2024年10月, 第18回次世代を担う若手医療薬科学シンポジウム 優秀発表賞　　　　　　　　　　　　　　　
アルデヒドオキシダーゼの薬物動態学的重要性および個体間差の解明
上田 一奈太、鳴海 克哉、岡本 敬介、古堅 彩子、小林 正紀

2024年08月, 第37回 北海道薬物作用談話会 若手研究者優秀発表賞（口頭発表）　　　　　　　　　　　　　　　
糖尿病治療薬がシスプラチン起因性腎障害に及ぼす影響の評価
坂田 浩太郎、岡本 敬介、齋藤 佳敬、古堅 彩子、鳴海 克哉、小林 正紀

2024年07月, 第40回日本TDM学会・学術大会 学生優秀演題賞 （口頭発表）　　　　　　　　　　　　　　　
オレキシン受容体拮抗薬のUPLC/MS/MS定量法構築とヒト乳汁移行性評価への応用
石川 陽菜、古堅 彩子、西村 あや子、馬詰 武、青栁 亮一、石川 修平、鳴海 克哉、岡本 敬介、武隈 洋、菅原 満、小林 正紀

2024年07月, 日本薬学会北海道支部第151回例会 学生優秀発表賞 （口頭発表）　　　　　　　　　　　　　　　
ヒト胎盤幹細胞におけるトランスポーター発現・機能評価
澤田 理子、古堅 彩子、植田 彩文、西村 あや子、馬詰 武、鳴海 克哉、岡本 敬介、小林 正紀

2023年07月, 医療薬学ﾌｫｰﾗﾑ2023／第31回ｸﾘﾆｶﾙﾌｧｰﾏｼｰｼﾝﾎﾟｼﾞｳﾑ優秀ポスター賞　　　　　　　　　　　　　　　
メトトレキサート誘発性肝障害におけるアルデヒドオキシダーゼの寄与 ―基礎および薬剤疫学的アプローチによる検証―
鳴海 克哉、森山 綾子、上田 一奈太、淺野 秀峰、古堅 彩子、小林 正紀

2023年03月, 日本薬学会 第143年会 優秀発表賞　　　　　　　　　　　　　　　
アザチオプリン誘発性血液障害および肝障害発現における Aldehyde oxidase 阻害薬剤の影響
上田 一奈太、鳴海 克哉、古堅 彩子、小林正紀

2022年08月, 第35回北海道薬物作用談話会 若手研究者優秀発表賞　　　　　　　　　　　　　　　
メトトレキサート誘発性肝障害におけるaldehyde oxidase 1の関与
森山 綾子;鳴海 克哉;上田 一奈太;古堅 彩子;小林 正紀

2022年06月, 日本医療薬学会 第5回 フレッシャーズ・カンファランス 優秀演題　　　　　　　　　　　　　　　
AOX1遺伝子のT755I多型は二量体形成および代謝活性を低下させる
上田 一奈太、鳴海 克哉、古堅 彩子、小林 正紀

2022年05月, 日本薬学会北海道支部第149回例会学生優秀発表賞 （口頭発表部門）　　　　　　　　　　　　　　　
肝がん由来細胞において乳酸輸送に寄与する輸送担体の同定に関する研究
向井 悠斗、山口 敦史、佐久間 智也、古堅 彩子、鳴海 克哉、小林正紀

2021年03月, 日本薬学会 第141年会 優秀発表賞　　　　　　　　　　　　　　　
シスプラチンの副作用と耐性化に着目したNSAIDsの効果の検証
岡本 敬介、齋藤 佳敬、上田 一奈太、古堅 彩子、鳴海 克哉、小林 正紀

2020年11月, 第14回次世代を担う若手医療薬科学シンポジウム 優秀発表賞　　　　　　　　　　　　　　　
クロザピン誘発性流涎症のリスク因子の同定と発現メカニズムの解明
石川 修平, 小林 正紀, 橋本 直樹, 久住 一郎

2017年01月, 北海道大学, 教育総長賞 (奨励賞)　　　　　　　　　　　　　　　
小林 正紀

2015年11月, 第２５回日本医療薬学会奨励賞　　　　　　　　　　　　　　　
MCTの役割に着目した疾患と副作用に関する研究
小林 正紀

2014年05月, 日本薬学会北海道支部 奨励賞　　　　　　　　　　　　　　　
脂質異常症治療薬の副作用・薬物相互作用におけるモノカルボン酸トランスポータの役割
小林 正紀

Validated UPLC-MS/MS method for quantification of melatonin receptor agonists and dual orexin receptor antagonists in human plasma and breast milk: Application to quantify suvorexant and lemborexant in clinical samples
Hina Ishikawa, Ayako Furugen, Ayako Nishimura, Takeshi Umazume, Shuhei Ishikawa, Ryoichi Aoyagi, Katsuya Narumi, Keisuke Okamoto, Yoh Takekuma, Mitsuru Sugawara, Masaki Kobayashi
Journal of Pharmaceutical and Biomedical Analysis, 251, 116432, 116432, Elsevier BV, 2024年12月, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）

Effects of famotidine use during pregnancy: an observational cohort study.
Ayako Nishimura, Ayako Furugen, Masaki Kobayashi, Yoh Takekuma, Naho Yakuwa, Mikako Goto, Masahiro Hayashi, Atsuko Murashima, Mitsuru Sugawara
Journal of pharmaceutical health care and sciences, 10, 1, 70, 70, 2024年11月08日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Famotidine, a histamine2-receptor antagonist (H2Ras), is widely used to treat and prevent gastrointestinal symptoms during pregnancy. Although several studies have reported the use of H2Ras during pregnancy, limited data on famotidine were included in these reports. Therefore, we analyzed pregnancy outcome data to evaluate the effects of famotidine use during pregnancy on the fetus. METHODS: Pregnancy outcome data were used for females enrolled in two Japanese facilities that provided counseling on drug use during pregnancy between April 1988 and December 2017. For the primary endpoint, the incidence of congenital malformations was calculated from the data of live birth to pregnant women who took famotidine (n = 330) or drugs considered to exert no teratogenic risk (control, n = 1,407) during the first trimester of pregnancy. Considering secondary endpoints, the incidence of obstetric outcomes, including preterm delivery, was calculated from data on the use of famotidine (n = 347) and controls (n = 1,476) during the entire pregnancy. The crude odds ratios (cORs) for the incidence of congenital malformations were calculated using univariate logistic regression analysis, with the control group used as the reference. Adjusted ORs (aORs) were calculated using multivariate logistic regression analysis adjusted for various other factors. RESULTS: The incidences of congenital malformations in the famotidine and control groups were 3.9% and 2.8%, respectively. There was no significant difference between the famotidine and control groups (cOR: 1.40 [95% CI:0.68-2.71], aOR: 1.06 [95% CI:0.51-2.16]). Conversely, the preterm delivery rates were 8.1% and 3.8% in the famotidine and control groups, respectively, indicating a significant difference (cOR: 2.00 [95% CI:1.20-3.27]). However, the multivariate analysis eliminated famotidine use as a confounding factor. CONCLUSIONS: This observational cohort study revealed that exposure to famotidine during the first trimester of pregnancy was not associated with an increased risk of congenital malformations in infants. Although a higher rate of preterm delivery was detected in famotidine users when compared with controls, this could be attributed to confounding factors, such as complications.

Reply to Accurate Risk Prediction Model for Surgical Site Infection After Lower Third Molar Surgery.
Akira Yamagami, Katsuya Narumi, Yoshitaka Saito, Ayako Furugen, Shungo Imai, Yoshimasa Kitagawa, Yoichi Ohiro, Ryo Takagi, Yoh Takekuma, Mitsuru Sugawara, Masaki Kobayashi
Oral diseases, 2024年11月03日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語

Relationship between magnesium dosage and the preventive effect on cisplatin-induced nephrotoxicity: meta-analysis and meta-regression analysis
Keisuke Okamoto, Yoshitaka Saito, Atsushi Yamaguchi, Katsuya Narumi, Masaki Kobayashi
International Journal of Clinical Oncology, Springer Science and Business Media LLC, 2024年09月24日, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）

Salmon Milt Extract Suppresses Glucose Uptake by Downregulating SGLT1 and GLUT2 Expression in Caco-2 Cells
Taichi Sato, Katsuya Narumi, Rin Taguchi, Komei Ishihara, Hiroshi Satoh, Takao Mori, Keisuke Okamoto, Ayako Furugen, Masaki Kobayashi
Biological and Pharmaceutical Bulletin, 47, 9, 1477, 1483, Pharmaceutical Society of Japan, 2024年09月04日, ［査読有り］, ［最終著者, 責任著者］, ［国内誌］
英語, 研究論文（学術雑誌）, Salmon milt extract (SME) is rich in nucleotides, especially deoxyribonucleoside monophosphates (dNMPs), which has the potential to exert anti-obesity effects. Sodium-dependent glucose transporter 1 (SGLT1) and glucose transporter 2 (GLUT2) are responsible for absorbing sugar from the small intestine. The purpose of this study was to examine the effects of SME on the functions of SGLT1 and GLUT2 and elucidate the mechanisms underlying the inhibition of glucose absorption by SME. We investigated the effect of SME on the expression and function of intestinal glucose transporters, using differentiated Caco-2 cells. SME treatment decreased the expression SGLT1 and GLUT2 mRNA and protein in Caco-2 cells. [14C]-Labelled methyl-α-D-glucopyranoside and [3H]-labelled 2-deoxy-D-glucose (DG) uptake into Caco-2 cells was significantly reduced by SME treatment. Similarly, the dNMP mixture containing the four mononucleotides 2'-deoxyadenosine 5'-monophosphate (dAMP), 2'-deoxyguanosine 5'-monophosphate (dGMP), 2'-deoxycytidine 5'-monophosphate (dCMP), and 2'-deoxythymidine 5'-monophosphate (dTMP) decreased SGLT1 and GLUT2 expression. dNMP mixture-induced reduction in the mRNA expression of these transporters was suppressed when exposed to the mixture without dTMP. Furthermore, dNMP mixture-induced alterations in the expression of hepatocyte nuclear factor (HNF)-1α and HNF1β, which have been characterized as modulators of both transporters also showed a similar trend. dTMP treatment alone decreased GLUT2 expression, resulting in reduced [3H] DG uptake by Caco-2 cells. SME decreased the expression of HNF1α, HNF1β, and its targets SGLT1 and GLUT2, resulting in reduced glucose uptake by Caco-2 cells. In addition, our results revealed that dTMP plays an important role in suppressing the expression of intestinal glucose transporters.

