Papers - Yonezawa, Atsushi

Clinical Outcomes of Vancomycin Therapy by the Protocol-Based Pharmacotherapy Management (PBPM) including TDM-Order

KATADA YOSHIKI, NAKAGAWA SHUNSAKU, TAUE HIROMI, TSUDA MASAHIRO, TSUZUKI TETSUNORI, HATA SACHIYO, ODAKA MIZUHO, YONEZAWA ATSUSHI, KAYANO YUICHIRO, YANO IKUKO, MINAKATA KENJI, SAKATA RYUZO, MATSUBARA KAZUO

医療薬学 ((一社)日本医療薬学会)  42 ( 1 ) 14 - 22 2016.01

Research paper (scientific journal), Joint Work, Accepted,  ISSN  1346-342X

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心臓血管外科病棟におけるバンコマイシン(VCM)治療に焦点を当てて、プロトコルの基づく薬物治療管理(PBPM)の臨床的アウトカムを定量化し、VCM治療における薬剤師介入の効果について検討した。VCMが開始され治療薬物モニタリング(TDM)を行った54例を対象とし、プロトコル導入以前にVCM治療を行った29例(男性21名、女性8名、70±18歳)(I群)、プロトコル導入後にVCM治療を行った25例(男性17名、女性8名、67±15歳)(II群)に分けた。急性腎障害発現はI群9例(31.0%)、II群2例(8.0%)であった。治療域維持率の中央値は、I群50%、II群80%、中毒域到達率の中央値は、I群17%、II群0%であった。治療域維持率及び中毒域到達率に関する2群間の差は、敗血症、肺炎、感染性心内膜炎の背景を有する場合において大きい値であった。VCM血中濃度の日内変動はI群で大きい傾向があった。

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Harnessing the innate immune system to treat cancer: Enhancement of antibody-dependent cellular cytotoxicity with anti-CD137 Ab

Atsushi Yonezawa, Cariad Chester, Narendiran Rajasekaran, Holbrook E. Kohrt

Chinese Clinical Oncology (AME Publishing Company)  5 ( 1 ) 5 2016

Research paper (scientific journal), Joint Work, Lead author, Accepted,  ISSN  23043865

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Background: Monoclonal antibodies are an invaluable therapeutic class when it comes to cancer therapy. One of the primary mechanisms of anti-tumor activity of monoclonal antibodies is antibody dependent cell-mediated cytotoxicity (ADCC) mediated by natural killer (NK) cells. Innate immune effector cells play important roles in generating and maintaining antitumor immunity to enhance ADCC. Methods: Efficacy of anti-CD137 mAb was examined by in vitro and in vivo experiments. Clinical trials of anti-CD137 mAb are also on-going. Results: CD137 is a costimulatory receptor that belongs to the tumor necrosis factor receptor (TNFR) superfamily. CD137 is expressed on T cell, activated NK cells and other immune cells. We recently showed that an agonistic anti-CD137 mAb activates NK cells, resulting in increased ADCC against cancer cells. Anti-CD137 agonistic mAb therapy may both increase ADCC due to mAb-activated NK cells and promote the proliferation and cytotoxicity of antigen-specific T cells induced by mAb treatment. Conclusions: We discuss some of the promising strategies that could potentially enhance ADCC with anti-CD137 mAbs.

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Population Pharmacokinetic Modeling of Levetiracetam in Pediatric and Adult Patients With Epilepsy by Using Routinely Monitored Data

Satoko Ito, Ikuko Yano, Sachiyo Hashi, Masahiro Tsuda, Mitsuhiro Sugimoto, Atsushi Yonezawa, Akio Ikeda, Kazuo Matsubara

THERAPEUTIC DRUG MONITORING (LIPPINCOTT WILLIAMS & WILKINS)  38 ( 3 ) 371 - 378 2016

Research paper (scientific journal), Joint Work, Accepted,  ISSN  01634356

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Background:Levetiracetam, a second-generation antiepileptic drug, is frequently used for managing partial-onset seizures. About 70% of the administered dose is excreted in urine unchanged, and dosage adjustment is recommended based on the individual's renal function. In this study, a population pharmacokinetic model of levetiracetam was developed using routinely monitored serum concentration data for individualized levetiracetam therapy.Methods:Patients whose serum concentrations of levetiracetam at steady-state were routinely monitored at Kyoto University Hospital from April 2012 to March 2013 were enrolled. The influence of patient characteristics on levetiracetam pharmacokinetics was evaluated using the nonlinear mixed-effects modeling (NONMEM) program.Results:A total of 583 steady-state concentrations from 225 patients were used for the analysis. The median patient age and estimated glomerular filtration rate (eGFR) were 38 (range: 1-89) years and 98 (15-189) mLmin(-1)1.73 m(-2), respectively. Serum concentration-time data of levetiracetam were well described by a 1-compartment model with first-order absorption. Oral clearance was allometrically related to the individual body weight and eGFR. An increase in the dose significantly increased oral clearance. No improvement in model fit was observed by including the covariate of any concomitant antiepileptic drugs. The population mean clearance for an adult weighing 70 kg and with a normal renal function was 4.8 and 5.9 L/h for 500 mg bis in die (bid) and 1500 mg bid, respectively.Conclusions:Oral clearance allometrically related with body weight and eGFR can well predict the routine therapeutic drug monitoring data from pediatric to aged patients with varying renal function. Dosage adjustments based on renal function are effective in controlling the trough and peak concentrations in similar ranges.

