Dihydropyrimidine dehydrogenase (DPD) expression is negatively regulated by certain microRNAs in human lung tissues.

Dihydropyrimidine dehydrogenase (DPD) is important to the antitumor effect of 5-fluorouracil (5-FU). DPD gene (DPYD) expression in tumors is correlated with sensitivity to 5-FU. Because the 5-FU accumulated in cancer cells is also rapidly converted into inactivated metabolites through catabolic pathways mediated by DPD, high DPD activity in cancer cells is an important determinant of the response to 5-FU. DPD activity is highly variable and reduced activity causes a high risk of 5-FU toxicity. Genetic variation in DPYD has been proposed as the main factor responsible for the variation in DPD activity. However, only a small proportion of the activity of DPD can be explained by DPYD mutations. In this study, we found that DPYD is a target of the following microRNAs (miRNA): miR-27a, miR-27b, miR-134, and miR-582-5p. In luciferase assays with HepG2 cells, the overexpression of these miRNAs was associated with significantly decreased reporter activity in a plasmid containing the 3'-UTR of DYPD mRNA. The level of DPD protein in MIAPaca-2 cells was also significantly decreased by the overexpression of these four miRNAs. The results suggest that miR-27a, miR-27b, miR-134, and miR-582-5p post-transcriptionally regulate DPD protein expression. The levels of miRNAs in normal lung tissue and lung tumors were compared; miR-27b and miR-134 levels were significantly lower in the tumors than normal tissue (3.64 ± 4.02 versus 9.75 ± 6.58 and 0.64 ± 0.75 versus 1.48 ± 1.39). DPD protein levels were significantly higher in the tumors. Thus, the decreased expression of miR-27b would be responsible for the high levels of DPD protein. This study is the first to show that miRNAs regulate the DPD protein, and provides new insight into 5-FU-based chemotherapy.

Interindividual differences in placental expression of the SLC22A2 (OCT2) gene: relationship to epigenetic variations in the 5'-upstream regulatory region.

Organic cation transporters (OCTs) mediate the transport of organic cations and some drugs (e.g., metformin and cimetidine). OCT1, OCT2, and OCT3 are located in the imprinting cluster of the insulin-like growth factor 2 receptor. It has been reported that OCT1 and OCT3 show a biallelic expression, whereas OCT2 undergoes maternal imprinting in the human placenta; however, a loss of the imprinting of OCT2 has recently been reported in some placental samples. This study investigated whether epigenetic mechanisms are involved in interindividual differences in the placental expression of OCT2. Because OCT2 mRNA levels were higher in biallelic samples than that in monoallelic samples, we compared the DNA methylation and chromatin modifications in the promoter regions. There was no remarkable difference in DNA methylation between the mono allelic samples and biallelic samples. In contrast, histone H3 acetylation (H3Ac) was increased in the biallelic samples. A significant negative correlation was observed between the trimethylation of lysine-9 on histone H3 (H3K9me3) and the OCT2 mRNA levels. Our results suggest that H3Ac plays a role in the allelic expression of OCT2. In addition, H3K9me3 in the OCT2 promoter may explain the interindividual differences in placental OCT2 mRNA levels.

Population pharmacokinetic and pharmacodynamic analysis of linezolid and a hematologic side effect, thrombocytopenia, in Japanese patients.

Linezolid is an antimicrobial agent to treat infections by Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA). While effective, linezolid treatment frequently is associated with hematological side effects, especially thrombocytopenia. However, little is known about the mechanism of this side effect and the exposure-response relationship. The present population pharmacokinetic/pharmacodynamic (PPK/PD) study was undertaken to elucidate the factors that determine linezolid levels, the relationship between exposure to linezolid and a decrease in platelet counts, and appropriate dosage adjustments based on exposure levels. In total, 50 patients (135 plasma samples) were used for the PPK analysis. The PPK analysis revealed that renal function and severe liver cirrhosis (Child Pugh grade C) significantly affect the pharmacokinetics of linezolid according to the equation clearance (liter/h)=2.85×(creatinine clearance/60.9)0.618×0.472CIR (CIR indicates cirrhosis status; 0 for noncirrhosis, 1 for cirrhosis patients). Using 603 platelet counts from 45 patients, a PPK/PD analysis with a semimechanistic pharmacodynamic model described the relationship between linezolid exposure and platelet counts quantitatively, and the newly constructed model was validated using external data (776 platelet counts from 60 patients). Simulation indicated considerable risks in patients with insufficient renal function (creatinine clearance, ≤30 ml/min) or severe liver cirrhosis. For these patients, a reduced dosage (600 mg/day) would be recommended for sufficient efficacy (area under the concentration-time curve over 24 h in the steady state divided by the MIC, >100) and safety.