Alteration in folate carrier expression via histone deacetylase inhibition in BeWo human placental choriocarcinoma cells.
Yuki Miyazawa, Ayako Furugen, Ryoichi Aoyagi, Haruna Kosugi, Ayako Nishimura, Takeshi Umazume, Katsuya Narumi, Masaki Kobayashi
Toxicology in vitro : an international journal published in association with BIBRA, 105934, 105934, 2024年09月03日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Folates are essential nutrients for fetal development during pregnancy. Valproic acid (VPA), an inhibitor of histone deacetylases (HDACs), alters the expression of folate carriers in placental cells; however, the underlying mechanisms remain unclear. Here, we aimed to determine the profiles of folate carriers (folate receptor alpha [FOLR1], solute carrier [SLC]-19A1, and SLC46A1) after inhibition of HDACs, especially class I and IIa HDACs, using different inhibitors and gene knockdown tests. Quantitative polymerase chain reaction revealed that BeWo cells (a trophoblast model) expressed HDACs and folate carriers, similar to human placental villi. FOLR1 expression was upregulated by VPA, apicidin, and trichostatin A, but downregulated by MS-275 after 24 h treatment. VPA and apicidin upregulated the expression of SLC46A1. These inhibitors downregulated SLC19A1 expression. TMP269 (a class IIa inhibitor) did not affect folate carrier levels. HDAC1/2 knockdown upregulated FOLR1 and SLC46A1 levels, whereas HDAC1/3 knockdown downregulated FOLR1 levels. Our findings suggest that the pharmacological inhibition of class I HDACs alters the expression of folate carriers in BeWo cells. By contrast, HDAC inhibitors exert different regulatory effects on folate carriers. Moreover, HDAC1/2 inhibition may be a potential mechanism involved in altering FOLR1 and SLC46A1 levels.

Contribution of aldehyde oxidase to methotrexate-induced hepatotoxicity: In Vitro and pharmacoepidemiological approaches.
Ayako Moriyama, Hinata Ueda, Katsuya Narumi, Shuho Asano, Ayako Furugen, Yoshitaka Saito, Masaki Kobayashi
Expert opinion on drug metabolism & toxicology, 2024年05月06日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Methotrexate (MTX) is partially metabolized by aldehyde oxidase (AOX) in the liver and its clinical impact remains unclear. In this study, we aimed to demonstrate how AOX contributes to MTX-induced hepatotoxicity in vitro and clarify the relationship between concomitant AOX inhibitor use and MTX-associated liver injury development using the U.S. Food and Drug Administration Adverse Event Reporting System (FAERS). METHODS: We assessed intracellular MTX accumulation and cytotoxicity using HepG2 cells. We used the FAERS database to detect reporting odds ratio (ROR)-based MTX-related hepatotoxicity event signals. RESULTS: AOX inhibition by AOX inhibitor raloxifene and siRNA increased the MTX accumulation in HepG2 cells and enhanced the MTX-induced cell viability reduction. In the FAERS analysis, the ROR for MTX-related hepatotoxicity increased with non-overlap of 95% confidence interval when co-administered with drugs with higher Imax, u (maximum unbound plasma concentration)/IC50 (half-maximal inhibitory concentration for inhibition of AOX) calculated based on reported pharmacokinetic data. CONCLUSION: AOX inhibition contributed to MTX accumulation in the liver, resulting in increased hepatotoxicity. Our study raises concerns regarding MTX-related hepatotoxicity when co-administered with drugs that possibly inhibit AOX activity at clinical concentrations.

Monocarboxylate Transporters 1 and 2 Are Responsible for L-Lactate Uptake in Differentiated Human Neuroblastoma SH-SY5Y Cells
Tomoya Sakuma, Yuto Mukai, Atsushi Yamaguchi, Yudai Suganuma, Keisuke Okamoto, Ayako Furugen, Katsuya Narumi, Shuhei Ishikawa, Yoshitaka Saito, Masaki Kobayashi
Biological and Pharmaceutical Bulletin, 47, 4, 764, 770, Pharmaceutical Society of Japan, 2024年04月04日, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）

Evaluation of Prediabetes in Cisplatin-induced Nephrotoxicity in the Short Hydration Method: A Subgroup Analysis
YOSHITAKA SAITO, TATSUHIKO SAKAMOTO, MASAKI KOBAYASHI, YOH TAKEKUMA, ISSEI HIGUCHI, KEISUKE OKAMOTO, JUN SAKAKIBARA-KONISHI, YASUSHI SHIMIZU, ICHIRO KINOSHITA, MITSURU SUGAWARA
In Vivo, 38, 2, 800, 806, Anticancer Research USA Inc., 2024年02月28日, ［査読有り］
研究論文（学術雑誌）

Breast milk concentrations of acetaminophen and diclofenac - unexpectedly high mammary transfer of the general-purpose drug acetaminophen.
Ryo Tamaki, Kiwamu Noshiro, Ayako Furugen, Ayako Nishimura, Hiroshi Asano, Hidemichi Watari, Masaki Kobayashi, Takeshi Umazume
BMC pregnancy and childbirth, 24, 1, 90, 90, 2024年01月29日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Breastfeeding is considered to be the most effective way of ensuring the health and survival of newborns. However, mammary transfer of drugs administered to mothers to breastfeeding infants remains a pressing concern. Acetaminophen and diclofenac sodium are widely prescribed analgesics for postpartum pain relief, but there have been few recent reports on the mammary transfer of these drugs, despite advances in analytic techniques. METHODS: We conducted a study on 20 postpartum mothers from August 2019-March 2020. Blood and milk samples from participants were analyzed using liquid chromatography-electrospray ionization tandem mass spectrometry within 24 hours after oral administration of acetaminophen and diclofenac sodium. The area under the concentration-time curve (AUC) was calculated from the concentration curve obtained by a naive pooled-data approach. RESULTS: For acetaminophen, AUC was 36,053 ng/mL.h and 37,768 ng/mL.h in plasma and breast milk, respectively, with a milk-to-plasma drug concentration ratio of 1.048. For diclofenac, the AUC was 0.227 ng/mL.h and 0.021 ng/mL.h, in plasma and breast milk, respectively, with a milk-to-plasma drug concentration ratio of 0.093. CONCLUSIONS: While diclofenac sodium showed low mammary transfer, acetaminophen showed a relatively high milk-to-plasma drug concentration ratio. Given recent studies suggesting potential connections between acetaminophen use during pregnancy and risks to developmental prognosis in children, we believe that adequate information regarding the fact that acetaminophen is easily transferred to breast milk should be provided to mothers.

Association Between Multisystem Immune-related Adverse Events and Progression-free Survivals in PD-1/PD-L1 Inhibitor Monotherapy.
Atsushi Yamaguchi, Yoshitaka Saito, Keisuke Okamoto, Ayako Furugen, Katsuya Narumi, Yoh Takekuma, Jun Sakakibara-Konishi, Yasushi Shimizu, Ichiro Kinoshita, Mitsuru Sugawara, Masaki Kobayashi
In vivo (Athens, Greece), 38, 6, 2886, 2896, 2024年, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND/AIM: Immune-related adverse events (irAEs) occur in various organs, and sometimes multiply following treatment with immune checkpoint inhibitors (ICIs). This study aimed to determine the association between the number of irAEs and clinical outcomes. PATIENTS AND METHODS: This was a retrospective study that included patients with lung cancer, melanoma, and head and neck cancer who were treated with anti-programmed cell death (ligand) 1 (PD-1/PD-L1) monotherapy. We evaluated the association between the number of irAEs and progression-free survival (PFS) in the simple Cox regression analysis. To eliminate the immortal-time bias, an additional landmark analysis was performed. RESULTS: In total, 92, 69, and 37 patients were allocated to the no, single, and multisystem irAEs groups, respectively. The multisystem irAEs were associated with better PFS compared to the no irAE group. In contrast, at the 12-week landmark, multisystem irAEs were associated with poor PFS compared to the no irAEs group. Furthermore, the rate of treatment suspension owing to irAEs in the multisystem irAEs group (62.5%) was higher than that in the single irAE group (17.3%) at the 12-week landmark. CONCLUSION: The incidence of multisystem irAEs was associated with improved clinical outcomes in patients with lung cancer, melanoma, and head and neck cancer treated with PD-1/PD-L1 inhibitor monotherapy. However, these results may be influenced by a potential immortal-time bias. When accounting for this bias, the early development of multisystem irAEs within 12 weeks was linked to treatment suspension and poorer clinical outcomes.

Regulation of Chloride Channels by Epidermal Growth Factor Receptor-Tyrosine Kinase Inhibitor-Induced α-Defensin 5.
Ippei Uemura, Natsuko Takahashi-Suzuki, Fumiya Kita, Masaki Kobayashi, Takehiro Yamada, Ken Iseki, Takashi Satoh
Biological & pharmaceutical bulletin, 47, 1, 159, 165, 2024年, ［査読有り］, ［国内誌］
英語, 研究論文（学術雑誌）, Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are used to treat non-small cell lung cancer with EGFR mutations. However, first-generation erlotinib and second-generation afatinib often cause diarrhea, which may develop because of the association between EGFR-TKIs and the chloride channel or abnormalities in the intestinal microbiota due to disruption of the intestinal immune system. As reports on the effects of EGFR-TKIs on intestinal immunity are lacking, we aimed to determine whether the intestinal immune system is involved in the molecular effects of EGFR-TKIs on chloride channels using Caco-2 cells. Initially, we evaluated the association of chloride channels with α-defensin 5 (DEFA5), a marker of intestinal immunity. Erlotinib and afatinib significantly increased the extracellularly secreted DEFA5 level and autophagy-related 16-like 1 and X-box binding protein 1 transcript levels, indicative of enhanced granule exocytosis. Conversely, intracellular DEFA5 and Toll-like receptor 4 protein expression and tumor necrosis factor-α transcript levels decreased significantly, suggesting that Toll-like receptor 4 suppression repressed DEFA5 production. Furthermore, among the chloride channels, DEFA5 was found to significantly increase the transcript levels of cystic fibrosis transmembrane conductance regulators. These results indicate that DEFA5 plays a significant role in the mechanism of chloride channel-mediated diarrhea induced by EGFR-TKIs. Therefore, we successfully elucidated the potential host action of DEFA5 in cancer therapy for the first time.

Comparative study on the occurrence of adverse effects in the concomitant use of azathioprine and aldehyde oxidase inhibitors.
Hinata Ueda, Katsuya Narumi, Shuho Asano, Yoshitaka Saito, Ayako Furugen, Masaki Kobayashi
Expert opinion on drug safety, 2023年12月14日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, OBJECTIVES: Aldehyde oxidase (AO) is a molybdenum-containing redox enzyme similar to xanthine oxidase that is involved in the thiopurine metabolism. This study investigated the effects of drug-drug interactions (DDIs) between azathioprine (AZA) and AO inhibitors on hematologic and hepatic disorders using the U.S. Food and Drug Administration Adverse Event Reporting System and the Japanese Adverse Drug Event Report database. METHODS: The presence of DDI was assessed using the interaction signal scores (ISSs) calculated via the reporting odds ratios and 95% confidence intervals. The study used reports of 'azathioprine' as a suspect drug for adverse effects. AO inhibitors were selected based on previous in vitro reports. RESULTS: Some drugs tested positive for ISSs in each database and type of adverse effect (hematologic or hepatic disorder) analysis. Among these drugs, chlorpromazine, clozapine, hydralazine, and quetiapine could inhibit AZA metabolism via AO, given the previously reported clinical blood concentration and inhibitory effects of each drug. CONCLUSION: Concomitant use of AO inhibitors increased the signals for AZA-induced adverse effects. To date, no studies have evaluated the clinical importance of AO as a drug-metabolizing enzyme, and further in vitro and clinical research is needed to clarify the contribution of AO to the pharmacokinetics of thiopurines.