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Pharmacokinetics and pharmacodynamics of mycophenolic acid in Nagase analbuminemic rats: Evaluation of protein binding effects using the modeling and simulation approach

Kazuaki Yoshimura, Ikuko Yano, Misaki Kawanishi, Shunsaku Nakagawa, Atsushi Yonezawa, Kazuo Matsubara

DRUG METABOLISM AND PHARMACOKINETICS (JAPANESE SOC STUDY XENOBIOTICS)  30 ( 6 ) 441 - 448 2015.12

Research paper (scientific journal), Joint Work, Accepted,  ISSN  13474367

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This study aimed to examine the pharmacokinetics and pharmacodynamics of mycophenolic acid (MPA) in Nagase analbuminemic rats (NARs) to evaluate the effect of protein binding on the associated inosine-5'-monophosphate dehydrogenase (IMPDH) activity. Free fractions of MPA in the control rats and NARs were 2.09 and 24.8%, respectively. Pharmacokinetic and pharmacodynamic parameters simultaneously obtained by the nonlinear mixed effects modeling program NONMEM explained reasonably well the concentrations of MPA and MPA glucuronide as well as IMPDH activity in both rats. NARs showed a higher clearance and a smaller volume of distribution based on the free MPA concentration than the controls did, besides the increase in free fraction. The half-maximal inhibitory concentration based on free MPA was estimated as 163 ng/mL for both rats. Simulations based on the obtained pharmacokinetic and pharmacodynamic parameters showed that the area under the IMPDH activity-time curve decreased non-linearly according to the increase in free fraction of MPA. In conclusion, the experimental data obtained from NARs followed by the modeling and simulation approach quantitatively clarified that the free MPA concentration was suitable for the biomarker of immunosuppressive effect of MPA. Dose adjustments based on the total MPA may cause unnecessary overexposure to MPA in patients with hypoalbuminemia. Copyright (C) 2015, The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

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Gefitinib and Erlotinib Lead to Phosphorylation of Eukaryotic Initiation Factor 2 Alpha Independent of Epidermal Growth Factor Receptor in A549 Cells

Satoshi Koyama, Tomohiro Omura, Atsushi Yonezawa, Satoshi Imai, Shunsaku Nakagawa, Takayuki Nakagawa, Ikuko Yano, Kazuo Matsubara

PLOS ONE (PUBLIC LIBRARY SCIENCE)  10 ( 8 ) e0136176 2015.08

Research paper (scientific journal), Joint Work, Accepted,  ISSN  1932-6203

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Gefitinib and erlotinib are anticancer agents, which inhibit epidermal growth factor receptor (EGFR) tyrosine kinase. Interstitial lung disease (ILD) occurs in patients with non-small cell lung cancer receiving EGFR inhibitors. In the present study, we examined whether gefitinib- and erlotinib-induced lung injury related to ILD through endoplasmic reticulum (ER) stress, which is a causative intracellular mechanism in cytotoxicity caused by various chemicals in adenocarcinomic human alveolar basal epithelial cells. These two EGFR inhibitors increased Parkinson juvenile disease protein 2 and C/EBP homologous protein mRNA expressions, and activated the eukaryotic initiation factor (eIF) 2 alpha/activating transcription factor 4 pathway without protein kinase R-like ER kinase activation in A549 cells. Gefitinib and erlotinib caused neither ER stress nor cell death; however, these agents inhibited cell growth via the reduction of cyclin-D1 expression. Tauroursodeoxycholic acid, which is known to suppress eIF2 alpha phosphorylation, cancelled the effects of EGFR inhibitors on cyclin-D1 expression and cell proliferation in a concentration-dependent manner. The results of an EGFR-silencing study using siRNA showed that gefitinib and erlotinib affected eIF2 alpha phosphorylation and cyclin-D1 expression independent of EGFR inhibition. Therefore, the inhibition of cell growth by these EGFR inhibitors might equate to impairment of the alveolar epithelial cell repair system via eIF2 alpha phosphorylation and reduced cyclin-D1 expression.

