Kosuke Doki

Co-administration of proton pump inhibitors delays elimination of plasma methotrexate in high-dose methotrexate therapy

AIM To assess whether or not co-administration of proton pump inhibitors (PPIs) is a risk factor for delayed elimination of plasma methotrexate (MTX) in high-dose MTX (HDMTX) therapy for malignant diseases. METHODS To assess the effects of PPI co-administration on elimination of plasma MTX, we examined plasma MTX concentration data on 171 cycles of HDMTX therapy performed in 74 patients. We performed multiple logistic regression analysis to evaluate PPI co-administration as a risk factor. Inhibitory potencies of omeprazole, lansoprazole, rabeprazole and pantoprazole on MTX transport via breast cancer resistance protein (BCRP, ABCG2) were also investigated in an in vitro study using membrane vesicles expressing human BCRP. RESULTS We identified co-administration of PPIs as a risk factor for delayed elimination (odds ratio 2.65, 95% confidence interval 1.03, 6.82) as well as renal and liver dysfunction. All four PPIs inhibited BCRP-mediated transport of MTX, with half-maximal inhibitory concentrations of 5.5-17.6 microM--considerably higher than the unbound plasma concentrations of the PPIs. CONCLUSIONS Our results support previous findings suggesting that PPI co-administration is associated with delayed elimination of plasma MTX in patients with HDMTX therapy. This drug interaction, however, cannot be explained solely by the inhibitory effects of PPIs on BCRP-mediated MTX transport.

Effects of CYP2D6 genotypes on age‐related change of flecainide metabolism: involvement of CYP1A2‐mediated metabolism

AIMS The aim of this study was to clarify the effects of CYP2D6 genotype on age-related change in flecainide metabolism in patients with supraventricular tachyarrhythmias. An in vitro study using microsomes was performed to identify other CYPs responsible for age-related change in flecainide metabolism. METHODS The study population comprised 111 genotyped patients: CYP2D6-homozygous extensive metabolizers (hom-EMs, n= 34), heterozygous EMs (het-EMs, n= 56), and intermediate and poor metabolizers (IMs/PMs, n= 21). Serum concentrations of flecainide and its metabolites [m-O-dealkylated flecainide (MODF) and m-O-dealkylated lactam of flecainide] were determined by use of a high-performance liquid chromatography. Metabolic ratio (MR) was expressed as serum concentrations of flecainide to its metabolites. In vitro formation of MODF was examined in human liver microsomes and cDNA-expressed CYP isoforms. RESULTS MR was higher in elderly patients (> or =70 years) than in middle-aged patients (<70 years). The increase of MR in elderly patients differed among CYP2D6 genotypes: 1.6-fold in het-EMs [4.3, 95% confidence interval (CI) 2.8, 5.7 vs. 2.7, 95% CI 2.3, 3.1, P < 0.05], 1.5-fold in IMs/PMs (6.0, 95% CI 4.5, 7.6 vs. 4.1, 95% CI 2.9, 5.4, P < 0.05), and no change in hom-EMs. The in vitro study using microsomes revealed that both CYP2D6 and CYP1A2 were involved in the formation of MODF. MODF formation in CYP2D6 PM microsomes increased as CYP1A2 activity increased. CONCLUSIONS The results suggest that patients with poor CYP2D6-mediated metabolism (het-EMs and IMs/PMs) showed age-related reduction in flecainide metabolism because metabolism was taken over by CYP1A2, whose activity decreases with age.

Drug interaction of tizanidine and fluvoxamine

hands this variability is in the region of 30%, similar to that of other cytotoxic agents. Moreover, other groups have shown a relationship between 5-FU plasma pharmacokinetics (but not dose) and response to treatment or toxicity, even though concentration was not measured at the site of action, suggesting that plasma pharmacokinetics is indicative of therapeutic activity. We report that we did not find a difference in plasma 5-FU pharmacokinetics in the presence of oxaliplatin, suggesting that the interaction does indeed take place at the cellular level. This would support further studies, with the use of methods such as those suggested by Drs Wolf and Presant, to investigate the anabolism or catabolism of 5-FU within the tumor. Such intratumoral methods have their own inherent problems, however, such as variation in the ratio of tumor to stroma or viable tissue to necrotic tissue, and are also less relevant to drug toxicity. In addition, although important in probing this interaction further at the clinical level, such approaches ask different questions than those posed in our own study. For example, it was important to establish that the increased rate of diarrhea seen with oxaliplatin–5-FU compared with the rate for 5-FU alone is not simply a result of reduced 5-FU clearance. In understanding the mechanism underlying clinically important drug interactions, we believe there is clearly a role for both types of study.

