Proteomic characterisation of perhexiline treatment on THP-1 M1 macrophage differentiation.

Background: Dysregulated inflammation is important in the pathogenesis of many diseases including cancer, allergy, and autoimmunity. Macrophage activation and polarisation are commonly involved in the initiation, maintenance and resolution of inflammation. Perhexiline (PHX), an antianginal drug, has been suggested to modulate macrophage function, but the molecular effects of PHX on macrophages are unknown. In this study we investigated the effect of PHX treatment on macrophage activation and polarization and reveal the underlying proteomic changes induced. Methods: We used an established protocol to differentiate human THP-1 monocytes into M1 or M2 macrophages involving three distinct, sequential stages (priming, rest, and differentiation). We examined the effect of PHX treatment at each stage on the polarization into either M1 or M2 macrophages using flow cytometry, quantitative polymerase chain reaction (qPCR) and enzyme linked immunosorbent assay (ELISA). Quantitative changes in the proteome were investigated using data independent acquisition mass spectrometry (DIA MS). Results: PHX treatment promoted M1 macrophage polarization, including increased STAT1 and CCL2 expression and IL-1ß secretion. This effect occurred when PHX was added at the differentiation stage of the M1 cultures. Proteomic profiling of PHX treated M1 cultures identified changes in metabolic (fatty acid metabolism, cholesterol homeostasis and oxidative phosphorylation) and immune signalling (Receptor Tyrosine Kinase, Rho GTPase and interferon) pathways. Conclusion: This is the first study to report on the action of PHX on THP-1 macrophage polarization and the associated changes in the proteome of these cells.

Study of the roles of cytochrome P450 (CYPs) in the metabolism and cytotoxicity of perhexiline.

Perhexiline is a prophylactic antianginal agent developed in the 1970s. Although, therapeutically, it remained a success, the concerns of its severe adverse effects including hepatotoxicity caused the restricted use of the drug, and eventually its withdrawal from the market in multiple countries. In the clinical setting, cytochrome P450 (CYP) 2D6 is considered as a possible risk factor for the adverse effects of perhexiline. However, the role of CYP-mediated metabolism in the toxicity of perhexiline, particularly in the intact cells, remains unclear. Using our previously established HepG2 cell lines that individually express 14 CYPs (1A1, 1A2, 1B1, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4, 3A5, and 3A7) and human liver microsomes, we identified that CYP2D6 plays a major role in the hydroxylation of perhexiline. We also determined that CYP1A2, 2C19, and 3A4 contribute to the metabolism of perhexiline. The toxic effect of perhexiline was reduced significantly in CYP2D6-overexpressing HepG2 cells, in comparison to the control cells. In contrast, overexpression of CYP1A2, 2C19, and 3A4 did not show a significant protective effect against the toxicity of perhexiline. Pre-incubation with quinidine, a well-recognized CYP2D6 inhibitor, significantly attenuated the protective effect in CYP2D6-overexpressing HepG2 cells. Furthermore, perhexiline-induced mitochondrial damage, apoptosis, and ER stress were also attenuated in CYP2D6-overexpressing HepG2 cells. These findings suggest that CYP2D6-mediated metabolism protects the cells from perhexiline-induced cytotoxicity and support the clinical observation that CYP2D6 poor metabolizers may have higher risk for perhexiline-induced hepatotoxicity.

The validation of an LC-MS/MS assay for perhexiline and major hydroxy metabolites, and its application to therapeutic monitoring in patient plasma.

AIM: Perhexiline (PEX), being developed to treat hypertrophic cardiomyopathy, is toxic at levels above the therapeutic range. Plasma level monitoring is therefore essential. The absence of a UV-absorbing chromophore has in the past required quantitative analysis of PEX in plasma using lengthy derivatization methods, followed by HPLC and fluorescence detection. The routine and urgent analysis of a large number of patient plasma samples necessitates faster and reliable analytical methodology. RESULTS: An LC-MS/MS method, using two novel internal standards, has been validated for the quantitative measurement of PEX and its major hydroxy metabolites in human plasma. CONCLUSION: The assay has been applied to therapeutic drug monitoring (TDM), where PEX and the ratio of the drug to cis-hydroxy perhexiline, were measured at designated intervals.

Development of Fluorinated Analogues of Perhexiline with Improved Pharmacokinetic Properties and Retained Efficacy.