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A novel protocol for antibiotic prophylaxis based on preoperative kidney function in patients undergoing open heart surgery under cardiopulmonary bypass

Mizuho Odaka, Kenji Minakata, Hideaki Toyokuni, Kazuhiro Yamazaki, Atsushi Yonezawa, Ryuzo Sakata, Kazuo Matsubara

General Thoracic and Cardiovascular Surgery (Springer Tokyo)  63 ( 8 ) 446 - 452 2015.08

Research paper (scientific journal), Joint Work, Accepted,  ISSN  18636705

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Objective: This study aimed to develop and assess the effectiveness of a protocol for antibiotic prophylaxis based on preoperative kidney function in patients undergoing open heart surgery. Methods: We established a protocol for antibiotic prophylaxis based on preoperative kidney function in patients undergoing open heart surgery. This novel protocol was assessed by comparing patients undergoing open heart surgery before (control group
n = 30) and after its implementation (protocol group
n = 31) at Kyoto University Hospital between July 2012 and January 2013. Results: Surgical site infections (SSIs) were observed in 4 control group patients (13.3 %), whereas no SSIs were observed in the protocol group patients (P &lt
 0.05). The total duration of antibiotic use decreased significantly from 80.7 ± 17.6 h (mean ± SD) in the control group to 55.5 ± 14.9 h in the protocol group (P &lt
 0.05). Similarly, introduction of the protocol significantly decreased the total antibiotic dose used in the perioperative period (P &lt
 0.05). Furthermore, antibiotic regimens were changed under suspicion of infection in 5 of 30 control group patients, whereas none of the protocol group patients required this additional change in the antibiotic regimen (P &lt
 0.05). Conclusion: Our novel antibiotic prophylaxis protocol based on preoperative kidney function effectively prevents SSIs in patients undergoing open heart surgery.

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Boosting Cancer Immunotherapy with Anti-CD137 Antibody Therapy

Atsushi Yonezawa, Suparna Dutt, Cariad Chester, Jeewon Kim, Holbrook E. Kohrt

CLINICAL CANCER RESEARCH (AMER ASSOC CANCER RESEARCH)  21 ( 14 ) 3113 - 3120 2015.07

Research paper (scientific journal), Joint Work, Lead author, Accepted,  ISSN  10780432

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In the past 5 years, immunomodulatory antibodies have revolutionized cancer immunotherapy. CD137, a member of the tumor necrosis factor receptor superfamily, represents a promising target for enhancing antitumor immune responses. CD137 helps regulate the activation of many immune cells, including CD4(+) T cells, CD8(+) T cells, dendritic cells, and natural killer cells. Recent studies indicate that the antitumor efficacy of therapeutic tumor-targeting antibodies can be augmented by the addition of agonistic antibodies targeting CD137. As ligation of CD137 provides a costimulatory signal in multiple immune cell subsets, combination therapy of CD137 antibody with therapeutic antibodies and/or vaccination has the potential to improve cancer treatment. Recently, clinical trials of combination therapies with agonistic anti-CD137 mAbs have been launched. In this review, we discuss the recent advances and clinical promise of agonistic anti-CD137 monoclonal antibody therapy. (C) 2015 AACR.

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Association of decreased mRNA expression of multidrug and toxin extrusion protein 1 in peripheral blood cells with the development of flutamide-induced liver injury

Kazuhiko Nakano, Hitoshi Ando, Shinsuke Kurokawa, Keiko Hosohata, Kentarou Ushijima, Makoto Takada, Masato Tateishi, Atsushi Yonezawa, Satohiro Masuda, Kazuo Matsubara, Ken-ichi Inui, Tatsuo Morita, Akio Fujimura

CANCER CHEMOTHERAPY AND PHARMACOLOGY (SPRINGER)  75 ( 6 ) 1191 - 1197 2015.06

Research paper (scientific journal), Joint Work, Accepted,  ISSN  03445704

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The anti-prostate cancer drug flutamide occasionally causes hepatotoxicity, and predictive biomarkers of flutamide-induced liver injury (FILI) are needed to improve safety of this drug. The aim of this prospective study was to identify such a biomarker by analyzing peripheral blood samples from patients before flutamide therapy.
Blood samples were obtained from 52 patients with prostate cancer before flutamide therapy. FILI was defined as treatment-related elevation of the serum concentration of aspartate or alanine aminotransferase to more than twice the upper limit of the reference range. The patients were monitored for at least 6 months regarding FILI. Microarray and quantitative real-time PCR analyses were conducted to compare gene expression profiles between the groups with and without FILI.
Seventeen patients developed FILI. Microarray analysis of the training set in 15 patients detected 11 annotated genes showing > twofold expression changes between the groups (p < 0.005). Quantitative PCR analysis of both the training set and validation set confirmed that mRNA levels of multidrug and toxin extrusion protein 1 (MATE1 or SLC47A1, encoded by 1 of the 11 genes) were significantly lower in patients with FILI. A small experiment on mice (three per group) showed that Mate1 knockout mice had an elevated serum concentration of 4-nitro-3-(trifluoromethyl)phenylamine, a major metabolite of flutamide, 2 h after administration of the drug, suggesting that Mate1 could affect the pharmacokinetics of flutamide.
The MATE1 mRNA level in peripheral blood is a possible negative predictive biomarker of FILI.