Use of lacosamide for focal epilepsy in a child with kidney failure undergoing peritoneal dialysis.
Yuki Ueda, Ayako Furugen, Masaki Kobayashi, Yasuyuki Sato, Yasuhiro Ueda, Asako Hayashi, Takeru Goto, Shuhei Kimura, Masashi Narugami, Sachiko Nakakubo, Midori Nakajima, Kiyoshi Egawa, Takayuki Okamoto, Atsushi Manabe, Hideaki Shiraishi
Brain & development, 2023年10月30日, ［査読有り］, ［国際誌］
英語, BACKGROUND: Lacosamide (LCM) has become commonly used for focal onset seizures due to its high tolerability and low drug interactions. Unlike patients on hemodialysis (HD), pharmacokinetic data and dosing recommendations for patients undergoing peritoneal dialysis (PD) are scant. CASE REPORT: A 2-year-old girl with end-stage kidney disease undergoing PD suffered prolonged focal onset seizures. The patient had congenital anomalies of the kidney and urinary tract associated with branchio-oto-renal syndrome due to an EYA1 gene mutation. She also had neurological sequelae from post-resuscitation encephalopathy at the age of one month. Antiseizure medication with few drug interactions, less impact on the neurodevelopmental state and possibility of intravenous administration was preferred. LCM met those criteria and was carefully administered. Although the patient had recurrent prolonged seizures during the titration periods, LCM could be continued without any apparent side effects. The blood levels of LCM increased linearly to the optimal level. We confirmed excretion of LCM in the PD fluid. Kidney transplantation was done three months after and her seizures were well controlled. CONCLUSIONS: LCM might be a promising option for patients undergoing PD. Due to the lower removal efficacy in PD compared with in HD, close attention should be paid to possible drug excess.

Risk factor analysis for cisplatin-induced nephrotoxicity with the short hydration method in diabetic patients.
Yoshitaka Saito, Masaki Kobayashi, Shinya Tamaki, Katsuyuki Nakamura, Daisuke Hirate, Kenta Takahashi, Yoh Takekuma, Jun Sakakibara-Konishi, Yasushi Shimizu, Ichiro Kinoshita, Mitsuru Sugawara
Scientific reports, 13, 1, 17126, 17126, 2023年10月10日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The occurrence of cisplatin (CDDP)-induced nephrotoxicity (CIN) has decreased with advancements in supportive care. In contrast, we reported that baseline diabetes mellitus (DM) complications significantly worsen CIN. This study aimed to determine further risk factors associated with CIN development in DM patients. Patients with thoracic cancer requiring DM pharmacotherapy, who received CDDP (≥ 60 mg/m2)-containing regimens using the short hydration method (n = 140), were enrolled in this retrospective multicenter observational study. The primary endpoint of the present study was the elucidation of risk factors (patient factors, DM medication influence, and treatment-related factors) associated with CIN development in patients with DM. Cisplatin-induced nephrotoxicity occurred in 22.1% of patients with DM. The median worst variation of serum creatinine levels and creatinine clearance (worst level - baseline level) was 0.16 mg/dL (range: - 0.12-1.41 mg/dL) and - 15.9 mL/min (- 85.5-24.3 mL/min), respectively. Multivariate logistic regression analyses identified female sex as the singular risk factor for CIN development in the DM population (adjusted odds ratio; 2.87, 95% confidence interval; 1.08-7.67, P = 0.04). Diabetes mellitus medication and treatment-related factors did not affect CIN development. In conclusion, our study revealed that female sex is significantly associated with CIN development in patients with DM and thoracic cancer.

Quantitative analysis of communication changes in online medication counseling -Using the Roter Interaction Analysis System.
Ayako Mori, Izumi Kato, Katsuya Narumi, Yoh Takekuma, Hitoshi Kashiwagi, Yuki Sato, Mitsuru Sugawara, Masaki Kobayashi
Research in social & administrative pharmacy : RSAP, 2023年10月04日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Quantitative analysis and objective evaluation of communication play an important role in medical communication education. In the process of developing an online methodology for medication counseling practice, we felt the necessity of conducting a quantitative evaluation to enhance its effectiveness. OBJECTIVES: This study aimed to quantitatively evaluate the communication in each scenario to comprehensively identify the differences between face-to-face and online communication in medication counseling practicum. In addition, we examined how patient satisfaction changes between face-to-face and online interactions. METHODS: Face-to-face and online role-playing were conducted between simulated patients (SPs) and students acting as pharmacists, and their dialogues were videotaped. The utterances in each recorded dialogue were categorized and analyzed by the Roter interaction analysis system (RIAS). The Japanese version of the Medical Interview Satisfaction Scale (MISS-21J) responses of the SPs were analyzed for the patient satisfaction survey. RESULT: The results of the RIAS analysis revealed that the socio-emotional category appeared significantly more frequently in face-to-face communication, with more utterances that were more attuned to the feelings of the other person and more considerate of his or her feelings. The ratio of the number of utterances between students and SPs suggested that the communication was more interactive. CONCLUSION: Based on the respective communication tendencies may have led to higher satisfaction in face-to-face than in online patient satisfaction surveys, less anxiety about illness and medications, and easier trusting relationships. Since it is difficult to grasp the mood of the other party and to open up to them due to the lack of nonverbal information in online dialogue, it is necessary to be more conscious of conversations that capture the feelings of patients in online medication counseling.

Molecular characteristic analysis of single-nucleotide polymorphisms in SLC16A9/hMCT9
Atsushi Yamaguchi, Yuto Mukai, Tomoya Sakuma, Yudai Suganuma, Ayako Furugen, Katsuya Narumi, Masaki Kobayashi
Life Sciences, 122205, 122205, Elsevier BV, 2023年10月, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）

Beneficial effects of a new neuroprotective compound in neuronal cells and MPTP-administered mouse model of Parkinson's disease.
Izumi Kato, Yudai Ogawa, Fumika Yakushiji, Jiro Ogura, Masaki Kobayashi, Naoya Shindo, Satoshi Ichikawa, Katsumi Maenaka, Masahiro Sakaitani
Chemical communications (Cambridge, England), 59, 82, 12306, 12309, 2023年09月27日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, A new compound, a derivative of 3,4,5-trimethoxy-N-phenyl benzamide bearing an 8''-methylimidazopyridine moiety, is found to demonstrate neuroprotective effects by preventing cell death caused by oxidative stress. The compound possesses high solubility and metabolic stability, and inhibits MPTP-induced effects in vivo, indicating high potential as a therapeutic drug for Parkinson's disease.

Development of a risk prediction model for surgical site infection after lower third molar surgery.
Akira Yamagami, Katsuya Narumi, Yoshitaka Saito, Ayako Furugen, Shungo Imai, Yoshimasa Kitagawa, Yoichi Ohiro, Ryo Takagi, Yoh Takekuma, Mitsuru Sugawara, Masaki Kobayashi
Oral diseases, 2023年09月27日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: There is little evidence regarding risk prediction for surgical site infection (SSI) after lower third molar (L3M) surgery. METHODS: We conducted a nested case-control study to develop a multivariable logistic model for predicting the risk of SSI after L3M surgery. Data were obtained from Hokkaido University Hospital from April 2013 to March 2020. Multiple imputation was applied for the missing values. We conducted decision tree (DT) analysis to evaluate the combinations of factors affecting SSI risk. RESULTS: We identified 648 patients. The final model retained the available distal space (Pell & Gregory II [p = 0.05], Pell & Gregory III [p < 0.01]), depth (Pell & Gregory B [p < 0.01], Pell & Gregory C [p < 0.01]), surgeon's experience (3-10 years [p = 0.25], <3 years [p < 0.01]), and simultaneous extraction of both L3M [p < 0.01]; the concordance-statistic was 0.72. The DT analysis demonstrated that patients with Pell and Gregory B or C and simultaneous extraction of both L3M had the highest risk of SSI. CONCLUSIONS: We developed a model for predicting SSI after L3M surgery with adequate predictive metrics in a single center. This model will make the SSI risk prediction more accessible.

Simple and validated method to quantify lacosamide in human breast milk and plasma using UPLC/MS/MS and its application to estimate drug transfer into breast milk.
Ayako Furugen, Ayako Nishimura, Takeshi Umazume, Hina Ishikawa, Katsuya Narumi, Masaki Kobayashi
Journal of pharmaceutical health care and sciences, 9, 1, 26, 26, 2023年09月01日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Epilepsy is a common neurological disorder. Lacosamide is a third-generation antiepileptic drug used to treat partial-onset seizures. Limited information is currently available on the transfer of lacosamide to breast milk. To facilitate studies on the safety of lacosamide use during breastfeeding, we aimed to develop a method to quantify lacosamide in human breast milk and plasma using ultra-performance liquid chromatography/tandem mass spectrometry. METHODS: Fifty microliters of breast milk or plasma was used, and samples were prepared by protein precipitation using methanol containing lacosamide-d3 as an internal standard (IS). Chromatography was performed using an ACQUITY HSS T3 column with an isocratic flow of 10 mM ammonium acetate solution/methanol (70:30, v/v). Lacosamide and IS were detected by multiple reaction monitoring in positive ion electrospray mode. The run time was 3.5 min. RESULTS: Calibration curves were linear and in the range of 0.5 to 100 ng/mL both in breast milk and plasma. The validation assessment indicated that precision, accuracy, matrix effects, selectivity, dilution integrity, and stability were acceptable. The developed method was successfully applied to quantify lacosamide in breast milk and plasma obtained from a volunteer who had been orally administered lacosamide twice a day (100 mg × 2). Relative infant dose of lacosamide was estimated to be 14.6% in breast milk at five time points. CONCLUSIONS: We developed a simple and robust method to quantify of lacosamide in human breast milk and plasma. This method could be useful for in future studies investigating the safety of lacosamide use during breastfeeding.

Atorvastatin Exerts More Selective Inhibitory Effects on hMCT2 than on hMCT1 and hMCT4.
Atsushi Yamaguchi, Yuto Mukai, Tomoya Sakuma, Ayako Furugen, Katsuya Narumi, Masaki Kobayashi
Anticancer research, 43, 7, 3015, 3022, 2023年07月, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND/AIM: Human monocarboxylate transporter 1 (hMCT1), hMCT2, and hMCT4 transport monocarboxylates, such as L-lactate and pyruvate, with pH dependency. They are often over-expressed in various cancer cells and mediate the energy balance and pH homeostasis. Therefore, hMCT inhibitors can potentially be used as anticancer drugs. However, isoform-selective inhibitors have not yet been well-characterized. In addition, several statins and 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors have been reported to inhibit hMCTs, but their selectivity has not yet been evaluated. In this study, we aimed to determine whether statins could inhibit hMCT1, hMCT2, and hMCT4. MATERIALS AND METHODS: We expressed hMCT1, hMCT2, and hMCT4 in a heterologous expression system of Xenopus oocytes and performed inhibitory experiments with various statins (fluvastatin, atorvastatin, simvastatin, rosuvastatin, pravastatin, and pitavastatin). As the three-dimensional structure of hMCT2 has been recently reported, docking simulations of statins and their structures were also performed to estimate the inhibition site. RESULTS: All statins inhibited the transport activities of hMCT1, hMCT2, and hMCT4. In addition, atorvastatin was found to be a potent isoform-selective inhibitor of hMCT2. Docking simulation indicated that atorvastatin could interact with a site surrounded by transmembrane (TM)-2, TM11, and intracellular helix in the TM6/7loop. Therefore, targeting this site may lead to the discovery of more potent hMCT2-selective inhibitors. CONCLUSION: Atorvastatin exerts selective inhibitory effects on hMCT2. These findings provide insights into the inhibitory mechanism of statins against hMCT1, hMCT2, and hMCT4 and may aid in the development of novel anticancer agents.