We designed and synthesized perhexiline analogues that have the same therapeutic profile as the parent cardiovascular drug but lacking its metabolic liability associated with CYP2D6 metabolism. Cycloalkyl perhexiline analogues 6a-j were found to be unsuitable for further development, as they retained a pharmacokinetic profile very similar to that shown by the parent compound. Multistep synthesis of perhexiline analogues incorporating fluorine atoms onto the cyclohexyl ring(s) provided a range of different fluoroperhexiline analogues. Of these, analogues 50 (4,4-gem-difluoro) and 62 (4,4,4',4'-tetrafluoro) were highly stable and showed greatly reduced susceptibility to CYP2D6-mediated metabolism. In vitro efficacy studies demonstrated that a number of derivatives retained acceptable potency against CPT-1. Having the best balance of properties, 50 was selected for further evaluation. Like perhexiline, it was shown to be selectively concentrated in the myocardium and, using the Langendorff model, to be effective in improving both cardiac contractility and relaxation when challenged with high fat buffer.

Comparison of CYP2D metabolism and hepatotoxicity of the myocardial metabolic agent perhexiline in Sprague-Dawley and Dark Agouti rats.

1. Perhexiline, a chiral anti-anginal agent, may be useful to develop new cardiovascular therapies, despite its potential hepatotoxicity. 2. This study compared Dark Agouti (DA) and Sprague-Dawley (SD) rats, as models of perhexiline's metabolism and hepatotoxicity in humans. Rats (n = 4/group) received vehicle or 200 mg/kg/d of racemic perhexiline maleate for 8 weeks. Plasma and liver samples were collected to determine concentrations of perhexiline and its metabolites, hepatic function and histology. 3. Median (range) plasma and liver perhexiline concentrations in SD rats were 0.09 (0.04-0.13) mg/L and 5.42 (0.92-8.22) ng/mg, respectively. In comparison, DA rats showed higher (p < 0.05) plasma 0.50 (0.16-1.13) mg/L and liver 24.5 (9.40-54.7) ng/mg perhexiline concentrations, respectively, 2.5- and 3.7-fold higher cis-OH-perhexiline concentrations, respectively (p < 0.05), and lower plasma metabolic ratio (0.89 versus 1.55, p < 0.05). In both strains, the (+):(-) enantiomer ratio was 2:1. Perhexiline increased plasma LDH concentrations in DA rats (p < 0.05), but had no effect on plasma biochemistry in SD rats. Liver histology revealed lower glycogen content in perhexiline-treated SD rats (p < 0.05), but no effects on lipid content in either strain. 4. DA rats appeared more similar to humans with respect to plasma perhexiline concentrations, metabolic ratio, enantioselective disposition and biochemical changes suggestive of perhexiline-induced toxicity.

CYP2B6, CYP2D6, and CYP3A4 catalyze the primary oxidative metabolism of perhexiline enantiomers by human liver microsomes.

The cytochrome P450 (P450)-mediated 4-monohydroxylations of the individual enantiomers of the racemic antianginal agent perhexiline (PHX) were investigated in human liver microsomes (HLMs) to identify stereoselective differences in metabolism and to determine the contribution of the polymorphic enzyme CYP2D6 and other P450s to the intrinsic clearance of each enantiomer. The cis-, trans1-, and trans2-4-monohydroxylation rates of (+)- and (-)-PHX by human liver microsomes from three extensive metabolizers (EMs), two intermediate metabolizers (IMs), and two poor metabolizers (PMs) of CYP2D6 were measured with a high-performance liquid chromatography assay. P450 isoform-specific inhibitors, monoclonal antibodies directed against P450 isoforms, and recombinantly expressed human P450 enzymes were used to define the P450 isoform profile of PHX 4-monohydroxylations. The total in vitro intrinsic clearance values (mean +/- S.D.) of (+)- and (-)-PHX were 1376 +/- 330 and 2475 +/- 321, 230 +/- 225 and 482 +/- 437, and 63.4 +/- 1.6 and 54.6 +/- 1.2 microl/min/mg for the EM, IM, and PM HLMs, respectively. CYP2D6 catalyzes the formation of cis-OH-(+)-PHX and trans1-OH-(+)-PHX from (+)-PHX and cis-OH-(-)-PHX from (-)-PHX with high affinity. CYP2B6 and CYP3A4 each catalyze the trans1- and trans2-4-monohydroxylation of both (+)- and (-)-PHX with low affinity. Both enantiomers of PHX are subject to significant polymorphic metabolism by CYP2D6, although this enzyme exhibits distinct stereoselectivity with respect to the conformation of metabolites and the rate at which they are formed. CYP2B6 and CYP3A4 are minor contributors to the intrinsic P450-mediated hepatic clearance of both enantiomers of PHX, except in CYP2D6 PMs.