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Enhancement of antibody-dependent cell mediated cytotoxicity: a new era in cancer treatment.

Rajasekaran N, Chester C, Yonezawa A, Zhao X, Kohrt HE

ImmunoTargets and therapy 4   91 - 100 2015

Research paper (scientific journal), Joint Work, Accepted,  ISSN  2253-1556

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Extracellular nucleotides from dying cells act as molecular signals to promote wound repair in renal tubular injury

Shunsaku Nakagawa, Tomohiro Omura, Atsushi Yonezawa, Ikuko Yano, Takayuki Nakagawa, Kazuo Matsubara

American Journal of Physiology - Renal Physiology (American Physiological Society)  307 ( 12 ) F1404 - F1411 2014.12

Research paper (scientific journal), Joint Work, Accepted,  ISSN  1522-1466

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Acute kidney injury (AKI) often correlates with poor prognosis and is followed by various severe unfavorable systemic outcomes. It is important to understand the pathophysiology of AKI for the development of novel therapeutic approaches toward promoting renal regeneration after injury. Recent studies have indicated that AKI-induced tubular cell death plays an active role in the onset of tissue regeneration
however, the mechanisms underlying renal tubular repair after injury have yet to be understood. In the present study, we explored molecules that might serve as “danger” signals in mediating tubular regeneration. Kidneys of rats systemically administered the nephrotoxicant cisplatin (to induce AKI) exhibited massive cell proliferation. The proportion of proliferating cells in the total cell distribution was highest in the outer stripe of the outer medulla coincided with where the tubular damage was the most severe in this study. This finding suggests that soluble factors may have been released from damaged cells to stimulate the proliferation of neighboring tubular epithelial cells. In elucidating the mechanism of dying cell-to-surviving cell communication using normal rat kidney NRK-52E epithelial cells, we found a significant increase in ATP levels in supernatants of these cells after the induction of cell death using ultraviolet irradiation. Furthermore, treatment of conditioned supernatants with apyrase or suramin, which inhibits purinergic signaling, resulted in significant decreases in cell proliferation and migration activities. These results demonstrate a novel role for extracellular nucleotides, probably as danger signals in aggravating tubular regeneration after AKI.

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A randomized controlled clinical trial of topical insulin-like growth factor-1 therapy for sudden deafness refractory to systemic corticosteroid treatment

Takayuki Nakagawa, Kozo Kumakawa, Shin-ichi Usami, Naohito Hato, Keiji Tabuchi, Mariko Takahashi, Keizo Fujiwara, Akira Sasaki, Shizuo Komune, Tatsunori Sakamoto, Harukazu Hiraumi, Norio Yamamoto, Shiro Tanaka, Harue Tada, Michio Yamamoto, Atsushi Yonezawa, Toshiko Ito-Ihara, Takafumi Ikeda, Akira Shimizu, Yasuhiko Tabata, Juichi Ito

BMC MEDICINE (BIOMED CENTRAL LTD)  12 ( 1 ) 219 2014.11

Research paper (scientific journal), Joint Work, Accepted,  ISSN  1741-7015

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Background: To date, no therapeutic option has been established for sudden deafness refractory to systemic corticosteroids. This study aimed to examine the efficacy and safety of topical insulin-like growth factor-1 (IGF-1) therapy in comparison to intratympanic corticosteroid therapy.
Methods: We randomly assigned patients with sudden deafness refractory to systemic corticosteroids to receive either gelatin hydrogels impregnated with IGF-1 in the middle ear (62 patients) or four intratympanic injections with dexamethasone (Dex; 58 patients). The primary outcome was the proportion of patients showing hearing improvement (10 decibels or greater in pure-tone average hearing thresholds) 8 weeks after treatment. The secondary outcomes included the change in pure-tone average hearing thresholds over time and the incidence of adverse events.
Results: In the IGF-1 group, 66.7% (95% confidence interval [CI], 52.9-78.6%) of the patients showed hearing improvement compared to 53.6% (95% CI, 39.7-67.0%) of the patients in the Dex group (P = 0.109). The difference in changes in pure-tone average hearing thresholds over time between the two treatments was statistically significant (P = 0.003). No serious adverse events were observed in either treatment group. Tympanic membrane perforation did not persist in any patient in the IGF-1 group, but did persist in 15.5% (95% CI, 7.3-27.4%) of the patients in the Dex group (P = 0.001).
Conclusions: The positive effect of topical IGF-1 application on hearing levels and its favorable safety profile suggest utility for topical IGF-1 therapy in patients with sudden deafness.