Effects of valproic acid on syncytialization in human placental trophoblast cell lines.
Nanami Ohyama, Ayako Furugen, Riko Sawada, Ryoichi Aoyagi, Ayako Nishimura, Takeshi Umazume, Katsuya Narumi, Masaki Kobayashi
Toxicology and applied pharmacology, 474, 116611, 116611, 2023年06月27日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, The placenta is a critical organ for fetal development and a healthy pregnancy, and has multifaceted functions (e.g., substance exchange and hormone secretion). Syncytialization of trophoblasts is important for maintaining placental functions. Epilepsy is one of the most common neurological conditions worldwide. Therefore, this study aimed to reveal the influence of antiepileptic drugs, including valproic acid (VPA), carbamazepine, lamotrigine, gabapentin, levetiracetam, topiramate, lacosamide, and clobazam, at clinically relevant concentrations on syncytialization using in vitro models of trophoblasts. To induce differentiation into syncytiotrophoblast-like cells, BeWo cells were treated with forskolin. Exposure to VPA was found to dose-dependently influence syncytialization-associated genes (ERVW-1, ERVFRD-1, GJA1, CGB, CSH, SLC1A5, and ABCC4) in differentiated BeWo cells. Herein, the biomarkers between differentiated BeWo cells and the human trophoblast stem model (TSCT) were compared. In particular, MFSD2A levels were low in BeWo cells but abundant in TSCT cells. VPA exposure affected the expression of ERVW-1, ERVFRD-1, GJA1, CSH, MFSD2A, and ABCC4 in differentiated cells (ST-TSCT). Furthermore, VPA exposure attenuated BeWo and TSCT cell fusion. Finally, the relationships between neonatal/placental parameters and the expression of syncytialization markers in human term placentas were analyzed. MFSD2A expression was positively correlated with neonatal body weight, head circumference, chest circumference, and placental weight. Our findings have important implications for better understanding the mechanisms of toxicity of antiepileptic drugs and predicting the risks to placental and fetal development.

Association between α-defensin 5 and the expression and function of P-glycoprotein in differentiated intestinal Caco-2 cells.
Genki Yasuda, Atsuhito Kubota, Keisuke Okamoto, Katsuya Narumi, Ayako Furugen, Izumi Kato, Ayako Mori, Yoshitaka Saito, Takashi Satoh, Natsuko Takahashi-Suzuki, Ken Iseki, Masaki Kobayashi
Biopharmaceutics & drug disposition, 2023年06月05日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, α-Defensin 5 is known to be secreted by Paneth cells in the small intestine and plays an important role in eliminating pathogenic microorganisms. It has been reported that a decrease in α-defensin 5 level in the human small intestine is a risk of inflammatory bowel disease (IBD). Furthermore, P-glycoprotein (P-gp), a member of the ATP-binding cassette transporter superfamily, encoded by the ABCB1/MDR1 gene, plays an important role in the front line of host defense by protecting the gastrointestinal barrier from xenobiotic accumulation and may contribute to the development and persistence of IBD. Therefore, we examined the relationship between α-defensin 5 and the expression and function of P-gp using a human gastrointestinal model cell line (Caco-2). We found that MDR1 mRNA and P-gp protein level were increased in Caco-2 cells as well as α-defensin 5 secretion corresponded with the duration of cell culture. Exposure to α-defensin 5 peptide and recombinant tumor necrosis factor-α (TNF-α) significantly increased the expression and function P-gp. The mRNA levels of interleukin (IL)-8, IL-6, TNF-α, IL-1β, and IL-2 were also increased following exposure to TNF-α, similar to α-defensin 5 treatment. These results suggest that α-defensin 5 regulates P-gp expression and function by increasing TNF-α expression in Caco-2 cells.

5-Oxoproline Enhances 4-Hydroxytamoxifen-induced Cytotoxicity by Increasing Oxidative Stress in MCF-7 Breast Cancer Cells.
Takanobu Nadai, Katsuya Narumi, Yuto Mukai, Hinata Ueda, Ayako Furugen, Yoshitaka Saito, Masaki Kobayashi
Anticancer research, 43, 3, 1113, 1120, 2023年03月, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND/AIM: Monocarboxylate transporters (MCTs) transport short-chain monocarboxylates, such as lactate, and have been reported to be related to poor prognosis in breast cancer. Our previous studies showed that a high glucose state altered MCT expression and changed the sensitivity of the tamoxifen active metabolite 4-hydroxytamoxifen (4-OHT) via hypoxia-inducible factor-1α (HIF-1α) protein expression. We hypothesized that MCT inhibitors affect 4-OHT-induced cytotoxicity under normal glucose conditions by decreasing HIF-1α protein expression. To test this hypothesis, we evaluated the combined effect of MCT inhibitor and 4-OHT using the estrogen receptor (ER)-positive breast cancer cell line MCF-7, under normal glucose conditions. MATERIALS AND METHODS: Expression of MCTs and oxidative stress markers was evaluated by real-time PCR. Cell viability was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Reactive oxygen species (ROS) were measured using the cell permeability probe 2',7'-dichlorodihydrofluorescein diacetate. RESULTS: MCT1 expression increased under normal glucose conditions. The MCT1 substrate/inhibitor, 5-oxoproline (5-OP), enhanced 4-OHT-induced cytotoxicity. Bindarit, a selective MCT4 inhibitor, decreased 4-OHT sensitivity, similar to results of our previous study under high glucose conditions. In contrast, the combination of 5-OP and 4-OHT decreased ATP levels compared with that by 4-OHT alone in MCF-7 cells. Furthermore, 5-OP significantly increased the ROS production induced by 4-OHT. CONCLUSION: 5-OP enhances 4-OHT-induced cytotoxicity in ER-positive breast cancer cells under normal glucose conditions.

Monocarboxylate transporter 4 involves in energy metabolism and drug sensitivity in hypoxia
Atsushi Yamaguchi, Yuto Mukai, Tomoya Sakuma, Katsuya Narumi, Ayako Furugen, Yuma Yamada, Masaki Kobayashi
Scientific Reports, 13, 1, Springer Science and Business Media LLC, 2023年01月27日, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）, Abstract

Metabolic reprogramming of cancer cells is a potential target for cancer therapy. It is also known that a hypoxic environment, one of the tumor microenvironments, can alter the energy metabolism from oxidative phosphorylation to glycolysis. However, the relationship between hypoxia and drug sensitivity, which targets energy metabolism, is not well known. In this study, A549 cells, a cell line derived from lung adenocarcinoma, were evaluated under normoxia and hypoxia for the sensitivity of reagents targeting oxidative phosphorylation (metformin) and glycolysis (α-cyano-4-hydroxycinnamic acid [CHC]). The results showed that a hypoxic environment increased the expression levels of monocarboxylate transporter (MCT) 4 and hypoxia-induced factor-1α (HIF-1α), whereas MCT1 and MCT2 expression did not vary between normoxia and hypoxia. Furthermore, the evaluation of the ATP production ratio indicated that glycolysis was enhanced under hypoxic conditions. It was then found that the sensitivity to metformin decreased while that to CHC increased under hypoxia. To elucidate this mechanism, MCT4 and HIF-1α were knocked down and the expression level of MCT4 was significantly decreased under both conditions. In contrast, the expression of HIF-1α was decreased by HIF-1α knockdown and increased by MCT4 knockdown. In addition, changes in metformin and CHC sensitivity under hypoxia were eliminated by the knockdown of MCT4 and HIF-1α, suggesting that MCT4 is involved in the phenomenon described above. In conclusion, it was shown that the sensitivity of reagents targeting energy metabolism is dependent on their microenvironment. As MCT4 is involved in some of these mechanisms, we hypothesized that MCT4 could be an important target molecule for cancer therapy.

Oral administration of linoleic acid immediately before glucose load ameliorates postprandial hyperglycemia.
Yuta Yamamoto, Katsuya Narumi, Naoko Yamagishi, Toshio Nishi, Takao Ito, Ken Iseki, Masaki Kobayashi, Yoshimitsu Kanai
Frontiers in pharmacology, 14, 1197743, 1197743, 2023年, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Introduction: Fatty acids are a major nutrient in dietary fat, some of which are ligands of long-chain fatty acid receptors, including G-protein-coupled receptor (GPR) 40 and GPR120. Pretreatment with GPR40 agonists enhanced the secretion of insulin in response to elevating blood glucose levels after glucose load in a diabetes model, but pretreatment with GPR120 agonist did not ameliorate postprandial hyperglycemia. This study examined whether oral administration of linoleic acid (LA), a GPR40 and GPR120 agonist, immediately before glucose load would affect the elevation of postprandial blood glucose levels in rats. Methods: Male rats and rats with type 1 diabetes administered streptozocin were orally administered LA, trilinolein, α-linolenic acid (α-LA), oleic acid, TAK-875, or TUG-891 immediately before glucose load. Blood glucose levels were measured before, then 15, 30, 60 and 120 min after glucose load. CACO-2 cells were used to measure the uptake of [14C] α-MDG for 30 min with or without LA. Gastric content from rats administered LA was collected 15 and 30 min after glucose load, and blood samples were collected for measurement of glucagon-like peptide 1 (GLP-1) and cholecystokinin concentrations. Results: The elevation of postprandial blood glucose levels was slowed by LA but not by trilinolein in rats without promotion of insulin secretion, and this effect was also observed in rats with type 1 diabetes. The uptake of α-MDG, an SGLT-specific substrate, was, however, not inhibited by LA. Gastric emptying was slowed by LA 15 min after glucose load, and GLP-1, but not cholecystokinin, level was elevated by LA 15 min after glucose load. TUG-891, a GPR120 agonist, ameliorated postprandial hyperglycemia but TAK-875, a GPR40 agonist, did not. Pretreatment with AH7614, a GPR120 antagonist, partially canceled the improvement of postprandial hyperglycemia induced by LA. α-LA, which has high affinity with GPR120 as well as LA, slowed the elevation of postprandial blood glucose levels, but oleic acid, which has lower affinity with GPR120 than LA, did not. Conclusion: Oral administration of LA immediately after glucose load ameliorated postprandial hyperglycemia due to slowing of gastric emptying via promotion of GLP-1 secretion. The mechanisms may be associated with GPR120 pathway.