Determination of the 4-monohydroxy metabolites of perhexiline in human plasma, urine and liver microsomes by liquid chromatography.

The use of perhexiline (PHX) is limited by hepatic and neurological toxicity associated with elevated concentrations in plasma that are the result of polymorphism of the cytochrome P450 2D6 isoform (CYP2D6). PHX is cleared by hepatic oxidation that produces three 4-monohydroxy metabolites: cis-OH-PHX, trans1-OH-PHX and trans2-OH-PHX. The current study describes an HPLC-fluorescent method utilising pre-column derivatization with dansyl chloride. Following derivatization, the metabolites were resolved on a C18 column with a gradient elution using a mobile phase composed of methanol and water. The method described is suitable for the quantification of the metabolites in human plasma and urine following clinical doses and for kinetic studies using human liver microsomes. The method demonstrates sufficient sensitivity, accuracy and precision between 5.0 and 0.01, 50.0 and 0.2 and 1.0 and 0.005 mg/l in human plasma, urine and liver microsomes, respectively, with intra-assay coefficients of variation and bias <15%, except at the lowest limit of quantification (<20%). The inter-assay coefficients of variation and bias were <15%. The application of this method to plasma and urine samples of five CYP2D6 extensive metaboliser (EM) patients at steady state with respect to PHX dosing determined that the mean (+/-S.D.) renal clearances of trans1-OH-PHX and cis-OH-PHX were 1.58+/-0.35 and 0.16+/-0.06l/h, respectively. The mean (+/-S.D.) dose recovered in urine as free and glucuronidated 4-monohydroxy PHX metabolites was 20.6+/-11.6%.

Metabolic therapeutics in angina pectoris: history revisited with perhexiline.

The ever-increasing burden of ischaemic heart disease and its common manifestation chronic angina pectoris calls for the exploration of other treatment options for those patients who despite the maximum conventional pharmacological and surgical interventions continue to suffer. Such exploration has led to the increasing use of new metabolically acting antianginal agents and the re-emergence of an old and somewhat forgotten pharmacological agent, perhexiline maleate. This review aims to update the cardiac nurse with knowledge to manage the care a patient receiving perhexiline maleate treatment and provide a brief review of three new metabolic agents: trimetazidine, ranolazine and etomoxir.

Correlation of CYP2D6 genotype with perhexiline phenotypic metabolizer status.

Perhexiline is metabolized by CYP2D6 and has concentration-related hepatoxicity and peripheral neuropathy. The risk of toxicity is reduced using therapeutic drug monitoring. CYP2D6 genotyping before therapy may allow earlier appropriate dosing. This study aimed to determine whether assessment of CYP2D6 genotype in patients on perhexiline could predict accurately metabolizer status as determined by the perhexiline metabolic ratio (MR). Blood samples from patients stabilized on perhexiline were analysed for CYP2D6 genotype and for concentrations of perhexiline and its hydroxy metabolite. The MR was determined. Of 74 patients, five were poor metabolizers (PM) defined by a MR<0.4, and the remainder were extensive metabolizers (EM). The genotypes were: *1/*1 (n=21), *1/*4 (n=18), *1/*2 (n=12), *1/*3 (n=2), *1/*5 (n=1), *1/*9 (n=2), *1/*10 (n=2), *2/*4 (n=4), *2/*2 (n=3), *4/*41 (n=3), *2/*41 (n=1), *41/*41 (n=1), *4/*9 (n=1), *4/*5 (n=1), *5/*6 (n=1) and *4/*6 (n=1). Allele frequencies were consistent with those reported in population studies. The 3 PMs with the lowest MR were predicted by genotype (*4/*5, *5/*6, *4/*6). The other 2 PMs had intermediate metabolizer genotypes and were on CYP2D6 inhibiting drugs. Amongst the EMs, the highest MR was associated with *1 and *2 allele combinations and the MR was progressively lower with the presence of alleles with intermediate function (*9, *10, *41) followed by alleles with no functional product (*3, *4, *5, *6). Thus, a gene-dose effect was observed. Genotype predicted PM phenotype and also intermediate metabolizers. Determination of CYP2D6 genotype before therapy with perhexiline may help predict perhexiline dose requirements and reduce the risk of perhexiline concentration-related toxicity.

