizes custom‐designed porous functionalized calcium carbonate (FCC) coatings and inkjet‐printed fluid barriers. All materials used are novel and carefully chosen to preserve biocompatibility. The design comprises separate zones for reaction, separation and detection, and an absorbent pad to keep the assay wet for extended periods (up to 10 min) even when heated to physiological temperature. The concept enables CYP assays to be made at lower cost than conventional well‐plate assays, while providing increased selectivity at equally high speed, owing to the possibility for simultaneous chromatographic separation of the reaction products from the reactants on the FCC coating. The developed concept provides a viable rapid prediction of the interaction risks related to metabolic clearance of drugs and other xenobiotics, and exemplifies a novel coating technology illustrating the opportunity to broaden application functionality.

Prediction of metabolic drug clearance in humans

Prediction of metabolic drug clearance in humans

New Ways of Detecting ADRs in Neonates and Children.

Severe adverse drug reactions (ADRs) cause 5-7% of all hospital admissions, an estimated 2,000,000 severe reactions, and over 100,000 deaths each year in the USA. A recent systematic review indicated that the overall incidence of ADRs was 11% in hospitalized children and 1% in outpatients. Detecting ADRs in neonates and children is challenging, particularly because there are fewer clinical trials involving children than adults and drug use in children is common without a labeled indication. Ontogeny and significant physiological changes related to age have an impact on metabolic drug clearance and pharmacodynamics in children compared to adults, as well as on drug action. A variety of strategies have been developed for the identification and further evaluation of ADRs, starting from case reports and advancing into more structured methodologies, such as active surveillance, for accumulating the necessary information. While each approach has merit, a comprehensive surveillance approach with different methods is required to monitor drug safety in the post-market period. Among the methods that have shown value in neonates and children are anecdotal reporting, voluntary organized reporting, prescription event monitoring, pharmacoepidemiology using administrative databases, and active surveillance. There is an urgent need to improve the evaluation of paediatric drug safety in the pre- and post-market phases of drug evaluation in order to better predict in whom serious harm may occur and better ensure the safe use of drugs for neonates and children.

Selective Inhibition of Cytochrome P450 2D6 by Sarpogrelate and Its Active Metabolite, M-1, in Human Liver Microsomes

The present study was performed to evaluate the in vitro inhibitory potential of sarpogrelate and its active metabolite, M-1, on the activities of nine human cytochrome (CYP) isoforms. Using a cocktail assay, the effects of sarpogrelate on nine CYP isoforms and M-1 were measured by specific marker reactions in human liver microsomes. Sarpogrelate potently and selectively inhibited CYP2D6-mediated dextromethorphan O-demethylation with an IC50 (Ki) value of 3.05 μM (1.24 μM), in a competitive manner. M-1 also markedly inhibited CYP2D6 activity; its inhibitory effect with an IC50 (Ki) value of 0.201 μM (0.120 μM) was more potent than that of sarpogrelate, and was similarly potent as quinidine (Ki, 0.129 μM), a well-known typical CYP2D6 inhibitor. In addition, sarpogrelate and M-1 strongly inhibited both CYP2D6-catalyzed bufuralol 1'-hydroxylation and metoprolol α-hydroxylation activities. However, sarpogrelate and M-1 showed no apparent inhibition of the other following eight CYPs: CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2E1, or CYP3A4/5. Upon 30-minute preincubation of human liver microsomes with sarpogrelate or M-1 in the presence of NADPH, no obvious shift in IC50 was observed in terms of inhibition of the nine CYP activities, suggesting that sarpogrelate and M-1 are not time-dependent inactivators. Sarpogrelate strongly inhibited the activity of CYP2D6 at clinically relevant concentrations in human liver microsomes. These observations suggest that sarpogrelate could have an effect on the metabolic clearance of drugs possessing CYP2D6-catalyzed metabolism as a major clearance pathway, thereby eliciting pharmacokinetic drug-drug interactions.

Significant increase in plasma 4β-hydroxycholesterol concentration in patients after kidney transplantation

Several previous studies have shown that renal failure decreases not only renal elimination but also metabolic clearance of drugs, particularly those metabolized by CYP3A. However, whether recovery of renal function results in recovery of hepatic CYP3A activity remains unknown. In this study, we evaluated the effect of renal function on CYP3A activity after kidney transplantation in patients with end-stage renal disease (ESRD) by measuring the change in CYP3A activity using plasma concentration of 4β-hydroxycholesterol as a biomarker. The study enrolled 13 patients with ESRD who underwent the first kidney allograft transplantation. Morning blood samples were collected before and 3, 7, 10, 14, 21, 30, 60, 90, 120, 150 and 180 days after kidney transplantation. Plasma concentration of 4β-hydroxycholesterol was measured using GC-MS. Compared with before kidney transplantation, creatinine clearance increased significantly from day 3 after kidney transplantation and stabilized thereafter. Plasma concentration of 4β-hydroxycholesterol was elevated significantly on days 90 and 180 after kidney transplantation. In conclusion, this study suggests the recovery of CYP3A activity with improvement in renal function after kidney transplantation in patients with ESRD.