Diabetes mellitus degenerates cisplatin-induced nephrotoxicity in short hydration method: a propensity score-matching analysis.
Yoshitaka Saito, Tatsuhiko Sakamoto, Yoh Takekuma, Masaki Kobayashi, Keisuke Okamoto, Naofumi Shinagawa, Yasushi Shimizu, Ichiro Kinoshita, Mitsuru Sugawara
Scientific Reports, 12, 1, 21819, 21819, 2022年12月17日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Cisplatin (CDDP)-induced nephrotoxicity (CIN) is dose-limiting. We revealed that co-administration of non-steroid anti-inflammatory drugs and baseline comorbidity of diabetes mellitus (DM) are associated with CIN development in the short hydration method; however, the results were accessorily obtained without appropriate power calculation. This study aimed to demonstrate the influence of DM complications on CIN incidence in a real-world setting. Lung cancer patients receiving CDDP (≥ 75 mg/m2)-containing regimens with a short hydration method (n = 227) were retrospectively evaluated. The patients were divided into control and baseline DM complication groups. The primary endpoint was the evaluation of CIN incidence between the groups. Propensity score-matching was performed to confirm the robustness of the primary analysis results. CIN occurred in 6.8% of control and 27.0% of DM patients, respectively, with a significant difference in all-patient populations (P = 0.001). In addition, variation of serum creatinine and creatinine clearance significantly worsened in DM patients. Similar results were obtained in a propensity-matched population. Multivariate logistic regression analysis found that DM complication is a singular risk factor for CIN development (adjusted odds ratio; 4.31, 95% confidence interval; 1.62-11.50, P = 0.003). In conclusion, our study revealed that baseline DM complications significantly worsen CIN.

Association between skin immune-related adverse events (irAEs) and multisystem irAEs during PD-1/PD-L1 inhibitor monotherapy.
Atsushi Yamaguchi, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Yoh Takekuma, Naofumi Shinagawa, Yasushi Shimizu, Hirotoshi Dosaka-Akita, Mitsuru Sugawara, Masaki Kobayashi
Journal of cancer research and clinical oncology, 149, 4, 1659, 1666, 2022年11月08日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, PURPOSE: Patients treated with immune checkpoint inhibitors (ICIs) often develop immune-related adverse events (irAEs) in various organs of the body. However, the patient factors associated with the development of multisystem irAEs are not well known. Skin irAEs most frequently occur and appear early after ICI treatment initiation. They may be a predictive marker for the development of multisystem irAEs, and their occurrence should be evaluated. METHODS: Data of patients receiving ICI monotherapy for lung cancer, melanoma, and head and neck cancer treatment were retrospectively evaluated (n = 207); the single irAE development group (n = 69) was compared with the multisystem irAE development group (n = 37). The primary endpoint was the comparison of the incidence of skin irAEs between the two groups. RESULTS: Skin, thyroid, and hepatic irAEs were associated with the development of multisystem irAEs (odds ratio: 3.30, 95% confidence interval: 1.27-8.52, p = 0.01 for skin; 5.07, 2.09-12.3, p = 0.0003 for thyroid; 10.63, 1.19-94.7, p = 0.03 for hepatic). Skin irAEs were the most common type (65.0% of total participants) and appeared earlier than other irAEs, except for gastrointestinal and ocular irAEs (median time to onset of skin irAEs: 7.5 weeks). Skin irAEs occurred more frequently in the multisystem irAE group (81.0%) than in the single irAE group (56.5%, p = 0.02). CONCLUSION: Skin irAEs can be a useful predictive marker for multisystem irAE development due to ICI treatment. Consequently, patients with skin irAEs should be treated and monitored for other types of irAEs.

Involvement of SLC16A1/MCT1 and SLC16A3/MCT4 in l-lactate transport in the hepatocellular carcinoma cell line.
Yuto Mukai, Atsushi Yamaguchi, Tomoya Sakuma, Takanobu Nadai, Ayako Furugen, Katsuya Narumi, Masaki Kobayashi
Biopharmaceutics & drug disposition, 43, 5, 183, 191, 2022年09月14日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Fourteen isoforms of the monocarboxylate transporter (MCT) have been reported. Among the MCT isoforms, MCT1, MCT2, and MCT4 play a role in l-lactate/proton co-transport and are involved in the balance of intracellular energy and pH. Therefore, MCT1, MCT2, and MCT4 are associated with energy metabolism processes in normal and pathological cells. In the present study, we evaluated the expression of MCT1, MCT2, and MCT4 and the contribution of these three MCT isoforms to l-lactate uptake in hepatocellular carcinoma (HCC) cells. In HepG2 and Huh-7 cells, l-lactate transport was pH-dependent, which is characteristic of MCT1, MCT2, and MCT4. Furthermore, l-lactate uptake was selectively inhibited by MCT1 and MCT4 inhibitors in HepG2 and Huh-7 cells. Kinetic analysis of HepG2 cells demonstrated that l-lactate uptake was biphasic. Although the knockdown of MCT1 and MCT4 in the HepG2 cells decreased the uptake of l-lactate, the knockdown of MCT2 had no effect on the uptake of l-lactate. Consequently, we concluded that both MCT1 and MCT4 were involved in the transport of l-lactate in HepG2 and Huh-7 cells at pH 6.0. In contrast, PXB-cells, freshly isolated hepatocytes from humanized mouse livers, showed lower MCT4 expression and l-lactate uptake at pH 6.0 compared to that in HCC cell lines. In conclusion, MCT4, which contributes to l-lactate transport in HCC cells, is significantly different in HCC compared to normal hepatocytes, and has potential as a target for HCC treatment. This article is protected by copyright. All rights reserved.

The rs35217482 (T755I) single-nucleotide polymorphism in aldehyde oxidase-1 attenuates protein dimer formation and reduces the rates of phthalazine metabolism
Hinata Ueda, Katsuya Narumi, Ayako Furugen, Yoshitaka Saito, Masaki Kobayashi
Drug Metabolism and Disposition, DMD, AR, American Society for Pharmacology & Experimental Therapeutics (ASPET), 2022年07月16日, ［査読有り］, ［最終著者, 責任著者］
研究論文（学術雑誌）

Suitability of Oral Rehydration Solution (ORS) for Use in the Cisplatin Short Hydration Method.
Yoshitaka Saito, Yoh Takekuma, Masaki Kobayashi, Naofumi Shinagawa, Takuro Noguchi, Satoshi Takeuchi, Yasushi Shimizu, Ichiro Kinoshita, Hirotoshi Dosaka-Akita, Mitsuru Sugawara
Anticancer research, 42, 6, 3185, 3193, 2022年06月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND/AIM: Short hydration is a method to change partial intravenous hydration to oral to administer cisplatin (CDDP); however, the most suitable form of oral hydration is unknown. This study aimed to determine whether oral rehydration solution (ORS) affects CDDP-induced nephrotoxicity (CIN) and electrolyte imbalance. PATIENTS AND METHODS: Lung cancer patients (n=200) who had received CDDP-including regimens (CDDP dosage ≥75 mg/m2) were retrospectively evaluated. We used logistic analysis to evaluate whether ORS intake could be a preventive factor for CIN (≥grade 2 serum creatinine elevation). Moreover, incidence of CIN and electrolyte imbalance and the variation in serum creatinine and electrolyte levels were compared between ORS and non-ORS (control) patients. RESULTS: CIN occurred in 9.8% of ORS patients, and 7.5% of non-ORS patients (p=0.79). The variation in serum creatinine level was also similar in both groups. Multivariate analysis suggested that ORS intake does not affect CIN, although CIN was associated with the coadministration of non-steroidal anti-inflammatory drugs and the presence of diabetes mellitus. The variations in serum electrolyte levels did not differ, and incidence of hyponatremia, hypokalemia, and hypochloremia was also similar between the groups. Moreover, patients in ORS group experienced significantly more anorexia compared to controls, and approximately 40% of the patients were unable to continue ORS intake. CONCLUSION: ORS intake in CDDP short hydration regimens does not affect CIN and CDDP-induced electrolyte imbalance; however, its intake is associated with the incidence of anorexia suggesting that ORS should not be used for oral hydration.

Uptake of antiepileptic drugs in forskolin-induced differentiated BeWo cells: Alteration of gabapentin transport.
Mai Koishikawa, Ayako Furugen, Nanami Ohyama, Katsuya Narumi, Shuhei Ishikawa, Masaki Kobayashi
Xenobiotica; the fate of foreign compounds in biological systems, 52, 4, 1, 30, 2022年06月01日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Previous studies have indicated that the expression levels of several transporters are altered during placental trophoblast differentiation. However, changes in the transport activities of therapeutic agents during differentiation must be comprehensively characterised. Antiepileptic drugs, including gabapentin (GBP), lamotrigine (LTG), topiramate, and levetiracetam, are increasingly prescribed during pregnancy. The objective of this study was to elucidate differences in the uptake of antiepileptic drugs during the differentiation process.Human placental choriocarcinoma BeWo cells were used as trophoblast models. For differentiation into syncytiotrophoblast-like cells, cells were treated with forskolin.The uptake of GBP and LTG was lower in differentiated BeWo cells than in undifferentiated cells. In particular, the maximum uptake rate of GBP transport was decreased in differentiated BeWo cells. Furthermore, GBP transport was trans-stimulated by the amino acids His and Met. We investigated the profiles of amino acids in undifferentiated and differentiated BeWo cells. Supplementation with His and Met, which demonstrated trans-stimulatory effects on GBP uptake, restored GBP uptake in differentiated cells. The findings of this study suggest that drug transport in BeWo cells can be altered before and after differentiation, and that the altered GBP uptake could be mediated by the intracellular amino acid status.

Evaluation of the strategies to reduce third-generation oral cephalosporins in dentistry at a Japanese academic hospital: An interrupted time series analysis.
Akira Yamagami, Katsuya Narumi, Yoshitaka Saito, Ayako Furugen, Shungo Imai, Yoshimasa Kitagawa, Yoichi Ohiro, Ryo Takagi, Yoh Takekuma, Mitsuru Sugawara, Masaki Kobayashi
Journal of clinical pharmacy and therapeutics, 47, 7, 1010, 1019, 2022年03月07日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, WHAT IS KNOWN AND OBJECTIVE: Third-generation oral cephalosporins, especially cefcapene-pivoxil (CFPN-PI), have been used frequently in the Japanese dental field. In December 2014 and April 2016, the newly published clinical guidelines recommended the use of amoxicillin (AMPC). Thus, it is important to evaluate the impact of these guidelines on the prescription profiles of prophylactic antibiotics, clinical outcomes and cost-effectiveness of antibiotics. METHODS: We conducted a retrospective study to analyse an interrupted time series analysis from April 2013 to March 2020 at the Department of Dentistry of Hokkaido University Hospital. A segmented regression model was used to estimate the changes in the incidence of infectious complications following tooth extraction. Prescribed antibiotic data were evaluated via days of therapy (DOT). Antibiotic costs were calculated in terms of the Japanese yen (JPY). RESULTS AND DISCUSSION: We identified 17,825 eligible patients. The incidence rates of infectious complications (SSI + dry socket) and SSI after tooth extraction were 3.2% and 2.2%, respectively, during the entire period. The extraction of impacted third molars corresponded to 5.0% and 3.4%, respectively. However, their incidence rates were not significantly different during this period. The use of prophylactic antibiotics and antibiotic cost showed consistent trends following the implementation of guidelines. The mean DOT of CFPN-PI decreased (ranging from 4893.6 DOTs/1000 patients [March 2013 to November 2014] to 3856.4 DOTs/1000 patients [December 2014 to March 2016]; p < 0.001, and from 3856.4 DOTs/1000 patients [December 2014 to March 2016] to 2293.9 DOTs/1000 patients [April 2016 to March 2020]; p < 0.001). In contrast, the mean DOT of AMPC was found to be increased (ranging from 1379.7 DOTs/1000 patients [March 2013 to November 2014] to 3236.3 DOTs/1000 patients [December 2014 to March 2016]; p < 0.001, and from 3236.3 DOTs/1000 patients [December 2014 to March 2016] to 4597.8 DOTs/1000 patients [April 2016 to March 2020]; p < 0.001). The mean monthly cost was decreased (ranging from 905.3 JPY [March 2013 to November 2014] to 788.7 JPY [December 2014 to March 2016]; p = 0.003, and from 788.7 JPY [December 2014 to March 2016] to 614.0 JPY [April 2016 to March 2020]; p < 0.001). WHAT IS NEW AND CONCLUSION: After December 2014, prophylactic antibiotics were switched from CFPN-PI to AMPC, and the incidence rate of infectious complications was not significantly different over time. However, changing antibiotics is useful from a cost-effectiveness perspective.