Polymorphic hydroxylation of perhexiline in vitro.

AIMS: The aims of this study were to examine the in vitro enzyme kinetics and CYP isoform selectivity of perhexiline monohydroxylation using human liver microsomes. METHODS: Conversion of rac-perhexiline to monohydroxyperhexiline by human liver microsomes was assessed using a high-performance liquid chromatography assay with precolumn derivatization to measure the formation rate of the product. Isoform selective inhibitors were used to define the CYP isoform profile of perhexiline monohydroxylation. RESULTS: The rate of perhexiline monohydroxylation with microsomes from 20 livers varied 50-fold. The activity in 18 phenotypic perhexiline extensive metabolizer (PEM) livers varied about five-fold. The apparent Km was 3.3 +/- 1.5 micro m, the Vmax was 9.1 +/- 3.1 pmol min-1 mg-1 microsomal protein and the in vitro intrinsic clearance (Vmax/Km) was 2.9 +/- 0.5 micro l min-1 mg-1 microsomal protein in the extensive metabolizer livers. The corresponding values in the poor metabolizer livers were: apparent Km 124 +/- 141 micro m; Vmax 1.4 +/- 0.6 pmol min-1 mg-1 microsomal protein; and intrinsic clearance 0.026 micro l min-1 mg-1 microsomal protein. Quinidine almost completely inhibited perhexiline monohydroxylation activity, but inhibitors selective for other CYP isoforms had little effect. CONCLUSIONS: Perhexiline monohydroxylation is almost exclusively catalysed by CYP2D6 with activities being about 100-fold lower in CYP2D6 poor metabolizers than in extensive metabolizers. The in vitro data predict the in vivo saturable metabolism and pharmacogenetics of perhexiline.

Method for the analysis of perhexiline and its hydroxy metabolite in plasma using high-performance liquid chromatography with precolumn derivatization.

A high-performance liquid chromatographic method for the analysis of perhexiline and its monohydroxy metabolite in plasma has been developed. After a simple extraction procedure, the analytes are derivatized over a 30-min period with trans-4-nitrocinnamoyl chloride. The derivatized products are monitored at 340 nm following separation on a 5-micron phenyl reversed-phase column under isocratic conditions. The limits of detection for perhexiline and its hydroxy metabolite are 0.03 and 0.02 mg/l, respectively. The between-day and within-day assay coefficients of variation for perhexiline and its hydroxy metabolite at concentrations of 0.2 and 1.0 mg/I were less than 10%. The method has proved robust and suitable for the routine monitoring of perhexiline and hydroxyperhexiline.

[Genetic polymorphism of the metabolism of drugs used in cardiac diseases]. / Polymorphisme génétique du métabolisme des médicaments utilisés en pathologie cardiovasculaire.

The genetic determinants of the metabolism of certain drugs used in cardiology is one predictable cause of variability in their pharmacokinetics and effects. Genetic polymorphism of the metabolism of drugs is characterised by the existence of several metabolic phenotypes, usually 2, which allow distinction between fast and slow metabolisers. Clinical identification of these phenotypes is relatively simple. The enzymatic deficiency in slow metabolisers concerns a specific metabolic pathway responsible for the biotransformation of the drug but it respects other eventual metabolic pathways. For a given drug and a given metabolic pathway, slow metabolisers are unable to eliminate the parent product by hepatic metabolism. When the drug is given orally, the plasma concentrations of the parent product in slow metabolisers are 5 to 25 times higher than those observed in fast metabolisers. Depending on the given drug, doses usually well tolerated by fast metabolisers may cause excessive effects in slow metabolisers. The pharmacodynamic consequences of genetic polymorphism on the metabolism of drugs depend essentially on the therapeutic index of the drug concerned and on the activity of the metabolites formed by the genetically determined pathway of the parent product. The pharmacokinetic and pharmacodynamic consequences of the two main genetic polymorphisms concerning drugs used in cardiology are discussed: the polymorphism of N acetylation and that of cytochrome P-450 IID6.

Synthesis and pharmacological properties of "soft drug" derivatives related to perhexiline.

