5-hydroxymethyl Metabolite Research Articles

Detection of antimuscarinic agents tolterodine and fesoterodine and their metabolite 5-hydroxymethyl tolterodine by ion transfer voltammetry at a polarized room-temperature ionic liquid membrane

Ion transfer voltammetry at a polarized hydrophobic room temperature ionic liquid (RTIL) membrane was used for evaluation of the diffusion coefficients and the standard Gibbs energies of ion transfer of the protonated antimuscarinic agents tolterodine (TOL), fesoterodine (FES), and their common metabolite 5-hydroxymethyl tolterodine (5-HMT), as well as for their determination in the aqueous samples and urine. An analysis of the pH effect provided the parameters characterizing their lipophilicity both in their ionic and neutral forms confirming a remarkably low lipophilicity of 5-HMT. The application of the ion transfer voltammetry for a monitoring of the enzymatic hydrolysis of FES to 5-HMT was demonstrated. For determination of protonated TOL, FES, and 5-HMT in the aqueous samples, linear calibration dependences were plotted in the range 2.0–12.5 μmol L−1 with the mean limit of detection (LOD) 0.43 μmol L−1. Analogous linear calibration dependences were constructed for 10times diluted urine spiked with 2.0–12.5 μmol L−1 TOL, FES, and 5-HMT with the mean LOD 0.65 μmol L−1. Unfortunately, determination of 5-HMT in diluted urine samples was interfered by an unknown ionic urine component, which somewhat increased its particular LOD.

Роль фезотеродина в лечении синдрома гиперактивного мочевого пузыря

The muscarinic receptors have been used as target in treatment of overactive bladder (OAB) for a long time. These patients have complaints of urgency, increased urinary frequency and nocturia, sometimes with urinary incontinence (involuntary urine leakage which is associated with abrupt and strong desire to void). Fesoterodine is a prodrug that is structurally and functionally associated with tolterodine and it is the novel drug for OAB treatment. As a result of fesoterodine cleavage by non-specific esterase, the active metabolite 5-hydroxymethyl tolterodine (5-HMT) is formed. Like other antimuscarinic drugs, fesoterodine allows to improve bladder storage function (reducing the frequency of urination) and urgency. For assessing an improvement of the quality of life after treatment by fesoterodine the KHQ questionnaire was used.

Physiological based pharmacokinetic modeling to estimate in vivo Ki of ketoconazole on renal P-gp using human drug-drug interaction study result of fesoterodine and ketoconazole

This study was conducted to estimate in vivo inhibition constant (Ki) of ketoconazole on renal P-glycoprotein (P-gp) using human drug-drug interaction (DDI) study result of fesoterodine and ketoconazole. Fesoterodine is a prodrug which is extensively hydrolyzed by non-specific esterases to the active metabolite 5-hydroxymethyl tolterodine (5-HMT). 5-HMT is then further metabolized via Cytochrome P450 (CYP) 2D6 and CYP3A4. It is reported that 5-HMT is a substrate of P-gp whereas fesoterodine is not. Renal clearance of 5-HMT is approximately two-times greater than renal glomerular filtration rate. This suggests the possibility that renal clearance of 5-HMT involves secretion by P-gp. Utilizing the available pharmacokinetic characteristics of fesoterodine and 5-HMT, we estimated in vivo Ki of ketoconazole on P-gp at kidney based on DDI study data using physiologically-based pharmacokinetic approach. The estimated in vivo Ki of ketoconazole for hepatic CYP3A4 (6.64 ng/mL) was consistent with the reported values. The in vivo Ki of ketoconazole for renal P-gp was successfully estimated as 2.27 ng/mL, which was notably lower than reported in vitro 50% inhibitory concentration (IC50) values ranged 223–2440 ng/mL due to different condition between in vitro and in vivo.