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Impact of cytochrome P450 3A5 polymorphism in graft livers on the frequency of acute cellular rejection in living-donor liver transplantation

Miwa Uesugi, Mio Kikuchi, Haruka Shinke, Tomohiro Omura, Atsushi Yonezawa, Kazuo Matsubara, Yasuhiro Fujimoto, Shinya Okamoto, Toshimi Kaido, Shinji Uemoto, Satohiro Masuda

PHARMACOGENETICS AND GENOMICS (LIPPINCOTT WILLIAMS & WILKINS)  24 ( 7 ) 356 - 366 2014.07

Research paper (scientific journal), Joint Work, Accepted,  ISSN  17446872

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ObjectiveWe investigated whether the cytochrome P450 3A5*3 (CYP3A5*3) genotype affects tacrolimus pharmacokinetics and the risk of acute cellular rejection in living-donor liver transplant patients in Japan.Materials and methodsBetween July 2004 and June 2011, we enrolled 410 living-donor liver transplant patients receiving tacrolimus. Biopsy specimens of intestinal mucosa and graft liver at surgery were obtained to examine the mRNA expression of CYP3A subfamilies as well as the genotyping of CYP3A5*3 polymorphism.ResultsThe CYP3A5 genotype in the native intestine had no significant effect on the occurrence of acute cellular rejection between postoperative days 14 and 23 in cases with identical or compatible ABO blood types (11.5% for the CYP3A5*1 allele vs. 7.4% for CYP3A5*3/*3; P=0.2643), although the concentration/dose ratio of tacrolimus was significantly higher in patients with the intestinal CYP3A5*3/*3 genotype than in those with the CYP3A5*1 allele for 5 post-transplant weeks. However, patients who received a graft liver with the CYP3A5*1 allele showed a higher rate of acute cellular rejection than those who received a graft liver with the CYP3A5*3/*3 genotype (14.5 vs. 5.7%; P=0.0134). The relative risk for acute cellular rejection associated with the CYP3A5*1 liver allele was 2.629 (P=0.018, Cox regression model). Consequently, graft liver CYP3A5*1 genotype might increase the risk for acute cellular rejection after living-donor liver transplantation, possibly by associating with the local hepatic tacrolimus concentration.ConclusionsThe target level of tacrolimus may be affected by the CYP3A5*3 genotype of the liver, rather than by that of the small intestine, after postoperative day 14.

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Assessment of Four Methodologies (Microparticle Enzyme Immunoassay, Chemiluminescent Enzyme Immunoassay, Affinity Column-Mediated Immunoassay, and Flow Injection Assay-Tandem Mass Spectrometry) for Measuring Tacrolimus Blood Concentration in Japanese Liver Transplant Recipients

S. Hashi, S. Masuda, M. Kikuchi, M. Uesugi, I. Yano, T. Omura, A. Yonezawa, Y. Fujimoto, K. Ogawa, T. Kaido, S. Uemoto, K. Matsubara

TRANSPLANTATION PROCEEDINGS (ELSEVIER SCIENCE INC)  46 ( 3 ) 758 - 760 2014.04

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00411345

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Therapeutic drug monitoring (TDM) and subsequent dosage adjustment for individual patients in the treatment with tacrolimus are required after liver transplantation to prevent rejection and over-immunosuppression, which leads to severe infection and adverse reactions including nephrotoxicity. The purpose of this study was to evaluate the analytical performance among commercially available immunoassay methods, which were microparticle enzyme immunoassay (MEIA), chemiluminescent enzyme immunoassay (CLIA), and affinity column-mediated immunoassay (ACMIA), compared with an assay using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, the flow injection assay (FIA-MS/MS) was also evaluated to determine whether it could be available as a new method of analysis in tacrolimus therapy. The blood tacrolimus concentrations in samples from liver transplant recipients (n = 102) were measured using MEIA, CLIA, ACMIA, and LC-MS/MS. Additional blood samples from liver transplant recipients (n = 54) were analyzed using both FIA-MS/MS and LC-MS/MS. Because the assay performance and characteristics of MEIA, CLIA, ACMIA, and FIA-MS/MS are relatively different, the measured data should be carefully considered depending on the methodology.