[Development of an Online Role-play-based Medical Interview Training Method for Fourth-year Pharmacy Students].
Ayako Mori, Izumi Kato, Hitoshi Kashiwagi, Shungo Imai, Katsuya Narumi, Yuki Sato, Ayako Furugen, Yuma Yamada, Masaki Kobayashi
Yakugaku zasshi : Journal of the Pharmaceutical Society of Japan, 142, 6, 661, 674, 2022年, ［査読有り］, ［最終著者, 責任著者］, ［国内誌］
日本語, 研究論文（学術雑誌）, With the coronavirus disease 2019 pandemic, businesses are rapidly expanding their online practices, and the online medical care system has been established and is growing. The field of pharmacy education is also looking for ways to conduct practical online training. Hence, we developed an online role-play-based medical interview training method for fourth-year pharmacy students. The purpose of this study was to describe in detail this method and to clarify the effect of online on medical interviewing practice. The training sessions were conducted using video teleconferencing software. Two settings were used for the role-play scenarios: the pharmacy and hospital. To evaluate the effectiveness of the sessions, a questionnaire was sent to the students, and the results were analyzed using text mining. The most important requirement for successfully conducting the interviews was a stable voice connection, and we reduced audio interruptions and delays by connecting the host personal computer to a wired local area network. We also solved the problem of howling when multiple terminals were installed in the same room by muting all devices in the room. Results of the analysis of the questionnaires suggested that students were more tense online. We also found that students perceived a difference between online and face-to-face interviews in terms of eye contact and the presentation of documents. In this way, we succeeded in conducting smooth online role-playing sessions while taking countermeasures against infection. In the future, it will be necessary to devise nonverbal communication methods and digital methods of presenting the training material.

5-Aminosalicylic Acid, A Weak Agonist for Aryl Hydrocarbon Receptor That Induces Splenic Regulatory T Cells.
Atsuhito Kubota, Masaru Terasaki, Rie Takai, Masaki Kobayashi, Ryuta Muromoto, Hiroyuki Kojima
Pharmacology, 107, 1-2, 1, 7, 2021年12月16日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, INTRODUCTION: 5-Aminosalicylic acid (5-ASA) is widely used as a key drug in inflammatory bowel disease. It has been recently reported that 5-ASA induces CD4 + Foxp3 + regulatory T cells (Tregs) in the colon via the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that regulates inflammation. However, the role of 5-ASA as an AhR agonist that induces Tregs in the spleen remains unknown. METHODS: In the present study, we investigated these themes using an AhR-mediated transactivation assay and flow cytometry analysis. The experiments were conducted by using DR-EcoScreen cells and C57BL/6 mice. RESULTS: The DR-EcoScreen cell-based transactivation assay revealed that 5-ASA acted as a weak AhR agonist at concentrations of ≥300 μM (1.31-1.45-fold), and that a typical AhR agonist, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), activated AhR at a concentration of 0.1 nM (22.8-fold). In addition, the treatment of mouse splenic cells with 300 μM 5-ASA in a primary culture assay significantly induced CD4+CD25 + Foxp3 + Tregs (control vs. 5-ASA: 9.0% vs. 12.65%, p < 0.05), while 0.1 nM TCDD also showed significant induction of Tregs (control vs. TCDD: 9.0% vs. 14.1%, p < 0.05). Interestingly, this induction was eliminated by co-treatment with an AhR antagonist, CH-223191. DISCUSSION: These results suggest that 5-ASA is a weak agonist of AhR and thereby induces Tregs in spleen cells. Our findings may provide useful insights into the mechanism by which 5-ASA regulates inflammation.

Diclofenac potentiates the antitumor effect of cisplatin in a xenograft mouse model transplanted with cisplatin-resistant cells without enhancing cisplatin-induced nephrotoxicity.
Keisuke Okamoto, Hinata Ueda, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Masaki Kobayashi
Drug metabolism and pharmacokinetics, 41, 100417, 100417, 2021年08月16日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Cisplatin (CDDP) is a well-known anticancer agent, and CDDP-induced nephrotoxicity (CIN) is one of the most serious adverse effects. Previously, we revealed that while celecoxib reduces CIN, diclofenac does not appear to enhance it. Furthermore, we reported that diclofenac additively enhances the cytotoxic effect of CDDP on CDDP-resistant A549 cells (A549/DDP cells) and their spheroids. In addition, celecoxib reduces the cytotoxic effect of CDDP on A549/DDP cells while demonstrating an anticancer effect; however, it enhanced the effect of CDDP cytotoxicity on spheroids. Therefore, we evaluated the effects of diclofenac or celecoxib on CIN and the antitumor effect of CDDP in a xenograft mouse model transplanted with A549/DDP cells. Although CDDP did not decrease tumor size and tumor weight, these parameters were significantly reduced following co-administration with diclofenac when compared with the control group. Conversely, celecoxib marginally suppressed the antitumor effect of CDDP. Moreover, CDDP increased the mRNA levels of kidney injury molecule 1 (Kim-1), a renal disorder marker, in the kidneys of xenograft mice; treatment with celecoxib and diclofenac did not impact Kim-1 mRNA levels increased by CDDP. In conclusion, diclofenac potentiated the antitumor effect of CDDP without enhancing CIN.

In vitro and in vivo evaluation of organic anion-transporting polypeptide 2B1-mediated pharmacokinetic interactions by apple polyphenols.
Yuka Takahashi, Katsuya Narumi, Takanobu Nadai, Hinata Ueda, Taiki Yamamura, Ayako Furugen, Masaki Kobayashi
Xenobiotica; the fate of foreign compounds in biological systems, 51, 11, 1, 36, 2021年08月16日, ［査読有り］, ［最終著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Organic anion-transporting polypeptide (OATP) 2B1 plays a critical role in the intestinal absorption of substrate drugs. Apple juice reportedly interacts with OATP2B1 substrate drugs. The purpose of this study was to investigate the effect of two apple polyphenols, phloretin and phloridzin, on OATP2B1-mediated substrate transport in vitro and to evaluate the effect of phloretin on rosuvastatin pharmacokinetics in rats.In vitro studies revealed that both polyphenols inhibited OATP2B1-mediated uptake of estrone-3-sulfate. Despite preincubation with phloretin and subsequent washing, the inhibitory effect was retained. Phloretin markedly decreased OATP2B1-mediated rosuvastatin uptake, with an IC50 value of 3.6 μM.On coadministering rosuvastatin and phloretin in rats, the plasma concentration of rosuvastatin 10 min after oral administration was significantly lower than that in the vehicle group. The area under the plasma concentration-time curve of rosuvastatin was not significant, showing a tendency to decrease in the phloretin group when compared with the vehicle group. The in-situ rat intestinal loop study revealed the inhibitory effect of phloretin on rosuvastatin absorption.Phloretin has potent and long-lasting inhibitory effects on OATP2B1 in vitro. Phloretin may inhibit OATP2B1-mediated intestinal absorption of rosuvastatin; however, it failed to significantly impact the systemic exposure of rosuvastatin in rats.

Antioxidant effect of ascorbic acid against cisplatin-induced nephrotoxicity and P-glycoprotein expression in rats.
Keisuke Okamoto, Fumi Kitaichi, Yoshitaka Saito, Hinata Ueda, Katsuya Narumi, Ayako Furugen, Masaki Kobayashi
European journal of pharmacology, 909, 174395, 174395, 2021年07月29日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Cisplatin (CDDP) is a highly potent anticancer drug that is widely used in the treatment of several cancers. CDDP-induced nephrotoxicity (CIN) is one of the most significant adverse effects, and oxidative stress is thought to be one of the mechanisms underlying CIN. Although there are some studies available on the variability in transporter expression in the kidney after a single CDDP dose, none have reported the change in renal transporter expression after multiple CDDP dose administrations. P-glycoprotein (P-gp), a transporter, is reported to be induced by oxidative stress. Ascorbic acid is a vitamin with antioxidant potential and therefore, may regulate the expression of P-gp transporter and affect CIN. In the present study, our aim was to assess the variability in expression of several renal transporters after multiple CDDP dose administrations and the antioxidant effect of ascorbic acid against transporter expression and CIN. Multiple doses of CDDP affected markers of kidney injury and antioxidants in the kidneys. Also, the expression of P-gp, breast cancer resistance protein, and multidrug resistance-associated protein 4 was upregulated by CDDP. Using a normal kidney cell line, we demonstrated that ascorbic acid attenuated CDDP-induced cytotoxicity due to its high superoxide scavenging ability. CDDP and ascorbic acid were injected into rats once a week for three weeks, and it was observed that co-administration of ascorbic acid attenuated CIN and regulated antioxidant marker. In addition, ascorbic acid reduced P-gp expression, which was upregulated by CDDP. In conclusion, ascorbic acid may attenuate CIN and reverse P-gp-mediated changes in drug pharmacokinetics.

Kinetic analysis of cystine uptake and inhibition pattern of sulfasalazine in A549 cells.
Keisuke Okamoto, Yoshitaka Saito, Hinata Ueda, Katsuya Narumi, Ayako Furugen, Masaki Kobayashi
Biopharmaceutics & drug disposition, 42, 8, 389, 392, 2021年07月20日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Cystine/glutamate transporter (xCT) is an antiporter involved in cystine uptake and glutamate efflux. However, there are very few reports regarding the kinetic analysis of xCT for cystine uptake using cancer cell lines, as well as the inhibition pattern of sulfasalazine, an inhibitor of xCT, for cystine uptake. Therefore, the purpose of this study was to clarify the kinetics of xCT in A549 cells, human lung cancer cells, and to reveal the inhibition pattern of sulfasalazine. Cystine uptake occurred in a time-dependent manner, with linear cystine uptake observed for 5 min. Additionally, sulfasalazine inhibited cystine uptake in a concentration-dependent manner, presenting an IC50 value of 24.7 ± 5.6 μM. Cystine uptake was saturated with increasing concentration, demonstrating Km and Vmax values of 179.4 ± 26.7 μM and 30.4 ± 2.3 nmol/min/mg protein, respectively. Moreover, during cystine uptake with sulfasalazine, Km and Vmax were >300 μM and 8.0 ± 1.5 nmol/min/mg protein, respectively, suggesting that sulfasalazine might demonstrate a mixed inhibition pattern. Furthermore, xCT siRNA decreased the xCT mRNA level and reduced cystine uptake. In conclusion, xCT was involved in the cystine uptake in A549 cells and sulfasalazine showed a mixed inhibition pattern to xCT.