Mass Spectrometric and Functional Aspects of Drug–Protein Conjugation

The covalent binding of drugs (metabolites) to proteins to form drug-protein adducts can have an adverse effect on the body. These adducts are thought to be responsible for idiosyncratic drug reactions including severe drug hypersensitivity reactions. Major advances in proteomics technology have allowed for the identification and quantification of target proteins for certain drugs. Human serum albumin (HSA) and Hb have been identified as accessible targets and potential biomarkers for drug-protein adducts formation, for numerous drugs (metabolites) including β-lactam antibiotics, reactive drug metabolites such as quinone imines (acetaminophen) and acyl glucuronides (diclofenac), and covalent inhibitors (neratinib). For example, MS/MS analysis of plasma samples from patients taking flucloxacillin revealed that flucloxacillin and its 5-hydroxymethyl metabolite formed covalent adducts with lysine residues on albumin via opening of the β-lactam ring. Other proteins such as P450 and keratin are also potential targets for covalent binding. However, for most drugs, the properties of these target proteins including their location, their quantity, the timing of conjugate generation, and their biological function are not well understood. In this review, currently available proteomic technologies including MS/MS analysis to identify antigens, precise location of modifications, and the immunological consequence of hapten-protein complex are illustrated. Moving forward, identification of the nature of the antigenic determinants that trigger immune responses to drug-protein adducts will increase our ability to predict idiosyncratic toxicity for a given compound.

Pharmacokinetic drug interaction study between overactive bladder drugs mirabegron and tolterodine in Japanese healthy postmenopausal females

Mirabegron, the first selective β3-adrenoceptor agonist for the treatment of overactive bladder (OAB), inhibits cytochrome P450 isozyme CYP2D6. This study was performed in Japanese healthy postmenopausal female volunteers to assess any pharmacokinetic drug interaction between mirabegron and tolterodine, another OAB drug and a sensitive substrate of CYP2D6. Tolterodine 4 mg was orally administered from Days 1–7 and co-administered with mirabegron 50 mg from Days 8–14. Mirabegron 50 mg increased maximum concentration (Cmax) and area under the concentration-time curve from zero to 24 h after dosing (AUC24h) of tolterodine by 2.06-fold (90% confidence interval [CI] 1.81, 2.34) and 1.86-fold (90% CI 1.60, 2.16), respectively, and increased Cmax and AUC24h of the metabolite 5-hydroxymethyl tolterodine by 1.36-fold (90% CI 1.26, 1.47) and 1.25-fold (90% CI 1.15, 1.37), respectively. This suggested a weak pharmacokinetic drug interaction between mirabegron and tolterodine. Mean change from baseline of Fridericia's QT correction formula (ΔQTcF) was slightly higher on Day 14 than on Day 7. No subject had QTcF >480 msec or ΔQTcF >60 msec. All the treatment-emergent adverse events were mild. Mirabegron 50 mg was considered to be safe and well tolerated when coadministered with tolterodine 4 mg in healthy postmenopausal female volunteers.

Electrochemical oxidation of fesoterodine and identification of its oxidation products using liquid chromatography and mass spectrometry

The electrochemical behavior of fesoterodine (FES), an antimuscarinic drug used for the treatment of urge incontinence and overactive bladder, was investigated using linear sweep and cyclic voltammetry at a stationary and rotating disc glassy carbon electrodes. A single two-electron anodic signal of FES was observed in neutral buffered aqueous methanolic solutions. Kinetics of alkaline hydrolysis of FES to its active metabolite 5-hydroxymethyl tolterodine was investigated by time dependent linear sweep voltammetry. Controlled potential electrolysis of FES solutions was performed at platinum gauze electrode in aqueous-methanolic media. Electrolyzed solutions were analyzed using ultra performance liquid chromatography with electrospray ionization quadrupole time-of-flight mass spectrometry. Two main products of electrochemical oxidation of fesoterodine were identified as 5-formyl fesoterodine (isobutyric acid 2-(3-diisopropylamino-1-phenyl-propyl)-4-formyl-phenyl ester) and N-desisopropylated fesoterodine (isobutyric acid 4-hydroxymethyl-2-(3-isopropylamino-1-phenyl-propyl)-phenyl ester). The mechanism of the electrochemical oxidation of FES has been proposed and confirmed using on-line electrochemistry/mass spectrometry with porous glassy carbon electrode.