Life-threatening toxicities in a patient with UGT1A1*6/*28 and SLCO1B1*15/*15 genotypes after irinotecan-based chemotherapy.

INTRODUCTION: To explore severe toxicities induced by irinotecan-based chemotherapy and UGT1A1*6/*28 and SLCO1B1*15/*15 genotypes. CASE REPORT: A 66-year-old Japanese male diagnosed with left pharyngeal carcinoma (T2N2bM0, stage IVA) was treated with irinotecan (70 mg/m(2)) on days 1, 8 and 15 in combination with docetaxel (60 mg/m(2)) on day 1 of a 28-day cycle. After the first cycle, he suffered marked toxicities, including grade 4 diarrhea and febrile grade 4 neutropenia. Plasma concentrations of irinotecan, SN-38 and SN-38G were measured, and extensive accumulation of SN-38 was observed. Genotyping of UGT1A1 and OATP1B1 proteins showed UGT1A1*6/*28 and SLCO1B1*15/*15, respectively, which are known to lead to extremely low glucuronidation and transport activities of substrate drugs. CONCLUSION: The severe toxicities in this patient are attributable to the extensive accumulation of SN-38, which may result from a synergistic or additive effect of low metabolic (UGT1A1*6/*28) and transport (SLCO1B1*15/*15) capabilities.

Suitable blending method of lipiodol-cisplatin in transcatheter arterial embolization for hepatocellular carcinoma: evaluation of sustained release and accumulation nature.

BACKGROUND/AIMS: Recently, fine powder cisplatin (IA-call; Nipponkayaku, Japan) was released, but there is no detailed study on an appropriate blending method of lipiodol-cisplatin for transcatheter arterial chemoembolization. We evaluated the sustained release and accumulation nature of lipiodol-cisplatin for hepatocellular carcinoma. METHODOLOGY: We prepared three types of mixture: a suspension of lipiodol and cisplatin powder, an emulsion of cisplatin powder dissolved with contrast medium and lipiodol, and a suspension-emulsion that was a suspension of lipiodol and cisplatin powder emulsified with contrast medium. In a basic study, a cisplatin release test was performed. In a clinical evaluation, transcatheter arterial chemoembolization with 3 lipiodol-cisplatin mixtures that had sustained release was performed in 60 consecutive patients with hepatocellular carcinoma as a randomized controlled trial. The density ratio of the tumor and treated liver tissue was measured as the accumulation nature. RESULTS: The suspension-emulsion and emulsion with a 7:3 mixture of lipiodol and contrast medium, and the suspension had better sustained release. The accumulation nature of the suspension-emulsion and emulsion were higher than the suspension. CONCLUSIONS: The pattern efficiently accumulated on hepatocellular carcinoma was the suspension-emulsion and emulsion. We suggest that a suspension-emulsion may be created more easily and is more suitable for clinical use.

Polymorphism in human organic cation transporters and metformin action.

Considerable interindividual variabilities in clinical efficacy and adverse events are sometimes recognized in the treatment of Type 2 diabetes mellitus with oral antihyperglycemic drugs. Metformin is the most commonly used biguanide in clinical practice, and also improves insulin resistance and reduces cardiovascular risk. However, certain patients taking metformin do not respond sufficiently. The molecular reasons for the variability in response to metformin are not clear. However, it has been recently suggested that genetic factors may be responsible for the variability. Metformin is not metabolized but is transported by at least two organic cation transporters (OCT), OCT1 and OCT2. Recently, genetic polymorphisms in OCT 1 and OCT2 have been found to be associated with changes in pharmacokinetic/pharmacodynamic responses to substrate drugs. This review focuses on the impact of the genetic polymorphism of organic cation transporters on transport activity, and the implications for the clinical efficacy of metformin.