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Successful Telaprevir Treatment in Combination of Cyclosporine against Recurrence of Hepatitis C in the Japanese Liver Transplant Patients

Mio Kikuchi, Yuki Okuda, Yoshihide Ueda, Yuki Nishioka, Miwa Uesugi, Emina Hashimoto, Tamotsu Takahashi, Tomoki Kawai, Sachiyo Hashi, Haruka Shinke, Tomohiro Omura, Atsushi Yonezawa, Takashi Ito, Yasuhiro Fujimoto, Toshimi Kaido, Tsutomu Chiba, Shinji Uemoto, Kazuo Matsubara, Satohiro Masuda

BIOLOGICAL & PHARMACEUTICAL BULLETIN (PHARMACEUTICAL SOC JAPAN)  37 ( 3 ) 417 - 423 2014.03

Research paper (scientific journal), Joint Work, Accepted,  ISSN  09186158

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Telaprevir (TVR) is a protease inhibitor used in combination with pegylated interferon alfa-2b and ribavirin for hepatitis C, and TVR strongly inhibits CYP3A4 and CYP3A5. We reported successful TVR treatment of liver transplant patients with recurrence of hepatitis C during receiving immunosuppressive therapy. Before initiation of triple therapy, all patients switched from tacrolimus to cyclosporine, which has a lower inhibitory effect on CYP3A4 and CYP3A5 than tacrolimus. To avoid graft failure, we measured the cyclosporine blood concentrations at 0, 2, and 6 h after administration to maintain the target level (150-200 ng/mL) within 1 week after initiation of TVR and adjusted the dose of cyclosporine. The dose of cyclosporine was decreased 0.24-0.40 fold in all patients after initiation of TVR treatment. In 3 patients, the dose of TVR was decreased two-thirds of starting dose because of adverse effects, including anorexia and skin rash. However, the HCV RNA level rapidly decreased to undetectable levels within 1 month. Furthermore, all patients completed the TVR therapy in 12 weeks and did not experience liver graft rejection. In addition, we found the rapid elimination of inhibitory effect of TVR on the disposition of cyclospirne in the all four cases and therefore, rapid increase in the dosage of cyclosporine would be required immediately after the end of TVR administration. These results suggest that frequent measurement of cyclosporine levels was important for successful TVR triple therapy and prevention of rejection.

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Functional involvement of RFVT3/SLC52A3 in intestinal riboflavin absorption

Hiroki Yoshimatsu, Atsushi Yonezawa, Yoshiaki Yao, Kumiko Sugano, Shunsaku Nakagawa, Tomohiro Omura, Kazuo Matsubara

AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY (AMER PHYSIOLOGICAL SOC)  306 ( 2 ) G102 - G110 2014.01

Research paper (scientific journal), Joint Work, Corresponding author, Accepted,  ISSN  01931857

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Riboflavin, also known as vitamin B2, is transported across the biological membrane into various organs by transport systems. Riboflavin transporter RFVT3 is expressed in the small intestine and has been suggested to localize in the apical membranes of the intestinal epithelial cells. In this study, we investigated the functional involvement of RFVT3 in riboflavin absorption using intestinal epithelial T84 cells and mouse small intestine. T84 cells expressed RFVT3 and conserved unidirectional riboflavin transport corresponding to intestinal absorption. Apical [H-3] riboflavin uptake was pH-dependent in T84 cells. This uptake was not affected by Na+ depletion at apical pH 6.0, although it was significantly decreased at apical pH 7.4. The [H-3] riboflavin uptake from the apical side of T84 cells was prominently inhibited by the RFVT3 selective inhibitor methylene blue and significantly decreased by transfection of RFVT3-small-interfering RNA. In the gastrointestinal tract, RFVT3 was expressed in the jejunum and ileum. Mouse jejunal and ileal permeabilities of [H-3] riboflavin were measured by the in situ closed-loop method and were significantly reduced by methylene blue. These results strongly suggest that RFVT3 would functionally be involved in riboflavin absorption in the apical membranes of intestinal epithelial cells.

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Investigation of Endogenous Compounds for Assessing the Drug Interactions in the Urinary Excretion Involving Multidrug and Toxin Extrusion Proteins

Koji Kato, Haruyuki Mori, Tomoko Kito, Miyu Yokochi, Sumito Ito, Katsuhisa Inoue, Atsushi Yonezawa, Toshiya Katsura, Yuji Kumagai, Hiroaki Yuasa, Yoshinori Moriyama, Ken-ichi Inui, Hiroyuki Kusuhara, Yuichi Sugiyama