Assessment of serum flecainide trough levels in patients with tachyarrhythmia.

The reported therapeutic range for trough flecainide concentration is 200–1000 ng mL−1. Severe adverse events, such as ventricular arrhythmias, have occurred occasionally in patients whose serum flecainide exceeded 1000 ng mL−1. However, the lower limit remains controversial. We have evaluated blood flecainide concentrations in patients with tachyarrhythmia who received the drug to control palpitation. We measured the flecainide trough levels and incidence and frequency of palpitation of 44 outpatients receiving oral flecainide (150–300 mg daily). Mean serum flecainide trough concentrations differed significantly between patients with (n = 14) and without (n = 30) palpitation (259.5 ± 85.2 vs 462.2 ± 197.7 ng mL−1, P < 0.01). The frequency of palpitation decreased as the serum flecainide concentration increased. The incidence of palpitation was 65% at serum flecainide concentrations < 300 ng mL−1 and 11% at ≥ 300 ng mL−1. QRS values were increased significantly in patients with serum flecainide ≥ 300 ng mL−1 compared with < 300 ng mL−1 (0.110 ± 0.016s vs 0.093 ± 0.019s, P < 0.05). We concluded that to control paroxysm in patients receiving flecainide for tachyarrhythmia serum flecainide concentrations should be maintained at ≥ 300 ng mL−1.

Impact of CYP3A5 genetic polymorphism on pharmacokinetics of tacrolimus in healthy Japanese subjects

Tacrolimus (TAC), a calcineurin inhibitor, is an important immunosuppressive agent for treating autoimmune disease, including myasthenia gravis and rheumatoid arthritis, as well as for preventing allograft rejection in organ transplantation [1]. Since the therapeutic range of blood TAC is narrow, current interest focused on this drug is to maintain optimal blood concentrations in therapeutic drug monitoring (TDM) based on pharmacogenomic data, such as CYP3A5 and MDR1 gene polymorphisms [2]. Fukudo et al.[2] have reported that both CYP3A5 genotype and MDR1 mRNA expression were important factors affecting TAC pharmacokinetics in paediatric living related liver transplantation. Choi et al.[3] have revealed the impact of CYP3A5 genotype on TAC pharmacokinetics in healthy Koreans, where the area under the concentration–time curve (AUC) of TAC in subjects with CYP3A5*3/*3 was 2.5-fold higher compared with CYP3A5*1 carriers. We have examined the impact of CYP3A5 and MDR1 genetic polymorphism on TAC pharmacokinetics in healthy Japanese by using population pharmacokinetics (PPK) analysis. This study was approved by the Ethical Committee of University of Tsukuba (Tsukuba, Japan) and written informed consent was obtained in each case. Twenty healthy subjects received a single dose of oral TAC (2 mg; Prograf; capsule Astellas Pharma Inc., Tokyo, Japan). Venous blood samples for determining blood TAC were collected before and 1, 2, 4 and 8 h after the administration. Blood TAC was determined by microparticle enzyme immunoassay. Since the blood TAC concentration at 8 h after dose was as low as the detection limit for this method (1.5 ng ml−1), we did not measure the concentration after 8 h. PPK analysis was performed using WinNonMix (Version 2.0; Pharsight, Mountain View, CA, USA) with a one-compartment model to calculate apparent clearance (CL/F) and volume of distribution (V/F). The absorption rate constant (ka) was fixed to a reported value (4.5 h−1) [4]. Age, body weight, sex, and CYP3A5 and MDR1 genotypes were evaluated as the covariates by conducting a forward and backward stepwise regression analysis. Genome DNA was isolated from peripheral blood by using the QIAamp DNA Mini Kit (Qiagen GmbH, Hilden, Germany). The genotyping of CYP3A5 and MDR1 at the position of 1236C→T, 2677G→A/T and 3435C→T, was conducted by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method [5, 6]. MDR1−1517a T→C polymorphism, which was reported by Takane et al.[7] as the predictive factor of the expression level of MDR1 mRNA (NCBI dbSNP ID: rs28381796), was also genotyped by using a PCR-RFLP method developed in our laboratory. TAC pharmacokinetic parameters are summarized in Table 1. The AUC0−8h in subjects with CYP3A5*3/*3 were 1.8-fold higher than that in CYP3A5*1 carriers (P = 0.03), which agreed with the report of Choi et al.[3]. MDR1−1517a T/C genotype (n = 4) tended to have lower AUC0−8h compared with −1517a T/T genotype (n = 10) (20.8 ± 11.1 vs. 29.2 ± 8.2 ng h ml−1) in CYP3A5*1 carriers, although the difference was not statistically significant because of insufficient power. Table 1 Effect of CYP3A5 polymorphism on tacrolimus pharmacokinetics in healthy Japanese We conducted PPK analysis to estimate the contribution of CYP3A5 genotype on individual variations of TAC pharmacokinetics. Among the covariates, body weight affected the V/F and CL/F, and CYP3A5*1 allele only affected CL/F (Table 2). The estimated CL/F in subjects with CYP3A5*1 carriers was 1.5 times higher than that in subjects with CYP3A5*3/*3 (Table 2), which was comparable to measured values. The interindividual variability in CL/F with final model 4 was reduced from that with model 3 including no genetic covariates (22.4% to 9.7%, base model 1; 40.3%). These results suggest that approximately 32% of the interindividual variation of TAC CL/F can be explained by CYP3A5 polymorphism in healthy Japanese. This value is greater than the 9% and 23% reported, which were obtained in paediatric [2] and adult [8] liver transplantation situations, respectively. The liver transplantation is very complex in estimating the efficacy of CYP3A5 polymorphism on TAC pharmacokinetics, because of several issues such as the mismatch in genotype between recipients and donors and change in enzyme activities due to liver function, which may explain the discrepancy between the present and reported values. Because these factors associated with liver transplantation do not affect TAC pharmacokinetics in patients with myasthenia gravis and rheumatoid arthritis as well as healthy subjects, it is considered that the impact of CYP3A5 genotype on TAC TDM for autoimmune diseases is important. We therefore suggest that it would be useful to include CYP3A5 genotyping in TAC therapy for autoimmune diseases, especially in the Asian population, which frequently carry the CYP3A5*1 allele. Table 2 Summary of analysis models estimating for pharmacokinetic parameters of tacrolimus