Severe toxicities after irinotecan-based chemotherapy in a patient with lung cancer: a homozygote for the SLCO1B1*15 allele.

Irinotecan is used widely in the treatment of several malignancies, but unpredictable severe toxicities such as myelosuppression and delayed-type diarrhea are sometimes experienced. Polymorphism of the UGT1A1 gene is one of the likely reasons for interindividual differences in irinotecan pharmacokinetics and severe toxicity. Also, polymorphic organic anion-transporting polypeptide 1B1 (OATP1B1, SLCO1B1) is reported to be involved in the hepatocellular uptake of SN-38. A 61-year-old man with lung cancer developed severe toxicities, including grade 3 diarrhea, grade 4 leukopenia, and grade 4 neutropenia, after the first cycle of irinotecan (60 mg/m) plus cisplatin chemotherapy. The irinotecan and SN-38 areas under the concentration-time curve from time zero to infinity in this patient were 43% and 87% higher than the corresponding mean values for 10 other patients with lung cancer treated with irinotecan (60-100 mg/m) normalized for the dose of irinotecan. Analysis of genetic variants in genes encoding the drug-metabolizing enzyme (UGT1A1) and transporter (SLCO1B1) involving irinotecan disposition revealed that this patient was homozygous for the SLCO1B1*15 allele, which may result in severe toxicities attributable to the extensive accumulation of SN-38. Screening of SLCO1B1*15 is suggested to be useful in irinotecan chemotherapy to avoid unpredicted severe toxicity, although the homozygous genotype is rare among the Japanese.

Human organic cation transporter (OCT1 and OCT2) gene polymorphisms and therapeutic effects of metformin.

Organic cation transporters (OCTs) are responsible for the hepatic and renal transport of metformin. In this study we analyzed variants of OCT1 and OCT2 genes in 33 patients (24 responders and nine non-responders) based on the hypothesis that polymorphisms in both genes contribute to large inter-patient variability in the clinical efficacy of metformin. The sequences of the 5'-flanking and coding regions of the two genes of interest were screened by single-strand conformation polymorphism (SSCP) analysis. To compare the causative factors between responders and non-responders, we performed stepwise discriminant functional analysis. Age, body mass index (BMI) and treatment with lipid-lowering agents were demonstrated as positive predictors, and two mutations in the OCT1 gene, -43T > G in intron 1 and 408Met > Val (1222A > G) in exon 7, were negative and positive predictors, respectively, for the efficacy of metformin; the predictive accuracy was 55.5% (P < 0.05). Subsequent study indicated that OCT1 mRNA levels tended to be lower in human livers with the 408Met (1222A) variant, though the differences did not reach the level of significance. In this study it is suggested that OCT1 and OCT2 gene polymorphisms have little contribution to the clinical efficacy of metformin.

Genetic polymorphisms of drug transporters: pharmacokinetic and pharmacodynamic consequences in pharmacotherapy.

There has been increasing appreciation of the role of drug transporters in pharmacokinetic and pharmacodynamic consequences in pharmacotherapy. The clinical relevance of drug transporters depends on the localisation in human tissues (i.e., vectorial movement), the therapeutic index of the substrates and inherent interindividual variability. With regard to variability, polymorphisms of drug transporter genes have recently been reported to be associated with alterations in the pharmacokinetics and pharmacodynamics of clinically useful drugs. A growing number of preclinical and clinical studies have demonstrated that the application of genetic information may be useful in individualised pharmacotherapy for numerous diseases. However, the reported effects of variants in certain drug transporter genes have been inconsistent and, in some cases, conflicting among studies. Furthermore, the incidence of almost all known variants in transporter genes tends to be racially dependent. These observations suggest the necessity of considering interethnic variability before extrapolating pharmacokinetic data obtained in one ethic group to another, especially in the early phase of drug development. This review focuses on the impact of genetic variations in the function of drug transporters (ABC, organic anion and cation transporters) and the implications of these variations for pharmacotherapy from pharmacokinetic and pharmacodynamic viewpoints.

Pharmacogenetic determinants of variability in lipid-lowering response to pravastatin therapy.