Selective Inhibitory Effect of Imidafenacin and 5-Hydroxymethyl Tolterodine on Capsaicin Sensitive C Fibers of the Primary Bladder Mechanosensitive Afferent Nerves in the Rat

Imidafenacin and fesoterodine are used to treat overactive bladder. Imidafenacin, fesoterodine and its active metabolite 5-hydroxymethyl tolterodine are muscarinic receptor antagonists. It is believed that these agents act on afferent nerves in addition to smooth muscle. We investigated the effects of imidafenacin and 5-hydroxymethyl tolterodine on single unit afferent activity of mechanosensitive capsaicin sensitive and insensitive primary bladder afferent nerve fibers in rats. Female Sprague Dawley® rats were anesthetized. Single unit afferent activity was recorded from the L6 dorsal roots and classified by conduction velocity as that of Aδ or C fibers. After measuring control single afferent activity during constant filling cystometry the procedure was repeated with intravenous administration of imidafenacin (0.3 to 30 μg/kg) or 5-hydroxymethyl tolterodine (0.01 to 1 mg/kg) at cumulative doses with or without intravesical capsaicin or oxotremorine-M instillation. A total of 139 single unit afferent fibers were isolated from 111 rats, including 19 Aδ and 120 C fibers. Neither imidafenacin nor 5-hydroxymethyl tolterodine significantly affected the overall single unit afferent activity of Aδ or C fibers. Based on capsaicin sensitivity C fibers were divided into capsaicin sensitive and insensitive groups. Each antimuscarinic inhibited the single unit afferent activity of capsaicin sensitive C fibers but not of capsaicin insensitive C fibers at the highest dose. Moreover, oxotremorine-M facilitated single unit afferent activity in a proportion of C fibers. The facilitated single unit afferent activity was significantly attenuated by the highest dose of imidafenacin. These findings demonstrate that imidafenacin and 5-hydroxymethyl tolterodine can selectively inhibit capsaicin sensitive C fibers among mechanosensitive bladder afferents by antagonizing bladder muscarinic receptors.

In vitro hepatic microsomal metabolism of meloxicam in koalas (Phascolarctos cinereus), brushtail possums (Trichosurus vulpecula), ringtail possums (Pseudocheirus peregrinus), rats (Rattus norvegicus) and dogs (Canis lupus familiaris)

Quantitative and qualitative aspects of in vitro metabolism of the non-steroidal anti-inflammatory drug meloxicam, mediated via hepatic microsomes of specialized foliage (Eucalyptus) eating marsupials (koalas and ringtail possums), a generalized foliage eating marsupial (brushtail possum), rats, and dogs, are described. Using a substrate depletion method, intrinsic hepatic clearance (in vitro Clint) was determined. Significantly, rates of oxidative transformation of meloxicam, likely mediated via cytochromes P450 (CYP), were higher in marsupials compared to rats or dogs. The rank order of apparent in vitro Clint was brushtail possums (n=3) (mean: 394μL/min/mg protein), >koalas (n=6) (50), >ringtail possums (n=2) (36) (with no significant difference between koalas and ringtail possums), >pooled rats (3.2)>pooled dogs (in which the rate of depletion, as calculated by the ratio of the substrate remaining was <20% and too slow to determine). During the depletion of meloxicam, at a first-order rate constant, 5-hydroxymethyl metabolite (M1) was identified in the brushtail possums and the rat as the major metabolite. However, multiple hydroxyl metabolites were observed in the koala (M1, M2, and M3) and the ringtail possum (M1 and M3) indicating that these specialized foliage-eating marsupials have diverse oxidation capacity to metabolize meloxicam. Using a well-stirred model, the apparent in vitro Clint of meloxicam for koalas and the rat was further scaled to compare with published in vivo Cl. The closest in vivo Cl prediction from in vitro data of koalas was demonstrated with scaled hepatic Cl(total) (average fold error=1.9) excluding unbound fractions in the blood and microsome values; whereas for rats, the in-vitro scaled hepatic Cl fu(blood, mic), corrected with unbound fractions in the blood and microsome values, provided the best prediction (fold error=1.86). This study indicates that eutherians such as rats or dogs serve as inadequate models for dosage extrapolation of this drug to marsupials due to differences in hepatic turnover rate. Furthermore, as in vivo Cl is one of the pharmacokinetic indexes for determining therapeutic drug dosages, this study demonstrates the utility of in vitro to in vivo scaling as an alternative prediction method of drug Cl in koalas.