PHARMACEUTICAL RESEARCH (SPRINGER/PLENUM PUBLISHERS)  31 ( 1 ) 136 - 147 2014.01

Research paper (scientific journal), Joint Work, Accepted,  ISSN  07248741

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Purpose Multidrug and toxin extrusion proteins (MATEs) are multispecific organic cation transporters mediating the efflux of various cationic drugs into the urine. The present study aimed at identifying endogenous compounds in human plasma and urine specimens as biomarkers to evaluate drug interactions involving MATEs in the kidney without administration of their exogenous probe drugs.
Methods An untargeted metabolomic analysis was performed using urine and plasma samples from healthy volunteers and mice treated with or without the potent MATE inhibitor, pyrimethamine. Plasma and urinary concentrations of candidate markers were measured using liquid chromatography-mass spectrometry. Transport activities were determined in MATE- or OCT2-expressing HEK293 cells. The deuterium-labeled compounds of candidates were administered to mice for pharmacokinetics study.
Results Urinary excretion of eleven compounds including thiamine and carnitine was significantly lower in the pyrimethamine-treatment group in humans and mice, whereas no endogenous compound was noticeably accumulated in the plasma. The renal clearance of thiamine and carnitine was decreased by 70%-84% and 90%-94% (p < 0.05), respectively, in human. The specific uptake of thiamine was observed in MATE1-, MATE2-K- or OCT2-expressing HEK293 cells with K-m of 3.5 +/- 1.0, 3.9 +/- 0.8 and 59.9 +/- 6.7 mu M, respectively. The renal clearance of carnitine-d (3) was decreased by 62% in mice treated with pyrimethamine.
Conclusions Our findings indicate that MATEs account for the efflux of thiamine and perhaps carnitine as well as drugs into the urine. The urinary excretion of thiamine is useful to detect drug interaction involving MATEs in the kidney.

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Treatable childhood neuronopathy caused by mutations in riboflavin transporter RFVT2

A. Reghan Foley, Manoj P. Menezes, Amelie Pandraud, Michael A. Gonzalez, Ahmad Al-Odaib, Alexander J. Abrams, Kumiko Sugano, Atsushi Yonezawa, Adnan Y. Manzur, Joshua Burns, Imelda Hughes, B. Gary McCullagh, Heinz Jungbluth, Ming J. Lim, Jean-Pierre Lin, Andre Megarbane, J. Andoni Urtizberea, Ayaz H. Shah, Jayne Antony, Richard Webster, Alexander Broomfield, Joanne Ng, Ann A. Mathew, James J. O'Byrne, Eva Forman, Mariacristina Scoto, Manish Prasad, Katherine O'Brien, Simon Olpin, Marcus Oppenheim, Iain Hargreaves, John M. Land, Min X. Wang, Kevin Carpenter, Rita Horvath, Volker Straub, Monkol Lek, Wendy Gold, Michael O. Farrell, Sebastian Brandner, Rahul Phadke, Kazuo Matsubara, Michael L. McGarvey, Steven S. Scherer, Peter S. Baxter, Mary D. King, Peter Clayton, Shamima Rahman, Mary M. Reilly, Robert A. Ouvrier, John Christodoulou, Stephan Zuechner, Francesco Muntoni, Henry Houlden

BRAIN (OXFORD UNIV PRESS)  137 ( 1 ) 44 - 56 2014.01

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00068950

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Childhood onset motor neuron diseases or neuronopathies are a clinically heterogeneous group of disorders. A particularly severe subgroup first described in 1894, and subsequently called Brown-Vialetto-Van Laere syndrome, is characterized by progressive pontobulbar palsy, sensorineural hearing loss and respiratory insufficiency. There has been no treatment for this progressive neurodegenerative disorder, which leads to respiratory failure and usually death during childhood. We recently reported the identification of SLC52A2, encoding riboflavin transporter RFVT2, as a new causative gene for Brown-Vialetto-Van Laere syndrome. We used both exome and Sanger sequencing to identify SLC52A2 mutations in patients presenting with cranial neuropathies and sensorimotor neuropathy with or without respiratory insufficiency. We undertook clinical, neurophysiological and biochemical characterization of patients with mutations in SLC52A2, functionally analysed the most prevalent mutations and initiated a regimen of high-dose oral riboflavin. We identified 18 patients from 13 families with compound heterozygous or homozygous mutations in SLC52A2. Affected individuals share a core phenotype of rapidly progressive axonal sensorimotor neuropathy (manifesting with sensory ataxia, severe weakness of the upper limbs and axial muscles with distinctly preserved strength of the lower limbs), hearing loss, optic atrophy and respiratory insufficiency. We demonstrate that SLC52A2 mutations cause reduced riboflavin uptake and reduced riboflavin transporter protein expression, and we report the response to high-dose oral riboflavin therapy in patients with SLC52A2 mutations, including significant and sustained clinical and biochemical improvements in two patients and preliminary clinical response data in 13 patients with associated biochemical improvements in 10 patients. The clinical and biochemical responses of this SLC52A2-specific cohort suggest that riboflavin supplementation can ameliorate the progression of this neurodegenerative condition, particularly when initiated soon after the onset of symptoms.