Pravastatin is mainly taken up from the circulation into the liver via organic anion-transporting polypeptide 1B1 (SLCO1B1 gene product). We examined the contribution of genetic variants in the SLCO1B1 gene and other candidate genes to the variability of pravastatin efficacy in 33 hypercholesterolemic patients. In the initial phase of pravastatin treatment (8 weeks), heterozygous carriers of the SLCO1B1*15 allele had poor low-density lipoprotein cholesterol (LDL-C) reduction relative to non-carriers (percent reduction: -14.1 vs -28.9%); however, the genotype-dependent difference in the cholesterol-lowering effect disappeared after 1 year of treatment. Cholesterol 7alpha-hydroxylase (CYP7A1) and apolipoprotein E (APOE) are known to contribute to lipid metabolism. Homozygous carriers of the CYP7A1 -204C allele or heterozygotes for both CYP7A1 -204C and APOE epsilon4 alleles showed significantly poorer LDL-C reduction compared to that in other genotypic groups after 1 year of treatment (-24.3 vs -33.1%). These results suggest that the SLCO1B1*15 allele is associated with a slow response to pravastatin therapy, and the combined genotyping of CYP7A1 and APOE genes is a useful index of the lipid-lowering effect of pravastatin.

Effects of organic anion transporting polypeptide 1B1 haplotype on pharmacokinetics of pravastatin, valsartan, and temocapril.

OBJECTIVE: Recent reports have shown that genetic polymorphisms in organic anion transporting polypeptide (OATP) 1B1 have an effect on the pharmacokinetics of drugs. However, the impact of OATP1B1*1b alleles, the frequency of which is high in all ethnicities, on the pharmacokinetics of substrate drugs is not known after complete separation of subjects with OATP1B1*1a and *1b. Furthermore, the correlation between the clearances of OATP1B1 substrate drugs in individuals has not been characterized. We investigated the effect of genetic polymorphism of OATP1B1, particularly the *1b allele, on the pharmacokinetics of 3 anionic drugs, pravastatin, valsartan, and temocapril, in Japanese subjects. METHODS: Twenty-three healthy Japanese volunteers were enrolled in a 3-period crossover study. In each period, after a single oral administration of pravastatin, valsartan, or temocapril, plasma and urine were collected for up to 24 hours. RESULTS: The area under the plasma concentration-time curve (AUC) of pravastatin in *1b/*1b carriers (47.4 +/- 19.9 ng.h/mL) was 65% of that in *1a/*1a carriers (73.2 +/- 23.5 ng.h/mL) (P = .049). Carriers of *1b/*15 (38.2 +/- 15.9 ng.h/mL) exhibited a 45% lower AUC than *1a/*15 carriers (69.2 +/- 23.4 ng.h/mL) (P = .024). In the case of valsartan we observed a similar trend as with pravastatin, although the difference was not statistically significant (9.01 +/- 3.33 microg.h/mL for *1b/*1b carriers versus 12.3 +/- 4.6 microg.h/mL for *1a/*1a carriers [P = .171] and 6.31 +/- 3.64 microg.h/mL for *1b/*15 carriers versus 9.40 +/- 4.34 microg.h/mL for *1a/*15 carriers [P = .213]). The AUC of temocapril also showed a similar trend (12.4 +/- 4.1 ng.h/mL for *1b/*1b carriers versus 18.5 +/- 7.7 ng.h/mL for *1a/*1a carriers [P = .061] and 16.4 +/- 5.0 ng.h/mL for *1b/*15 carriers versus 19.0 +/- 4.1 ng.h/mL for *1a/*15 carriers [P = .425]), whereas that of temocaprilat (active form of temocapril) was not significantly affected by the haplotype of OATP1B1. Interestingly, the AUC of valsartan and temocapril in each subject was significantly correlated with that of pravastatin (R = 0.630 and 0.602, P < .01). The renal clearance remained unchanged for each haplotype for all drugs. CONCLUSION: The major clearance mechanism of pravastatin, valsartan, and temocapril appears to be similar, and OATP1B1*1b is one of the determinant factors governing the interindividual variability in the pharmacokinetics of pravastatin and, possibly, valsartan and temocapril.

Different contributions of polymorphisms in VKORC1 and CYP2C9 to intra- and inter-population differences in maintenance dose of warfarin in Japanese, Caucasians and African-Americans.