Multiple-dose pharmacokinetics of fesoterodine sustained-release in healthy Korean volunteers

Fesoterodine is a pro-drug of the active metabolite 5-hydroxymethyl tolterodine (5-HMT), a muscarinic receptor antagonist. This study aimed to evaluate the safety profile and pharmacokinetic characteristics of multiple oral doses of sustained-release fesoterodine (fesoterodine SR) in healthy Korean males. A randomized, double-blind, placebo-controlled, multiple-dose study with two oral doses (4 mg and 8 mg) was conducted in healthy Korean male participants. The study drug was administered once daily for 5 days. The plasma concentration of 5-HMT was measured up to 72 hours after the last drug administration. The CYP2D6 genotype was analyzed using polymerase chain reaction (PCR) methods to assess the effect of genetic polymorphisms on the pharmacokinetic parameters. 20 participants completed the study. The mean (SD) areas under the plasma concentration-time curves during the dosing interval (AUCτ) of the 4 mg and 8 mg dose groups were 26.1 (8.0) and 64.2 (30.5) μg·h/ml and the mean peak concentrations (Cmax) were 2.6 (0.7) and 6.0 (2.0) μg/ml, respectively, at steady-state. The mean AUCτ and Cmax of 5-HMT increased in approximately the same proportion as the dose increased. Fesoterodine SR was well tolerated without any serious adverse events or abnormal clinical laboratory findings. Systemic 5-HMT exposure showed dose-proportional characteristics in the 4 mg to 8 mg dose range in healthy Korean males. Thus, 4 mg or 8 mg doses of fesoterodine SR taken once-daily were tolerable in healthy Korean males.

Clinical pharmacology of functional disorders of the urogenital system

Clinical pharmacology of functional disorders of the urogenital system

Pharmacokinetics of Tolterodine in Japanese and Koreans: Physiological and Stochastic Assessment of Ethnic Differences

Tolterodine is known as a drug which exhibits ethnic differences in pharmacokinetics between Japanese and Koreans despite genetic similarities among the populations of East Asian countries. Tolterodine is mainly metabolized by CYP2D6 to a 5-hydroxymethyl metabolite (5-HM), and 5-HM is also metabolized by CYP2D6. The reduced-function allele CYP2D6*10 is frequently observed in Asian populations. We investigated differences in the pharmacokinetics of tolterodine between small Japanese and Korean study populations by physiological and stochastic approaches with consideration of the CYP2D6 genotype. The genotype frequencies of CYP2D6*10/*10 and CYP2D6*5/*10 were found to be higher in Koreans than in Japanese, which suggested that this frequency difference occurred incidentally. The effects of CYP2D6 genotype and ethnicity on the intrinsic clearance of tolterodine by CYP2D6 were tested and only genotype was found to be a significant factor by ANCOVA. A simulation was conducted to confirm whether the observed differences in tolterodine exposure could be explained by the differences in genotype frequency found in this study. It was confirmed that the variability of intrinsic clearance could be responsible for the incidental exposure differences. In conclusion, apparent differences in exposure were found between small Japanese and Korean study populations because of the variability of intrinsic clearances and genotype frequencies.

LC–MS–MS Separation and Simultaneous Determination of Tolterodine and its Active Metabolite, 5-Hydroxymethyl Tolterodine in Human Plasma

A selective, sensitive and high throughput LC–MS–MS method has been developed and validated for the chromatographic separation and quantitation of tolterodine (TOL) and its metabolite 5-hydroxymethyl TOL in human plasma. Sample clean-up concerned liquid–liquid extraction of the drug, metabolite and their respective labelled internal standards from 300 μL human plasma. Both the analytes were chromatographically separated on a Symmetry C18 (100 mm × 4.6 mm, 5 μm particle size) analytical column using 10 mM ammonium formate (pH 5.0 ± 0.1, adjusted with formic acid) and acetonitrile (35:65, v/v) as the mobile phase with a resolution factor of 2.72. The method was validated over the concentration range of 0.025–10.0 ng mL−1 for both analytes. The process efficiency found for TOL and its metabolite was 98.3 and 99.5%, respectively. The method was successfully applied to a pivotal bioequivalence study in 41 healthy human subjects after oral administration of a 2 mg tablet formulation under fasting conditions.