CYP2D6 genotype affects age-related decline in flecainide clearance: a population pharmacokinetic analysis.

Objective To investigate the association between age-related decline in flecainide clearance and CYP2D6 genotype, we conducted a population pharmacokinetic analysis of flecainide using routine therapeutic drug monitoring data. Methods Population pharmacokinetic analysis was performed on retrospective data from 163 genotyped patients treated with oral flecainide for supraventricular tachyarrhythmias. The CYP2D6 genotype was categorized as CYP2D6 homozygous extensive metabolizers (hom-EMs; n=57), heterozygous extensive metabolizers (het-EMs; n=79), and intermediate metabolizers and poor metabolizers (IMs/PMs; n=27). Results Population pharmacokinetic analysis revealed that estimated glomerular filtration rate, body weight, female sex, and aging were important factors for estimating flecainide clearance. The metabolic clearance was decreased age dependently in a curvilinear fashion, where the lower clearance was observed in greater than 60 years for het-EMs and greater than 55 years for IMs/PMs. The reduction in metabolic clearance in elderly (70 years) patients compared with middle-aged (52 years) patients was different among the CYP2D6 genotype groups: 22.1 and 49.5% in CYP2D6 het-EMs and IMs/PMs, respectively, and no change in hom-EMs. A 11.4% reduction in estimated glomerular filtration rate in elderly patients compared with middle-aged patients corresponded to 6.1% decline in flecainide clearance. Overall, the age-related decline in flecainide clearance was 6.1% in hom-EMs, 16.3% in het-EMs, and 28.9% in IMs/PMs groups. Conclusion This study suggests that CYP2D6 genotype is a determinant factor of age-related decline in flecainide clearance.

Difference in blood tacrolimus concentration between ACMIA and MEIA in samples with low haematocrit values.

Objectives  The aim was to compare blood tacrolimus concentrations in anaemic patients between affinity column‐mediated immunoassay (ACMIA) and microparticle enzyme immunoassay (MEIA).