OBJECTIVE: To investigate pharmacokinetic and pharmacodynamic factors associated with population differences in warfarin doses needed to achieve anticoagulation, in particular the possible involvement of genetic variability in vitamin K epoxide reductase (VKOR) and CYP2C9. METHODS: Warfarin maintenance dose, unbound plasma S-warfarin concentration [Cu(S)] and INR were determined in 157 Caucasians, 172 Japanese, and 36 African-Americans stably anticoagulated patients. In a subset (n = 166), fully carboxylated plasma normal prothrombin levels (NPT) were also measured. Genotyping for seven CYP2C9 (CYP2C9*1 through 6 and *11) and seven VKORC1 variants were performed in 115 Caucasians and 64 Japanese patients and 66 healthy African-Americans. Multivariate analysis was performed to identify covariates associated with warfarin requirement. RESULTS: The relationship between NPT and Cu(S) indicated Japanese are more susceptible to inhibition of NPT production by S-warfarin than the other two populations. VKORC1 1173 C > T had a greater frequency in Japanese (89.1%) than Caucasians (42.2%) and African-Americans (8.6%). CYP2C9 variants with reduced metabolizing ability were less frequent in Japanese compared to the other two populations. The median warfarin dose was significantly higher in Caucasians than Japanese patients (5.5 versus 3.5 mg/day), however, when matched for CYP2C9*1 homozygosity, no difference in dose was observed between VKORC1 genotype-matched groups. Furthermore, VKORC1 1173C > T and CYP2C9 (*2/*3/*11) genotypes, age and weight were identified as independent covariates contributing to interpatient variability in warfarin dosage. CONCLUSIONS: Both VKORC1 and CYP2C9 polymorphisms contribute to inter-population difference in warfarin doses among the three populations, but their contribution to intra-population variability may differ within each population.

Interaction magnitude, pharmacokinetics and pharmacodynamics of ticlopidine in relation to CYP2C19 genotypic status.

OBJECTIVES: The aim of this study was to investigate the impact of CYP2C19 polymorphism on the extent of the interaction and on the pharmacokinetics and pharmacodynamics of ticlopidine. METHODS: Homozygous (hmEMs) and heterozygous extensive metabolizers (htEMs), and poor metabolizers (PMs, n = 6 each) took an oral dose (20 mg) of omeprazole. After a 1-week washout period, each subject received ticlopidine (200 mg) for 8 days, and ticlopidine pharmacokinetics were studied on days 1 and 7. On day 8, omeprazole was given again and its kinetic disposition was compared with that in the first dose. ADP-induced platelet aggregation was measured as a pharmacodynamic index. RESULTS: In contrast to the PMs, whose mean kinetic parameters were not altered by the repeated dosings of ticlopidine, an eight- to 10-fold increase in the mean AUC ratio of omeprazole to 5-hydroxyomeprazole was observed in both the EM groups. No significant intergenotypic differences in the pharmacokinetic parameters of ticlopidine were observed, although the accumulation ratio tended to be greater in hmEMs than in PMs (2.4 +/- 0.2 versus 1.7 +/- 0.2). A significantly positive correlation (P = 0.031) was observed between the individual percent inhibition of platelet aggregation and AUC0-24 of ticlopidine regardless of the CYP2C19 polymorphism. CONCLUSIONS: Ticlopidine is a potent inhibitor for CYP2C19 and may be associated with the phenocopy when CYPC19 substrates are co-administered to EMs. Whether and to what extent CYP2C19 would be involved in the metabolism of ticlopidine remain unanswered from the present in-vivo study.

Functional assessment of ABCG2 (BCRP) gene polymorphisms to protein expression in human placenta.