Predicted metabolic drug clearance with increasing adult age.

To determine the effect of increasing adult age on predicted metabolic drug clearance. Predicted metabolic drug clearances (CLPT ) were determined using in vitro-in vivo extrapolation coupled with physiological-based pharmacokinetic modelling and simulation (IVIVE-PBPK) in Simcyp®. Simulations were conducted using CYP-selective 'probe' drugs with subjects in 5 year age groups (20-25 to 90-95 years). CLPT values were compared with human pharmacokinetic data stratified according to age (young = 20-40 years and elderly = 65-85 years) and gender. Age-related changes in the physiological parameters used for IVIVE of CLPT were described. Predicted metabolic drug clearances decreased with increasing adult age to approximately 65-70 years: caffeine from 1.5 to 1.0 ml min(-1) kg(-1) (a 33% decrease), S-warfarin from 0.100 to 0.064 ml min(-1) kg(-1) (36%), S-mephenytoin from 4.1 to 2.5 ml min(-1) kg(-1) (39%), desipramine from 10.6 to 7.3 ml min(-1) kg(-1) (31%) and midazolam from 5.4 to 3.9 ml min(-1) kg(-1) (27%). Except for S-mephenytoin, predictions were within 3.5-fold of clearances from clinical studies when stratified by age and gender. A trend towards higher CLPT was observed in females, but this was only statistically significant in larger virtual trials. Physiological parameters that determine CLPT decreased with increasing adult age: mean microsomal protein g(-1) of liver, liver weight, hepatic blood flow and human serum albumin concentration. Decreased metabolic clearance in the elderly was predicted by Simcyp® and was generally consistent with limited clinical data for four out of five drugs studied and the broader literature for drugs metabolized by CYP enzymes. IVIVE-PBPK may be increasingly useful in predicting metabolic drug clearance in the elderly.

Comparative Assessment of In Vitro–In Vivo Extrapolation Methods used for Predicting Hepatic Metabolic Clearance of Drugs

The purpose of this study was to perform a comparative analysis of various in vitro--in vivo extrapolation (IVIVE) methods used for predicting hepatic metabolic clearance (CL) of drugs on the basis of intrinsic CL data determined in microsomes. Five IVIVE methods were evaluated: the "conventional and conventional bias-corrected methods" using the unbound fraction in plasma (fu(p) ), the "Berezhkovskiy method" in which the fu(p) is adjusted for drug ionization, the "Poulin et al. method" using the unbound fraction in liver (fu(liver) ), and the "direct scaling method," which does not consider any binding corrections. We investigated the effects of the following scenarios on the prediction of CL: the use of preclinical or human datasets, the extent of plasma protein binding, the magnitude of CL in vivo, and the extent of drug disposition based on biopharmaceutics drug disposition classification system (BDDCS) categorization. A large and diverse dataset of 139 compounds was collected, including those from the literature and in house from Genentech. The results of this study confirm that the Poulin et al. method is robust and showed the greatest accuracy as compared with the other IVIVE methods in the majority of prediction scenarios studied here. The difference across the prediction methods is most pronounced for (a) albumin-bound drugs, (b) highly bound drugs, and (c) low CL drugs. Predictions of CL showed relevant interspecies differences for BDDCS class 2 compounds; the direct scaling method showed the greatest predictivity for these compounds, particularly for a reduced dataset in rat that have unexpectedly high CL in vivo. This result is a reflection of the direct scaling method's natural tendency to overpredict the true metabolic CL. Overall, this study should facilitate the use of IVIVE correlation methods in physiologically based pharmacokinetics (PBPK) model.

The effect and clinical consequences of hypoxia on cytochrome P450, membrane carrier proteins activity and expression

Introduction: Chronic pulmonary disease and heart failure reduce drug clearance and consequently enhance adverse drug reactions. The mechanisms of action underlying the regulation of cytochrome P450 (CYP) isoforms and membrane carrier proteins by hypoxia, and the clinical consequences of the regulation of CYP by hypoxia, alone or combined with other conditions have been elucidated in the last decades. Overall, a reduced drug clearance appears to be associated with hypoxemia.Areas covered: In this review, the mechanisms of action underlying hypoxia-induced regulation of CYP enzymes are discussed. The authors also revise the effects of hypoxia on serum mediators, signal transduction pathways, orphan nuclear receptors, transcription factors and post-transcriptional mechanisms regulating CYP and membrane carrier proteins expression. Additionally, the paper also discusses the clinical repercussions of hypoxia-induced changes in CYP and membrane carrier proteins activity.Expert opinion: Acute systemic hypoxia down-regulates selected CYP isoforms and up-regulates CYP3A4 and P-glycoprotein, changing the metabolic clearance of drugs and endogenous compounds biotransformed by these isoforms as well as the kinetics. In patients with acute hypoxia, the dosage of drugs, biotransformed by CYP isoforms, may need to be adjusted. Tissue hypoxia enhances the expression of efflux membrane carrier proteins, increasing the probability of drug resistance.