Risk factor analysis for taxane-associated acute pain syndrome under the dexamethasone prophylaxis.
Yoshitaka Saito, Yoh Takekuma, Masaki Kobayashi, Tatsuhiko Sakamoto, Hiroko Yamashita, Mitsuru Sugawara
Supportive care in cancer : official journal of the Multinational Association of Supportive Care in Cancer, 29, 12, 8059, 8067, 2021年07月06日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: Taxane-associated acute pain syndrome (T-APS) reportedly occurs in approximately 70% of patients undergoing therapy. We have previously reported that additional dexamethasone (DEX) administration attenuates T-APS. The aim of this study was to reveal risk factor(s) associated with the incidence of T-APS under prophylactic DEX administration. METHODS: In total, 143 patients with breast cancer who received docetaxel (75 mg/m2) or paclitaxel (175 mg/m2)-containing treatment regimens were enrolled. DEX (4-8 mg) was orally administered on days 2-4. Risk factors for the incidence of ≥ G2 and all-grade T-APS, as well as T-APS incidence between taxane-containing regimens in the first cycle, were retrospectively evaluated. RESULTS: Approximately 90% of the patients received taxanes for adjuvant or neoadjuvant chemotherapy. Overall, 55% of patients administered 4 mg DEX, whereas 45% received 8 mg DEX. Pegfilgrastim was administered in 27% of patients. Incidence of ≥ G2 and all-grade T-APS was 23.8%, and 69.2%, respectively. Univariate and multivariate analyses revealed that administration of pegfilgrastim is an independent risk factor for the incidence of ≥ G2 and all-grade T-APS; age younger than 55 years is also a risk factor for all-grade T-APS. Moreover, the incidence of ≥ G2 and all-grade T-APS was 45.5% and 81.8% in a paclitaxel regimen, and 22.0% and 68.2% in docetaxel-including regimens, respectively, revealing increased tendency with paclitaxel administration, with no significant differences. CONCLUSION: Pegfilgrastim co-administration is an independent risk factor for ≥ G2 and all-grade T-APS, and age younger than 55 years is a risk factor of all-grade T-APS under prophylactic DEX administration.

Preexisting autoimmune disease is a risk factor for immune-related adverse events: a meta-analysis.
Atsushi Yamaguchi, Yoshitaka Saito, Keisuke Okamoto, Katsuya Narumi, Ayako Furugen, Yoh Takekuma, Mitsuru Sugawara, Masaki Kobayashi
Supportive care in cancer : official journal of the Multinational Association of Supportive Care in Cancer, 2021年06月23日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: Patients with preexisting autoimmune disease (PAD) are often excluded from clinical trials assessing immune checkpoint inhibitors (ICIs). Therefore, the safety of ICI therapy in patients with PAD remains unclear. Herein, we evaluated the incidence of immune-related adverse events (irAEs) in patients with PAD when compared with non-PAD patients. METHODS: We searched MEDLINE/PubMed, Web of Science, and Google Scholar for eligible studies from inception to January 2021. Observational studies reporting the incidence of irAEs in patients with and without PAD were included. We then performed a meta-analysis of eligible studies using forest plots. The primary endpoint of this study was the incidence rate of irAEs between patients with and without PAD. RESULTS: We identified three prospective and three retrospective studies involving 206 patients with PAD and 3078 patients without PAD. In the meta-analysis, 128 patients with PAD (62.1%) experienced irAEs, which occurred in 51.9% of non-PAD patients, resulting in an odds ratio (OR) of 2.14 (95% confidence interval [CI] 1.58-2.89). In the subgroup analysis, the incidence of irAEs was significantly higher in patients with PAD (OR = 2.19, 95% CI [1.55-3.08]). Furthermore, no significant heterogeneity or publication bias was detected, indicating that our meta-analysis could be generalized to clinical settings. CONCLUSION: This meta-analysis demonstrated that PAD was a risk factor for irAE incidence. These results suggest that monitoring the occurrence of irAEs in patients with PAD is required to manage irAEs appropriately.

Effects of valproate, an HDAC inhibitor, on the expression of folate carriers and folate metabolism-related genes in the placenta of rats.
Ayako Furugen, Yuki Kanno, Nanami Ohyama, Yuko Kurosawa, Naoko Jinno, Katsuya Narumi, Ken Iseki, Masaki Kobayashi
Drug metabolism and pharmacokinetics, 40, 100409, 100409, 2021年06月07日, ［査読有り］, ［最終著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Valproate (VPA), an antiepileptic drug, is known to inhibit histone deacetylases (HDACs). Exposure to VPA during pregnancy increases several fetal risks. The maintenance of folate level during pregnancy is essential for adequate fetal development, and the placenta plays a critical role in supplying nutrients to the fetus. The aim of this study was to elucidate the effects of VPA on the gene expression of folate carriers and metabolizing enzymes in the rat placenta at both mid and late gestation periods. Pregnant rats were orally administered VPA on a single day or 4 days (repeated administration). Gene expression of folate carriers (Folr1, Slc19a1, Slc46a1) and metabolizing enzymes (Cth, Mtr, Mtrr, Mthfr, Dhfr) was assessed in the placenta on gestational day (GD) 13 or GD20. In the control rats, the expression of Folr1, Slc46a1, Cth, and Mthfr tended to be upregulated, whereas that of Mtrr and Dhfr was downregulated during gestation; the expression of Slc19a1 and Mtr did not change. Repeated VPA administration reduced the placental expression of Folr1and Mtr on GD20 and increased the expression of Dhfr on GD13 compared with the control. These findings indicate that administration of VPA alters the placental gene expression of folate carriers and metabolism-related enzymes.

Characterization of deoxyribonucleoside transport mediated by concentrative nucleoside transporters.
Taiki Yamamura, Katsuya Narumi, Tsukika Ohata, Hiroshi Satoh, Takao Mori, Ayako Furugen, Masaki Kobayashi, Ken Iseki
Biochemical and biophysical research communications, 558, 120, 125, 2021年04月25日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Human concentrative nucleoside transporters (CNTs) are responsible for cellular uptake of ribonucleosides; however, although it is important to better characterize CNT-subtype specificity to understand the systemic disposition of deoxyribonucleosides (dNs) and their analogs, the involvement of CNTs in transporting dNs is not fully understood. In this study, using COS-7 cells that transiently expressed CNT1, CNT2, or CNT3, we investigated if CNTs could transport not only ribonucleosides but also dNs, i.e., 2'-deoxyadenosine (dAdo), 2'-deoxyguanosine (dGuo), and 2'-deoxycytidine (dCyd). The cellular uptake study demonstrated that dAdo and dGuo were taken up by CNT2 but not by CNT1. Although dCyd was taken up by CNT1, no significant uptake was detected in COS-7 cells expressing CNT2. Similarly, these dNs were transported by CNT3. The apparent Km values of their uptake were as follows: CNT1, Km ＝ 141 μM for dCyd; CNT2, Km ＝ 62.4 μM and 54.9 μM for dAdo and dGuo, respectively; CNT3, Km ＝ 14.7 μM and 34.4 μM for dGuo and dCyd, respectively. These results demonstrate that CNTs contribute not only to ribonucleoside transport but also to the transport of dNs. Moreover, our data indicated that CNT1 and CNT2 selectively transported pyrimidine and purine dNs, respectively, and CNT3 was shown to transport both pyrimidine and purine dNs.

Transport function, regulation, and biology of human monocarboxylate transporter 1 (hMCT1) and 4 (hMCT4).
Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Pharmacology & therapeutics, 226, 107862, 107862, 2021年04月21日, ［査読有り］, ［筆頭著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporter 1 (hMCT1) and 4 (hMCT4) are involved in the proton-dependent transport of monocarboxylates such as L-lactate, which play an essential role in cellular metabolism and pH regulation. hMCT1 and 4 are overexpressed in a number of cancers, and polymorphisms in hMCT1 have been reported to be associated with the prognosis of some cancers. Accordingly, recent advances have focused on the inhibition of these transporters as a novel therapeutic strategy in cancers. To screen for MCT inhibitors for clinical application, it is important to study MCT function and regulation, and the effect of compounds on them, using human-derived cells. In this review, we focus on the transport function, regulation, and biology of hMCT1 and hMCT4, and the effects of genetic variation in these transporters in humans.

The association between SLC16A11 haplotype and lipid metabolism in Japanese patients with type 2 diabetes.
Yuki Kimura, Issei Higuchi, Masaki Kobayashi, Ayako Furugen, Katsuya Narumi, Yuya Suzuki, Hideaki Miyoshi, Akinobu Nakamura, Tatsuya Atsumi, Ken Iseki
Drug metabolism and pharmacokinetics, 37, 100376, 100376, 2021年04月, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Solute carrier (SLC) 16A11 has been reported as a risk gene for type 2 diabetes (T2D). However, the physiological function of SLC16A11 has not yet been clarified, and the relationship between SLC16A11 and T2D condition remains unclear. Therefore, we performed an association analysis between the SLC16A11 genotype and T2D pathology. The SLC16A11 genotype was determined by direct sequencing in 85 Japanese patients with T2D. The genotypes were analyzed by Mann-Whitney's U test and Chi-square test. Six single nucleotide polymorphisms (SNPs) were detected in the SLC16A11 gene, and five of them formed a haplotype (5SNP haplotype). The 5SNP haplotype carriers had significantly higher fasting plasma glucose (FPG), total cholesterol (T-CHO), and low-density lipoprotein cholesterol (LDL-C) than the noncarriers. The SLC16A11 genotype affected the values of laboratory parameters for T2D, particularly of blood lipids. The function of SLC16A11 may be related to lipid metabolism.

Anticancer effects of non-steroidal anti-inflammatory drugs against cancer cells and cancer stem cells.
Keisuke Okamoto, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Ken Iseki, Masaki Kobayashi
Toxicology in vitro : an international journal published in association with BIBRA, 74, 105155, 105155, 2021年03月27日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Certain non-steroidal anti-inflammatory drugs (NSAIDs) are known to have anticancer effects. However, it is unclear whether all NSAIDs have anticancer effects, and thus far, very few studies have compared the antitumor effects among multiple NSAIDs. Therefore, we aimed to identify NSAIDs that enhance the anticancer effect of cisplatin (CDDP); the effects of 17 NSAIDs in lung cancer cells and their spheroids as cancer stem cells (CSCs) were evaluated. Some of the NSAIDs showed cytotoxic effects against A549 and SBC-3 cells and their CDDP-resistant cell lines (A549/DDP and SBC-3/DDP cells, respectively). In addition, co-addition of CDDP and celecoxib, which showed cytotoxic effects, increased the resistance to CDDP by increasing SLC7A11, which is one of the CDDP resistance mechanisms, in A549/DDP and SBC-3/DDP cells. On the other hand, celecoxib also showed antitumor effects on the spheroids of A549/DDP and SBC-3/DDP cells, and enhanced the antitumor effect of CDDP while increasing the mRNA levels of SLC7A11. Moreover, diclofenac was also cytotoxic and enhanced the cytotoxic effect of CDDP in cancer cells and CSCs. In conclusion, some NSAIDs including celecoxib and diclofenac may enhance the therapeutic efficacy of CDDP.