The aim of the present study was to assess the contribution of polymorphisms in the breast cancer resistance protein/ATP-binding cassette transporter G2 (BCRP/ABCG2) gene to the placental expression from a new perspective, allelic imbalance. Polymorphisms were screened by polymerase chain reaction (PCR)-single-strand conformation polymorphism analysis followed by sequencing with DNA extracted from 100 placentas. To examine whether polymorphisms of the BCRP gene correlate with the placental BCRP expression, we determined mRNA and protein levels by quantitative real-time PCR and Western blotting, respectively. In placentas, G34A (Val(12)Met) and C421A (Gln(141)Lys) were frequently observed (18-36%), but C376T, which creates a stop codon (Gln(126) stop codon), was found with an allelic frequency of 1%. The mean of the BCRP protein level was significantly lower (p < 0.05) in homozygotes for the A421 allele than in those for the C421 allele, and heterozygotes had an intermediate value. To evaluate whether the C421A polymorphism acts as a cis-element in BCRP transcription, allelic imbalance was determined using informative lymphoblasts and 56 samples of placental cDNA. In most of the placental samples we tested, the difference in expression levels between the two alleles was small, and only two samples indicated a monoallelic expression (i.e., preferential expression of one allele). These results suggest that 1) the predominant allelic expression pattern of BCRP in placental samples is biallelic, and 2) the mutation C421A is not a genetic variant acting in cis, but is considered to influence the translation efficiency.

Influence of common variants in the pharmacokinetic genes (OATP-C, UGT1A1, and MRP2) on serum bilirubin levels in healthy subjects.

To assess the contribution of OATP-C to the hepatobiliary transport of bilirubin, a pharmacogenomic evaluation with regard to polymorphisms of three candidate genes, OATP-C, MRP2, and UGT1A1, was performed. Serum total and direct (conjugated) bilirubin levels were used as phenotypic indexes. Pharmacokinetic variables of pravastatin, a typical substrate for OATP-C, were obtained from our previous study. Among 23 volunteers, two variants (Val417Ile and Ser789Phe) were observed in the MRP2 gene. While there was no apparent effect of these two variants and the UGT1A1*28 on direct bilirubin levels, the OATP-C variants were associated with differences in unconjugated bilirubin levels. Subjects with the OATP-C*15 allele had higher bilirubin levels; unconjugated bilirubin levels in *1b/*1b (n = 3), *1b/*15 (n= 7), and *15/*15 (n = 1) subjects were 0.40 +/- 0.10, 0.77 +/- 0.35, and 0.70 (mg/dL), respectively. In addition, the correlation between unconjugated bilirubin levels and pharmacokinetic parameters of pravastatin revealed that the subjects with higher bilirubin levels had lower non-renal clearance values, and then higher serum concentrations of pravastatin. Large clinical studies are needed to confirm a role of OATP-C in the carrier-mediated uptake of bilirubin in the human liver.

Functional analysis of single nucleotide polymorphisms of hepatic organic anion transporter OATP1B1 (OATP-C).

OBJECTIVE: Two kinds of single nucleotide polymorphism (SNP; Asn130Asp and Val174Ala) are frequently observed in the liver specific transporter, organic anion transporting polypeptide 1B1 (OATP1B1/OATP-C) gene. Although these two SNPs occur independently in European-Americans, Val174Ala is mostly associated with Asn130Asp in Japanese. Our previous in-vivo studies in Japanese subjects indicated that the non-renal clearance of pravastatin was decreased to 13% of that in wild-type subjects (Nishizato et al. Clin Pharmacol Ther 2003;73(6):554-564). The purpose of the present study is to characterize the function of SNPs variants of OATP1B1 in cDNA transfected cells. METHODS: The localization and transport activity were analyzed in HEK293 cells stably expressing wild-type OATP1B1 (OATP1B1*1a), OATP1B1*1b (Asn130Asp), OATP1B1*5 (Val174Ala) and OATP1B1*15 (Asn130Asp and Val174Ala). To characterize the intrinsic Vmax, observed Vmax in uptake study were normalized by the expression level estimated from Western blotting. RESULTS: All SNP variants are predominantly located on the cell surface. No significant alteration was observed in Km values for the transport of 17beta-estradiol 17beta-d-glucuronide (E217betaG), a typical substrate of OATP1B1, among these SNP variants. However, the normalized Vmax value for OATP1B1*15 was drastically decreased to less than 30% compared with OATP1B1*1a. In contrast, the transport activity of OATP1B1*1b (Asn130Asp) and OATP1B1*5 (Val 174Ala) was similar to that of OATP1B1*1a. CONCLUSIONS: These results are consistent with the results of our previous clinical studies. It is thus suggested that in-vivo disposition may be predicted from in-vitro results using recombinant transporters.