Spinal Effects of the Fesoterodine Metabolite 5-Hydroxymethyl Tolterodine and/or Doxazosin in Rats With or Without Partial Urethral Obstruction

The combination of muscarinic receptor and alpha(1)-adrenoceptor antagonists is increasingly used for benign prostatic hyperplasia related lower urinary tract symptoms. In addition to the well established peripheral site of action, little is known about the central effects of muscarinic receptor antagonists and muscarinic receptor/alpha(1)-adrenoceptor antagonist combinations on bladder function, partly due to poor brain penetration after systemic administration. We assessed the effects of intrathecal 5-hydroxymethyl tolterodine, an active metabolite of fesoterodine, in obstructed and nonobstructed rats, and of combined intrathecal 5-hydroxymethyl tolterodine/doxazosin in a rat model of partial urethral obstruction. We used 80 male Sprague-Dawley rats to test various doses of intrathecal 5-hydroxymethyl tolterodine and/or intrathecal doxazosin on urodynamic parameters. Urodynamic evaluation without anesthesia was done 3 days after bladder and intrathecal catheterization. Two weeks before urodynamics 40 rats underwent partial urethral obstruction. Intrathecal 5-hydroxymethyl tolterodine had no urodynamic effects in nonobstructed rats. Compared to controls obstructed rats had increased bladder pressure and weight, and voiding frequency. In obstructed rats 5-hydroxymethyl tolterodine restored urodynamic parameters to those seen in nonobstructed animals. Doxazosin had effects similar to those of intrathecal 5-hydroxymethyl tolterodine. When the 2 drugs were combined, at the doses used only small additional effects were observed. Urodynamic changes in obstructed rats can be normalized by intrathecal 5-hydroxymethyl tolterodine and by intrathecal doxazosin. The central pathways on which the 2 drugs act seem to be up-regulated with partial urethral obstruction and less relevant under normal conditions.

Characterisation of flucloxacillin and 5‐hydroxymethyl flucloxacillin haptenated HSA in vitro and in vivo

Flucloxacillin is a synthetic penicillin used in the treatment of Staphylococcal infections. Adverse reactions to the drug are believed to arise through covalent modification of proteins, with tissue damage occurring secondary to an immune reaction. Serum proteins have been shown by adduct-specific antibodies to be modified by flucloxacillin, but the nature and sites of modification have not been characterised. Here, in vitro studies on HSA have shown by MS that the modification of protein lysine residues occurs in a dose-, time- and site-dependent manner. Affinity, cation exchange and reversed phase chromatography prior to MS revealed in vivo modification of HSA with flucloxacillin in tolerant patients, with up to nine modified lysine residues being detected in each patient, and with modification of Lys190 and Lys212 being detected in 8/8 patients. It was also revealed for the first time that plasma proteins could be modified with the 5-hydroxymethyl metabolite of flucloxacillin, and that essentially the same Lys residues were targeted by both the parent drug and its metabolite. This study provides a detailed characterisation of sites of chemical modification of an endogenous target and reveals candidate peptides for T-cell and antibody assays of flucloxacillin hypersensitivity.

Determination of tolterodine and its 5-hydroxymethyl metabolite in human plasma by hydrophilic interaction liquid chromatography–tandem mass spectrometry

A rapid and reliable method was developed to quantitate tolterodine and its 5-hydroxymethyl metabolite in human plasma using liquid chromatography–electrospray tandem mass spectrometry. The assay was based on liquid–liquid extraction of the compounds from plasma with tert-butylmethylether and hydrophilic interaction chromatography performed on a silica column (30 mm × 4.6 mm, 3 μm particles), the mobile phase consisted of acetonitrile-20 mM ammonium acetate (70:30, v/v). Quantification was through positive-ion mode and selected reaction monitoring at m/ z 326 → 147 for tolterodine, 342 → 223 for the 5-hydroxymethyl metabolite and 260 → 183 for the internal standard propranolol, respectively. The lower limit of quantitation was 49 and 46 pg/ml using 0.5 ml of plasma for the parent drug and its metabolite, respectively and linearity was observed up to 30 ng/ml. Within-day and between-day precision expressed by relative standard deviation was less than 11% and inaccuracy did not exceed 7% at all levels. The assay was applied to the analysis of samples from a pharmacokinetic study.