In the hope of reducing the toxicity of perhexiline, a series of 27 cyclohexylaralkylamines II based on the "soft drug" concept and incorporating an amide function were synthesized. In a preliminary screening, compounds were evaluated for their alpha-adrenolytic activities. Several derivatives, especially N-(cyclohexylphenylmethyl)-2-(cyclohexyl-methylamino)acetamide (3), N-(cyclohexylphenylmethyl)-2-(homoveratrylmethylamino)acetam ide (7), and N-[2-(cyclohexylamino)ethyl]-alpha-cyclohexylbenzeneacetamide (23) had the same activity range as perhexiline in vitro in rat aorta strips. The in vitro metabolism of these three molecules was then investigated and compared to that of perhexiline. The effect upon the alpha-adrenolytic activity of introducing various N-aralkylamine groups on II was examined. Structure/activity relationships are discussed.

The enterohepatic circulation of perhexiline metabolites in the male Wistar rat.

1. The biliary excretion of some perhexiline metabolites has been assessed in male Wistar rats with biliary cannulation. 2. After intragastric administration of perhexiline maleate (2 mg/kg body weight) multiple perhexiline metabolites were detected in bile. 3. When aliquots of this metabolite-laden bile were administered intraduoduodenally to further 'recipient' rats with biliary cannulation, similar metabolites were detected in the bile of these rats, but at reduced concentrations equivalent to 30-35% of those present in the bile of 'donor' rats. 4. These findings indicate that in the male Wistar rat, there may be substantial enterohepatic circulation of some perhexiline metabolites.

Studies on the metabolism of perhexiline in man.

We have studied the disposition of perhexiline and its two major metabolites, M1 and M3, in healthy volunteers and in patients with biliary T-tube drains after cholecystectomy. In healthy volunteers the genetic control for impaired hepatic oxidation is identical for debrisoquine, sparteine, and perhexiline. Poor metabolizers demonstrate markedly reduced production and excretion of the major metabolite, M1. Their production of M3 is also reduced, but to a lesser degree than for M1, confirming substrate stereoselectivity by hepatic oxidases. Biphasic urinary elimination of M1 and M3 is seen in intact extensive oxidizers, whereas only the first phase is apparent in patients with biliary T-tube drainage. This suggests the possibility of enterohepatic recycling of these compounds, which may account for their prolonged elimination. More than 90% of an ingested dose of perhexiline maleate remains unaccounted for at 24 h after ingestion, even in extensive metabolizers. A careful, radiolabelled tissue-distribution study is warranted to elucidate the complicated metabolic fate of perhexiline.

Perhexiline maleate treatment for severe angina pectoris--correlations with pharmacokinetics.

Perhexiline maleate, which causes inhibition of myocardial fatty acid catabolism with a concomitant increase in glucose utilization, is particularly useful in the management of patients with severe angina pectoris. While perhexiline exerts no significant negative inotropic or dromotropic effects, its short- and long-term use has hitherto been restricted because of complex pharmacokinetics and the eventual development, in many patients, of hepatitis and peripheral neuropathy. Correlations between perhexiline dose, plasma drug concentrations, efficacy and development of toxicity were examined prospectively in 3 groups of patients. The first group (n = 29) were patients in whom perhexiline was added to previously prescribed anti-anginal medication for short-term (pre-surgical or post-myocardial infarction) control of angina pectoris. Over a mean treatment period of 18 +/- 2 (SEM) days, 13 patients experienced a marked reduction in frequency and severity of attacks. No adverse effects occurred. A second group of patients (n = 19) were treated chronically with 50-400 mg/day of perhexiline, dosage being adjusted to minimize symptoms. Over a mean treatment period of 8.8 +/- 1.7 months, 5 patients became asymptomatic, while 9 developed evidence of hepatitis or neurotoxicity, with concomitant plasma perhexiline concentrations of 720-2680 ng/ml. Subsequently, a further group of similar patients (n = 22) were treated for 12.4 +/- 2.6 months, perhexiline dosage being adjusted to maintain plasma perhexiline concentrations below 600 ng/ml. Nine patients became asymptomatic, while none developed adverse effects. It is concluded that perhexiline is useful both as a short-term adjunct to anti-anginal therapy and in the long-term management of patients unsuitable for coronary artery bypass grafting. The risk of long-term toxicity can be reduced markedly by maintenance of plasma drug concentrations below 600 ng/ml without significantly compromising anti-anginal efficacy.