Allelic expression imbalance of the human CYP3A4 gene and individual phenotypic status.

The human cytochrome P450 3A4 (CYP3A4) plays a dominant role in the metabolism of numerous clinically useful drugs. Alterations in the activity or expression of this enzyme may account for a major part of the variation in drug responsiveness and toxicity. However, it is generally accepted that most of the known single nucleotide polymorphisms in the coding and 5'-flanking regions are not the main determinants for the large inter-individual variability of CYP3A4 expression and activity. We show that the allelic variation is critically involved in determining the individual total hepatic CYP3A4 mRNA level and metabolic capability. There exists a definite correlation between the total CYP3A4 mRNA level and allelic expression ratio, the relative transcript level ratio derived from the two alleles. Individuals with a low expression ratio, exhibiting a large difference of transcript level between the two alleles, revealed extremely low levels of total hepatic CYP3A4 mRNA, and thus low metabolic capability as assessed by testosterone 6beta-hydroxylation. These results present a new insight into the individualized CYP3A4-dependent pharmacotherapy and the importance of expression imbalance to human phenotypic diversity.

Urodynamic effects and safety of modified intravesical oxybutynin chloride in patients with neurogenic detrusor overactivity: 3 years experience.

BACKGROUND: Intravesical oxybutynin chloride with hydroxypropylcellulose (HPC) (modified intravesical oxybutynin) has been reported to be effective for treatment of overactive bladder. We reported the short-term effects of modified intravesical oxybutynin previously. In the present article, we detail the results of a 3-year follow-up study of patients from our previous analysis and report the efficacy and side-effects of modified intravesical oxybutynin. METHODS: Modified intravesical oxybutynin (5 mg/10 mL, twice a day) was applied for more than 3 years to six neurogenic overactive detrusor patients (three men and three women, average age 53.3 years) who were not satisfied with oral anticholinergic agents or the other therapy. A cystometogram (CMG) was performed before, 1 week after and 3 years after the start of modified intravesical oxybutynin treatment. We evaluated the patient's satisfaction of this treatment after 4 weeks and again after 3 years. We compared the patients' answers before and after the therapy (excellent, good, fair, unchanged and worse). We also monitored systemic and topical side-effects in these patients during this period. RESULTS: CMG studies showed that two of six patients no longer exhibited uninhibited contraction 1 week after the treatment and that the cystocapacity of patients before, 1 week after and 3 years after the initial modified intravesical oxybutynin was 129.7 +/- 19.4, 283.5 +/- 40.4 and 286.8 +/- 38.1 mL, respectively. For the evaluation of patients' satisfaction with this treatment, four patients considered the therapy excellent and one patient described it as good after both 4 weeks and after 3 years. Two patients dropped out of the study; one developed left ureteral cancer (2.25 years) and the other developed ileus (1.5 years). Dry mouth and acute cystitis were observed in both patients. CONCLUSION: Modified intravesical oxybutynin is an effective and relatively safe option of therapy for overactive bladder patients. However, this therapy requires careful observation for emergent side-effects.

Haplotype-oriented genetic analysis and functional assessment of promoter variants in the MDR1 (ABCB1) gene.

Recently, a number of nucleotide variants have been described in the multidrug resistance 1 (MDR1/ABCB1) gene; however, most studies have focused on the coding region. In the present study, we identified promoter variants of the MDR1 gene and evaluated their phenotypic consequences using a reporter gene assay and the real-time polymerase chain reaction method. Ten allelic variants were detected in the promoter region (approximately 2 kilobases), seven of which were newly identified. Certain mutations occurred simultaneously, and a total of 10 haplotypes were observed. These promoter polymorphisms were found more frequently in Japanese than Caucasians. Some haplotypes were associated with changes in luciferase activity and placental and hepatic mRNA levels. We also determined DNA methylation status in the proximal promoter region of the MDR1 gene. The promoter region around potential binding sites for transcription factors was found to be hypomethylated and thus likely to be independent of the gene expression. Nucleotide and/or haplotype variants not only in the coding region but also in the promoter region of the MDR1 gene may be important for interindividual differences of P-glycoprotein expression.