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Safety and Efficacy of Sustained Release of Basic Fibroblast growth Factor Using Biodegradable Gelatin Hydrogel in Patients With Critical Limb Ischemia

Motoyuki Kumagai, Akira Marui, Yasuhiko Tabata, Takahide Takeda, Masaya Yamamoto, Atsushi Yonezawa, Shiro Tanaka, Shigeki Yanagi, Toshiko Ito-Ihara, Takafumi Ikeda, Toshinori Murayama, Satoshi Teramukai, Toshiya Katsura, Kazuo Matsubara, Koji Kawakami, Masayuki Yokode, Akira Shimizu, Ryuzo Sakata

CIRCULATION (LIPPINCOTT WILLIAMS & WILKINS)  128 ( 22 ) 16154.0 2013.11

Research paper (scientific journal), Joint Work, Accepted,  ISSN  0009-7322

Impact of P-glycoprotein and breast cancer resistance protein on the brain distribution of antiepileptic drugs in knockout mouse models

Haruka Nakanishi, Atsushi Yonezawa, Kazuo Matsubara, Ikuko Yano

European Journal of Pharmacology (1-3)  710 ( 1-3 ) 20 - 28 2013.06

Research paper (scientific journal), Joint Work, Accepted,  ISSN  00142999

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Refractory epilepsy is reportedly associated with an overexpression of ATP-binding cassette transporters such as P-glycoprotein (Pgp) and breast cancer resistance protein (Bcrp). In this study, we examined the contribution of Pgp and Bcrp to the brain distribution of 12 antiepileptic drugs (AEDs) in Mdr1a/1b(-/-) and Mdr1a/1b(-/-)/Bcrp(-/-) mice within a therapeutic concentration range. The blood concentrations were sequentially determined, and the brain concentrations were measured at 60 min after intravenous administration. The plasma concentration profiles for each AED in the Mdr1a/1b(-/-) mice were equivalent to those in the wild-type mice. In contrast, the plasma concentration profiles of phenytoin, lamotrigine, topiramate, tiagabine, and levetiracetam in the Mdr1a/1b(-/-)/Bcrp(-/-) mice were significantly lower than the corresponding ones in the wild-type mice. The brain-to-plasma concentration ratio (Kpbrain) values of phenytoin, topiramate, and tiagabine in the Mdr1a/1b(-/-) mice were significantly higher than the corresponding ones in the wild-type mice. In contrast, the Kp brain values of phenobarbital, clobazam, zonisamide, gabapentin, tiagabine, and levetiracetam in the Mdr1a/1b(-/-)/Bcrp(-/-) mice were significantly higher than the corresponding ones in Mdr1a/1b(-/-) mice. The Kpbrain values of the 12 AEDs in the Mdr1a/1b(-/-)/Bcrp(-/-) mice, but not wild-type mice, significantly correlated with the corresponding molecular weight values. These findings suggest that both Pgp and Bcrp restrict brain access for several AEDs. Taken together, information on the contribution of each transporter may be useful in the development of strategic treatments of refractory epilepsy. © 2013 Elsevier B.V.

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Novel riboflavin transporter family RFVT/SLC52: Identification, nomenclature, functional characterization and genetic diseases of RFVT/SLC52

Atsushi Yonezawa, Ken-ichi Inui

MOLECULAR ASPECTS OF MEDICINE (ELSEVIER SCIENCE BV)  34 ( 2-3 ) 693 - 701 2013.04

Research paper (scientific journal), Joint Work, Lead author, Corresponding author, Accepted,  ISSN  00982997

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Riboflavin, a water-soluble vitamin also known as vitamin B2, is essential for normal cellular functions. Riboflavin transporters play important roles in its homeostasis. Recently, three novel riboflavin transporters were identified, and designated as RFT1, RFT2 and RFT3. Because the RFTs did not show similarity to other SLC transporters, and RFT1 and RFT3 are similar in sequence and function, they were assigned into a new SLC family, SLC52. Subsequently, RFT1/GPR172B, RFT3/GPR172A and RFT2/C20orf54 were renamed as RFVT1/SLC52A1, RFVT2/SLC52A2 and RFVT3/SLC52A3, respectively. In this review, we summarize recent findings on the cloning, nomenclature, functional characterization and genetic diseases of RFVT1/SLC52A1, RFVT2/SLC52A2 and RFVT3/SLC52A3. (C) 2012 Elsevier Ltd. All rights reserved.

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