Stereoselective pharmacokinetics of perhexiline.

Blood plasma and urine excretion pharmacokinetics of the (+) and (-) enantiomers of perhexiline have been determined in oral single-dose studies in eight human volunteers, and compared with the pharmacokinetics of the racemate drug in the same subjects. The (-) enantiomer is more rapidly metabolized and eliminated, and is stereoselectively hydroxylated to the cis-monohydroxy-perhexiline. The peak plasma concn of unchanged perhexiline is greater, while that of the cis-monohydroxy-perhexiline metabolite is lower, after administration of the (+) enantiomer than after the (-) enantiomer or the racemate. Similarly, the AUC values for unchanged perhexiline and for the trans-monohydroxy-perhexiline metabolite are greatest and the AUC value for the cis-monohydroxy-perhexiline metabolite is lowest for the (+) enantiomer. The three stereoisomeric forms of perhexiline all had the same times to peak plasma concn of the unchanged drug or of the cis-metabolite, and all three forms had a similar plasma elimination half-life for unchanged perhexiline. Metabolism of racemic perhexiline to the cis-monohydroxy metabolite is the major mechanism of elimination of the drug in man and has been shown to be polymorphic in human populations. The (-) enantiomer which shows stereoselective metabolism to the cis metabolite might therefore show a greater polymorphic effect. Studies with rat-liver microsomal preparations in vitro showed that, in contrast to the human studies in vivo, hydroxylation of perhexiline yields mostly the trans-monohydroxy metabolite. The DA strain of rats exhibited slower rates of hydroxylation in vitro than Wistar or Lewis strains of rats.

Further studies on the pharmacokinetics of perhexiline maleate in humans.

We have performed single-dose pharmacokinetic studies on perhexiline in eight young volunteers, each given 300 mg of Pexid orally, using an h.p.l.c. method for the separation and quantification of the drug and its monohydroxy metabolites in plasma and urine. The plasma concentration of the cis-monohydroxyperhexiline (peak of 473 +/- 43 ng/ml at 7.5 +/- 2.0 h) was always higher than for unchanged perhexiline (peak of 112 +/- 20 ng/ml at 6.5 +/- 2.0 h) whereas the concentration of the transmetabolite was either low or undetectable in plasma. These findings indicate the occurrence of stereospecific pre-systemic metabolism of perhexiline which reduces the bioavailability of the parent drug. The plasma elimination half-life of perhexiline was 12.4 +/- 6.1 h (range 7-23 h) while that for cis-monohydroxyperhexiline was 19.9 +/- 7.7 h (range 10-29 h). Not more than 0.3% of unchanged perhexiline was excreted in the urine over five days in eight subjects. Between 3 and 23% of the orally administered drug was excreted as the cis- or trans-monohydroxy metabolites, the ratio of trans to cis metabolites being 0.52 +/- 0.20.

[Clinical pharmacology of calcium inhibitors]. / Pharmacocinétique clinique des inhibiteurs calciques.

The pharmacokinetics of the commercially available calcium antagonists, diltiazem (Tildiem), nifedipine (Adalate), perhexiline (Pexid), and verapamil (Isoptine) are well known; the pharmacokinetics of bepridil (Cordium) need further study. The properties of nicarpidine, a molecule currently being tested, will also be described. These products are well absorbed from the gastrointestinal tract but undergo variable degrees of transformation during the first passage through the liver. The bioavailabilities of bepridil, diltiazem and nifedipine are of the order of 40 to 60%; those of verapamil and nicarpidine are lower, 10-20% and 15-30%, respectively. The rates of absorption vary according to the derivatives and galenic preparations; in general, they are rapid; peak plasma concentrations are usually obtained one to four hours after administration. Protein binding is high but does not interfere in the distribution; the volumes of distribution of bepridil, diltiazem and verapamil are large (4-5 l/kg); those of nifedipine and nicardipine are smaller (l l/kg). The halflives of diltiazem, nifedipine, nicardipine and verapamil are short (1 to 5 hours); those of bepridil and perhexiline are longer (2 to 3 days). The main method of elimination is by hepatic transformation with high plasma clearance rates: diltiazem and verapamil have pharmacologically active derivatives whose contributions to the overall activities of the drugs are not fully understood. Physiopathological changes of the pharmacokinetic properties of diltiazem and verapamil (elderly patients, hepatic failure) have been described.