ANALYSIS OF POLYCYCLIC AROMATIC HYDROCARBONS IN AMNIOTIC FLUID SAMPLES FROM SMOKERS AND NONSMOKERS

Previous studies from this laboratory have focused on the characterization of blood protein adducts formed in utero as a result of maternal smoking during pregnancy. These biological samples, obtained during the third trimester of pregnancy, at delivery, have clearly shown a correlation between maternal smoking histories and exposure of the fetus to tobacco smoke carcinogens, including 4-aminobiphenyl and benzo(a)pyrene. In the present study, we examined exposure of the fetus during the first trimester of pregnancy to various environmental carcinogens, particularly those found in tobacco smoke. Amniotic fluid samples were obtained from women undergoing routine amniocentesis between 16 and 20 weeks gestational age. Amniotic fluid, produced by the fetal lungs and kidneys, is an important part of pregnancy and fetal development and this fluid surrounds the fetus throughout pregnancy. In these studies, samples of amniotic fluid were obtained from nonsmokers as well as 0.5 pack per day smokers, 1.0 pack per day smokers, and greater than 2.0 pack per day smokers. Maternal smoking status was determined by questionnaire as well as assessment of amniotic fluid for cotinine via immunoassay. Amniotic fluid samples were extracted and analyzed for the presence of polycyclic aromatic hydrocarbons (PAHs). A clear correlation was found between levels of maternal smoking and PAHs in the amniotic fluid. Amniotic fluid 1-hydroxypyrene levels ranged from 1.54 ± 0.12 μ g/L in nonsmokers to 11.72 ± 0.67 μ g/L in women smoking greater than 2 pks/da, indicating approximately a 10X increase over nonsmokers. Similar results were found 3-hydroxybenzo(a)pyrene, 6-hydroxybenzo(a)pyrene, and 3,6-dihydroxybenzo(a)pyrene, metabolites of the carcinogen benzo(a)pyrene as well as with the 9-hydroxy and 9,10-dihydroxy metabolites of anthracene. The 5-hydroxymethyl metabolite of 5-methylchrysene was found to range in concentrations from 1.65 ± 0.11 μ g/L in nonsmokers to 12.67 ± 0.79 μ g/L in greater than 2 packs per day smokers. These results demonstrate that amniotic fluid can serve as a biological marker of exposure to tobacco related polycyclic aromatic hydrocarbons. Identification of potentially harmful compounds detected at an early stage of pregnancy may prevent subsequent exposures to the fetus and as a result decrease the risk of potential genotoxic as well as teratogenic events.

Tolterodine does not affect memory assessed by passive-avoidance response test in mice

Antimuscarinics are first-line pharmacotherapy for the treatment of overactive bladder. However, because central nervous system cholinergic neurotransmission is involved in cognition, and the central nervous system-permeable antimuscarinics scopolamine and oxybutynin affect memory, cognitive impairment has been noted as a possible side effect of these drugs. We evaluated the effect of tolterodine, an antimuscarinic for overactive bladder, in a mouse passive-avoidance model of memory. Mice were chosen because like humans, mice but not rats, form the pharmacologically active 5-hydroxymethyl metabolite of tolterodine, DD01. In the passive-avoidance test, tolterodine at 1 or 3 mg/kg had no effect on memory; the latency to cross and percentage of animals crossing were comparable to controls. In contrast, scopolamine induced a memory deficit; the latency to cross was decreased, and the number of animals crossing was increased. Therefore, at a dose exceeding therapeutic exposure by six-fold, tolterodine had no effect on memory in the mouse passive-avoidance model, indicating that tolterodine does not disrupt cognitive function in this testing paradigm.

Comparative Evaluation of Central Muscarinic Receptor Binding Activity by Oxybutynin, Tolterodine and Darifenacin Used to Treat Overactive Bladder