Virtual bioequivalence for achlorhydric subjects: The use of PBPK modelling to assess the formulation-dependent effect of achlorhydria

&NA; Majority of bioequivalence studies are conducted in healthy volunteers. It has been argued that bioequivalence may not necessarily hold true in relevant patient populations due to a variety of reasons which affect one formulation more than the other for instance in achlorhydric patients where elevated gastric pH may lead to differential effects on formulations which are pH‐sensitive with respect to release or dissolution. We therefore examined achlorhydria‐related disparity in bioequivalence of levothyroxine and nifedipine formulations using virtual bioequivalence within a physiologically‐based pharmacokinetic (PBPK) modelling framework. The in vitro dissolution profiles at neutral pH were incorporated into PBPK models to mimic the achlorhydria with in vitro–in vivo relationship established using bio‐relevant pH media. The PBPK models successfully reproduced the outcome of the bioequivalence studies in healthy volunteers under the normal conditions as well as under proton pump inhibitor‐induced achlorhydria. The geometric mean test/reference ratios for Cmax and AUC between levothyroxine tablet and capsule in patients receiving proton pump inhibitor were 1.21 (90%CI, 1.13–1.29) and 1.09 (90%CI, 1.02–1.17), respectively. Extension of the virtual bioequivalence study to Japanese elderly, who show high incidence of achlorhydria, indicated bio‐inequivalence which Cmax and AUC ratios between nifedipine control‐released reference and test formulations were 3.08 (90%CI, 2.81–3.38) and 1.57 (90%CI, 1.43–1.74), respectively. Virtual bioequivalence studies through the PBPK models can highlight the need for conduct of specific studies in elderly Japanese populations where there are discrepancies in pH‐sensitivity of dissolution between the test and reference formulations. Graphical abstract Figure. No caption available.

SCN5A promoter haplotype affects the therapeutic range for serum flecainide concentration in Asian patients

Objective An increased slowing of cardiac conduction induced by sodium channel blockers is remarkably observed in carriers of an Asian-specific promoter haplotype [haplotype B (HapB)] of the cardiac sodium channel gene (SCN5A). We investigated the effect of HapB on the therapeutic range for serum flecainide concentration in Asian patients. Patients and methods We examined the serum concentration and antiarrhythmic efficacy of flecainide, together with the SCN5A promoter haplotype, in 146 patients with supraventricular tachyarrhythmias. Trough serum flecainide concentrations were determined by HPLC. The antiarrhythmic efficacy of flecainide was assessed for at least 2 months through examination of symptomatology, ECG, and Holter monitoring. Results The serum flecainide concentration did not differ between the wild-type HapA homozygotes and HapB carriers under treatment with the usual dose. A genetic difference in the antiarrhythmic efficacy of flecainide was observed between the HapA homozygotes and HapB carriers at serum flecainide concentrations less than 300 ng/ml (42.9 vs. 68.8%; P=0.022). PR prolongation and QRS widening were observed more commonly among the HapB carriers with serum flecainide concentrations of at least 300 ng/ml than in the HapA homozygotes (PR, 210±25 vs. 195±25 ms; P=0.036; and QRS, 112±10 vs. 105±9 ms; P=0.030). Conclusion These findings suggest that the therapeutic range for serum flecainide concentration is lower in HapB carriers than in HapA homozygotes.

Stereoselective analysis of flecainide enantiomers using reversed-phase liquid chromatography for assessing CYP2D6 activity

Stereoselective analyses of flecainide enantiomers were performed using reversed-phase high-performance liquid chromatography (HPLC) equipped with a polysaccharide-based chiral column (Chiralpak AS-RH) and fluorescence detector. Excitation and emission wavelengths were set at 300 and 370 nm, respectively. Flecainide enantiomers in serum and urine were extracted using diethyl ether. The mobile phase solution, comprising 0.1 m potassium hexafluorophosphate and acetonitrile (65:35, v/v), was pumped at a flow rate of 0.5 mL/min. The recoveries of flecainide enantiomers were greater than 94%, with the coefficients of variation (CVs) <6%. The calibration curves of flecainide enantiomers in serum and urine were linear in the concentration range 5-500 ng/mL and 0.75-15 µg/mL (r > 0.999), respectively. CVs in intra-day and inter-day assays were 1.8-5.8 and 3.4-7.5%, respectively. In a pharmacokinetic study, the ratios of (S)- to (R)-flecainide (S/R ratio) in the area under the curve and the amount of flecainide enantiomers excreted in urine were lower in a subject carrying CYP2D6*10/*10 than in subjects carrying CYP2D6*1/*2. The S/R ratio of trough serum flecainide concentration ranged from 0.79 to 1.16 in patients receiving oral flecainide. The present HPLC method can be used to assess hepatic flecainide metabolism in a pharmacokinetic study and therapeutic drug monitoring.