Spironolactone and Canrenone Inhibit UGT2B7-Catalyzed Human Liver and Kidney Microsomal Aldosterone 18β-Glucuronidation: A Potential Drug Interaction

Elevated plasma concentrations of aldosterone (ALDO) are observed in patients treated with spironolactone. Because ALDO is eliminated via UGT2B7-catalyzed 18beta-glucuronidation, this study aimed to determine whether spironolactone and its primary metabolites, canrenone and canrenoic acid, inhibit ALDO 18beta-glucuronidation by recombinant UGT2B7 and by human liver (HLM) and human kidney cortical (HKCM) microsomes. Initial experiments characterized the effects of all three compounds on 4-methylumbelliferone and ALDO glucuronidation by recombinant human UGT2B7. IC(50) values for spironolactone and canrenone ranged from 26 to 50 microM, whereas canrenoic acid was a weak inhibitor. Inhibitor constant (K(i)) values for spironolactone and canrenone inhibition of ALDO 18beta-glucuronidation were subsequently determined with HLM, HKCM, and UGT2B7 as the enzyme sources. Spironolactone and canrenone were competitive inhibitors of ALDO 18beta-glucuronidation by HLM, HKCM, and UGT2B7. Mean (+/-) K(i) values for spironolactone were 52 +/- 22 (HLM) and 34 +/- 4 microM (HKCM), and mean (+/-) K(i) values for canrenone were 41 +/- 19 (HLM) and 23 +/- 2 microM (HKCM). K(i) values for spironolactone and canrenone inhibition of ALDO 18beta-glucuronidation by recombinant UGT2B7 were 23 and 11 microM, respectively. "Actual" K(i) values for spironolactone and canrenone inhibition of ALDO 18beta-glucuronidation, which take into account the role of endogenous microsomal inhibitors, are predicted to be 3 to 5 and 2 to 4 microM, respectively. The data indicate that the elevated ALDO concentrations observed in patients treated with spironolactone may be due, at least in part, to a pharmacokinetic interaction, and spironolactone and canrenone should be considered to be potential inhibitors of the UGT2B7-mediated metabolic clearance of drugs in both liver and kidney.

Free Drug Metabolic Clearance in Elderly People

The question of whether metabolic drug clearance is decreased in elderly people has been the subject of considerable debate and is very important because clearance is a determinant of dosing. Drug clearance has been shown to be consistently impaired for flow-limited (high-clearance) drugs, but there have been conflicting results for capacity-limited (low-clearance) drugs. A limitation of the studies of capacity-limited drugs is that most have estimated clearance based on total drug concentrations (protein-bound plus free). Total drug clearance reflects both the intrinsic clearance of free drug and the extent of protein binding. Total clearance is a valid measure for capacity-limited drugs with low protein binding and appears to be consistently impaired in elderly subjects. For phenazone [antipyrine] (fraction unbound [f(u)] >0.9), seven studies have demonstrated statistical reductions in clearance of 20-52%. For theophylline (f(u) 0.6), five studies have demonstrated reductions in clearance of 22-35%. For paracetamol [acetaminophen] (f(u) 0.8), the clearance of which has been quoted as unchanged, four studies have demonstrated reductions in clearance of 19-35%. For highly protein-bound drugs, total clearance is not the appropriate parameter. Free drug clearance is more appropriate since it is independent of changes in protein binding. The literature was reviewed to test the hypothesis that in elderly people, capacity-limited drugs with high protein binding will show decreased free clearance even in the absence of a decrease in total clearance. For these drugs, data for free drug clearance based on measurement of actual free drug concentrations are limited, but suggest that the intrinsic metabolic clearance is impaired in elderly subjects. Four studies of naproxen (f(u) <0.01) have shown reduced free drug clearance of 50% or more. Two studies of valproic acid (f(u) 0.1-0.2) have shown reduced free clearance of 39% and 65%. Two studies of ibuprofen (f(u) <0.01) have shown reduced free clearance of S-ibuprofen of 21% and 28%. There is some indirect evidence for reduced clearance of the highly protein-bound drugs oxaprozin, temazepam, lorazepam, diazepam, phenytoin and warfarin, although studies measuring free concentrations are lacking. Together, the above studies support the hypothesis that the intrinsic metabolic drug clearance is impaired in elderly subjects, in the order of 20-60%, and that this effect is masked if highly protein-bound drugs are assessed using total drug clearance. If the findings are confirmed in future well-designed studies of free drug clearance, there are profound and beneficial implications for dosing of drugs in elderly people. Lower doses are likely to achieve appropriate concentrations, allowing full efficacy but decreased dose-related adverse effects.