We characterized muscarinic receptor binding in the mouse cerebral cortex after oral administration of anticholinergic agents used to treat overactive bladder. Muscarinic receptors in the mouse cerebral cortex and bladder after oral administration of anticholinergic agents were measured using [(3)H]N-methylscopolamine. In vitro binding affinities of tolterodine and its metabolite 5-hydroxymethyl metabolite in the mouse cerebral cortex and bladder were considerably greater than those of oxybutynin and darifenacin. Also, muscarinic receptor binding affinity of oxybutynin and its metabolite N-desethyl-oxybutynin in the cerebral cortex compared with that in the bladder was 2 to 3 times higher, whereas that of tolterodine and 5-hydroxymethyl metabolite was approximately 2 times lower. Oral administration of oxybutynin (76.1 micromol/kg), tolterodine (6.31 micromol/kg) and darifenacin (59.1 micromol/kg) showed binding activity that was approximately equal to that of bladder muscarinic receptors. Oral administration of oxybutynin (76.1 micromol/kg) showed significant binding of cerebral cortical muscarinic receptors in mice, as indicated by about a 2-fold increase in K(d) values for specific [(3)H]N-methylscopolamine binding 0.5 and 2 hours later. On the other hand, tolterodine and darifenacin given at oral doses that would exert a similar extent of bladder receptor binding activity as oxybutynin showed only a low level of binding activity of central muscarinic receptors in mice. Significant binding of brain muscarinic receptors in mice was observed by the oral administration of oxybutynin but not tolterodine and darifenacin.

Characterization of muscarinic receptor binding and inhibition of salivation after oral administration of tolterodine in mice

The current study was undertaken to characterize the effects of oral administration of tolterodine on muscarinic receptor binding in the bladder and submaxillary gland and on salivation in mice. In the in vitro experiment, tolterodine and its metabolite (5-hydroxymethyl metabolite: 5-HM) competed concentration-dependently with [ N-methyl- 3H]-scopolamine ([ 3H]NMS) in the mouse bladder, submaxillary gland and heart, and the potencies of both agents were greater than that of oxybutynin. After oral administration of tolterodine (6.31, 21.0 μmol/kg) and oxybutynin (76.1 μmol/kg), there was a dose and time-dependent increase in K d values for specific [ 3H]NMS binding in the bladder, prostate, submaxillary gland, heart, colon and lung, compared with control values, suggesting significant muscarinic receptor binding in each tissue. The K d increase in each tissue by oral oxybutynin reached a maximum value of 0.5 h after oral administration and then rapidly declined, while that by tolterodine was greatest 2 h after the administration and it was maintained for at least 6 or 12 h, depending on the dose and on the tissue. Thus, muscarinic receptor binding of oral tolterodine was slower in onset and of a longer duration than that of oxybutynin. Also, oral oxybutynin showed relatively greater receptor binding in the submaxillary gland as compared with other tissues, but such high selectivity to the exocrine gland muscarinic receptors was not observed by oral tolterodine. Oral administration of tolterodine and oxybutynin reduced significantly the pilocarpine-induced salivary secretion in mice, and the attenuation of oral tolterodine appeared more slowly and it was more persistent than that of oral oxybutynin. The antagonistic effect of oral tolterodine on the dose–response curves to pilocarpine was significantly weaker than that of oxybutynin. These data suggest that oral tolterodine, compared with the case of oral oxybutynin, binds more selectively to muscarinic receptors in the mouse bladder than in the submaxillary gland, which may be advantageous in treating patients with overactive bladder.

Effect of the Proton Pump Inhibitor Omeprazole on the Pharmacokinetics of Extended‐Release Formulations of Oxybutynin and Tolterodine

This study assessed the effect of the proton pump inhibitor omeprazole on the bioavailability of the extended-release formulations of oxybutynin and tolterodine. Forty-four healthy volunteers received each of 4 treatments in a 4-period crossover design. The treatments consisted of osmotically controlled extended-release oxybutynin chloride tablets at 10 mg/d or extended-release tolterodine tartrate capsules at 4 mg/d, with and without preceding treatment with 20 mg omeprazole daily for 4 days. Blood samples collected predose and at scheduled time points for 36 hours postdose were analyzed for oxybutynin and its active metabolite, N-desethyloxybutynin, or tolterodine and its active 5-hydroxymethyl metabolite, as appropriate. The AUCinfinity ratios for oxybutynin and its metabolite with and without prior omeprazole fell within the 80% to 125% range (accepted as the criterion for bioequivalence), as did those for tolterodine and its active moiety. The peak concentration ratios for oxybutynin and metabolite also conformed to this range; those for tolterodine did not. Increasing gastric pH with omeprazole does not substantially alter the pharmacokinetic properties of extended-release oxybutynin but may alter those of extended-release tolterodine.