Impact of histamine type-2 receptor antagonists on the anticancer efficacy of gefitinib in patients with non-small cell lung cancer.
Yoshitaka Saito, Yoh Takekuma, Masaki Kobayashi, Naofumi Shinagawa, Yasushi Shimizu, Ichiro Kinoshita, Hirotoshi Dosaka-Akita, Ken Iseki, Mitsuru Sugawara
European journal of clinical pharmacology, 77, 3, 381, 388, 2021年03月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: Gefitinib is one of the standard treatments for non-small cell lung cancer (NSCLC) with epidermal growth factor receptor mutations. It has been reported that acid suppressants (AS) decrease the anti-tumor effect of gefitinib by reducing its solubility. AS is sometimes necessary in cancer patients; however, previous reports have not shown the most compatible AS with gefitinib administration in cancer patients. This study was conducted to determine if histamine type 2 receptor antagonists (H2RAs) can affect the anti-tumor efficacy of gefitinib. METHODS: Eighty-seven patients with NSCLC who were administered gefitinib were retrospectively investigated. Patients who were co-administered H2RA were compared with non-AS control patients. H2RA was administered once a day at about 3-5 or 8-12 h after gefitinib intake. The primary endpoint of this study was progression-free survival (PFS), and secondary endpoints were overall survival (OS), overall response rate (ORR), and adverse effects. RESULTS: Median PFS in H2RA group and control group was 8.0 months and 9.0 months, respectively, with no significant difference (p = 0.82). The incidence of liver dysfunction was significantly less in patients administered H2RA, whereas there were no differences between the two groups with regard to skin toxicity and diarrhea. Multivariate analysis suggested that H2RA co-administration is not a risk factor for worse PFS and OS (hazard ratio of 0.95, 0.86; 95% confidence interval of 0.60-1.48, 0.52-1.43; p = 0.82 and 0.60, respectively). CONCLUSION: This study demonstrated that concomitant administration of H2RA with gefitinib does not affect the efficacy of gefitinib.

Benzodiazepine Concentrations in the Breast milk and Plasma of Nursing Mothers: Estimation of Relative Infant Dose.
Ayako Nishimura, Ayako Furugen, Takeshi Umazume, Seika Kitamura, Mayuko Soma, Kiwamu Noshiro, Yoh Takekuma, Mitsuru Sugawara, Ken Iseki, Masaki Kobayashi
Breastfeeding medicine : the official journal of the Academy of Breastfeeding Medicine, 16, 5, 424, 431, 2021年01月15日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Objective:
Benzodiazepines are common therapies for mental illness and insomnia, and are used during pregnancy and lactation. Although benzodiazepines have been shown to be transferred into breast milk, the amount transferred is small and compatible with breastfeeding. However, information is not available for all drugs. Therefore, we aimed to determine the milk to plasma (M/P) ratio and relative infant dose (RID), which are used as indicators of drug transfer to breast milk, to determine the safety of such drugs for lactating women and breastfeeding infants.
Methods:
The study comprised of 11 pregnant women who visited the obstetrics department of Hokkaido University Hospital (approval number: 017-0131) and Tenshi Hospital (approval number: 103) for childbirth. The samples were analyzed using liquid chromatography-tandem mass spectrometry, and the M/P ratio and RID were calculated. The condition of the mother and baby at 1 month after delivery was determined from the clinical information. The target benzodiazepines were alprazolam, brotizolam, clonazepam, clotiazepam, etizolam, ethyl loflazepate, flunitrazepam, and lorazepam.
Results:
For all drugs, the M/P ratios were <1 and remained constant over time. For drugs other than ethyl loflazepate, the RID values were <10%, which are considered safe; however, even with ethyl loflazepate, it was only slightly >10%. No abnormalities were found in breastfeeding infants whose mothers were receiving these medications.
Conclusions:
The RID results of this study suggest that drug exposure through breast milk is small; thus, maternal drug treatment and breastfeeding are compatible.

Pharmacological Inhibition of MCT4 Reduces 4-Hydroxytamoxifen Sensitivity by Increasing HIF-1α Protein Expression in ER-Positive MCF-7 Breast Cancer Cells.
Takanobu Nadai, Katsuya Narumi, Ayako Furugen, Yoshitaka Saito, Ken Iseki, Masaki Kobayashi
Biological & pharmaceutical bulletin, 44, 9, 1247, 1253, 2021年, ［査読有り］, ［最終著者, 責任著者］, ［国内誌］
英語, 研究論文（学術雑誌）, The rate of glycolysis in cancer cells is higher than that of normal cells owing to high energy demands, which results in the production of excess lactate. Monocarboxylate transporters (MCTs), especially MCT1 and MCT4, play a critical role in maintaining an appropriate pH environment through lactate transport, and their high expression is associated with poor prognosis in breast cancer. Thus, we hypothesized that inhibition of MCTs is a promising therapeutic target for adjuvant breast cancer treatment. We investigated the effect of MCT inhibition in combination with 4-hydroxytamoxifen (4-OHT), an active metabolite of tamoxifen, using two estrogen receptor (ER)-positive breast cancer cell lines, MCF-7 and T47D. Lactate transport was investigated in cellular uptake studies. The cytotoxicity of 4-OHT was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In both cell lines evaluated, MCT1 and MCT4 were constitutively expressed at the mRNA and protein levels. [14C]-L-lactate uptake by both cells was significantly inhibited by bindarit, a selective MCT4 inhibitor, but weakly affected by 5-oxoploline (5-OP), a selective MCT1 inhibitor. The results of the MTT assay showed that combination with bindarit, but not 5-OP, decreased 4-OHT sensitivity. Bindarit significantly increased the levels of hypoxia-inducible factor-1α (HIF-1α) in MCF-7 cells. Moreover, HIF-1α knockdown significantly increased 4-OHT sensitivity, whereas induction of HIF-1α by hypoxia decreased 4-OHT sensitivity in MCF-7 cells. In conclusion, pharmacological MCT4 inhibition confers resistance to 4-OHT rather than sensitivity, by increasing HIF-1α protein levels. In addition, HIF-1α inhibition represents a potential therapeutic strategy for enhancing 4-OHT sensitivity.

Analysis of α-Defensin 5 Secretion in Differentiated Caco-2 Cells: Comparison of Cell Bank Origin.
Genki Yasuda, Masaki Kobayashi, Atsuhito Kubota, Katsuya Narumi, Ayako Furugen, Yoshitaka Saito, Takashi Satoh, Natsuko Suzuki, Ken Iseki
Biological & pharmaceutical bulletin, 44, 2, 275, 278, 2021年, ［査読有り］, ［責任著者］, ［国内誌］
英語, 研究論文（学術雑誌）, α-Defensin 5 has a particularly broad antibacterial spectrum; it eliminates pathogenic microorganisms and regulates intestinal flora. Although Caco-2 cells are similar to small intestinal cells, it is unclear whether they secrete α-defensin 5. Therefore, we investigated whether Caco-2 cells secrete α-defensin 5 and determined the secretion mechanism using cells from three cell banks (ATCC, DSMZ, and RIKEN). The Caco-2 cell proliferation rate increased with the number of culture days, irrespective of cell bank origin. On the other hand, the alkaline phosphatase activity, which affects cell differentiation and the mRNA levels of several cytokines, such as interleukin 8 (IL-8), IL-6, IL-1β, tumor necrosis factor-α (TNF-α), and IL-2, in the Caco-2 cells fluctuated with the number of culture days, and differed for each cell bank. α-Defensin 5 secretion was detected in all three cell bank Caco-2 cells; particularly, the ATCC Caco-2 cells grew linearly depending on the cell culture day as well as the levels of IL-8 and TNF-α mRNA. This suggested that α-defensin 5 secretion in the ATCC Caco-2 cells was associated with fluctuations in the mRNA levels of various cytokines, such as IL-8 and TNF-α. In conclusion, Caco-2 cells may be a simple model for screening health food components and drugs that affect α-defensin 5 secretion.

Detection of risk factors related to administration suspension and severe neutropenia in gemcitabine and nab-paclitaxel treatment.
Yoshitaka Saito, Yoh Takekuma, Masaki Kobayashi, Yoshito Komatsu, Mitsuru Sugawara
Supportive care in cancer : official journal of the Multinational Association of Supportive Care in Cancer, 29, 6, 3277, 3285, 2020年10月26日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: The combination of gemcitabine (GEM) and nanoparticle albumin-bound paclitaxel (nab-PTX) is an effective chemotherapeutic regimen for locally advanced and metastatic pancreatic cancer. The dose-limiting toxicities (DLTs) of this treatment are sepsis and neutropenia, while the relative dose intensity (RDI) of GEM is approximately 75% and of nab-PTX is 70-80%. In this study, we evaluated the risk factor(s) regarding treatment suspension, which leads to reduction in the RDI of these agents, enabling appropriate schedule management. METHODS: Two hundred patients with pancreatic cancer who received GEM + nab-PTX were retrospectively investigated. Frequency and risk factor(s) of suspension of the treatment and grade 3/4 neutropenia in the first course were evaluated. RESULTS: The frequency of treatment suspension in the first course was 61%. The frequency of grade 3/4 neutropenia was 51%, while that of thrombocytopenia was 7.5%. The RDI was 78.0% for GEM and 77.7% for nab-PTX. Univariate and multivariate analyses to identify risk or preventive factors related to treatment suspension suggested that low platelet count at baseline was a risk factor, whereas dose reduction from the treatment initiation was a preventive factor. The most common cause of abeyance was grade 3/4 neutropenia (83.6%), the risk factors of which were low platelet count and age ≥ 65 years at baseline, while dose reduction was a preventive factor. CONCLUSION: We found that a low platelet level at baseline was a risk factor, whereas dose reduction from initiation was a preventive factor in regard to treatment suspension and severe neutropenia occurrence in GEM + nab-PTX treatment.

Identification of the essential extracellular aspartic acids conserved in human monocarboxylate transporters 1, 2, and 4.
Atsushi Yamaguchi, Katsuya Narumi, Ayako Furugen, Ken Iseki, Masaki Kobayashi
Biochemical and biophysical research communications, 529, 4, 1061, 1065, 2020年09月03日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporters (hMCTs) 1-4 transport monocarboxylates, such as l-lactate and pyruvate, as well as H+ across the plasma membrane. hMCT1, 2, and 4 play important roles in energy balance, pH homeostasis. However, the molecular mechanism of these transporters, especially their pH dependency, remains unknown. The aim of this study was to identify the residues involved in the pH dependence of hMCT1, 2, and 4. Firstly, we focused on the effects of extracellular acids of hMCT1. l-Lactate uptake assay and site-directed mutagenesis revealed that the aspartic acid of hMCT1 (hMCT1 D414) was an important residue conserved in MCT1, 2, and 4 (hMCT2 D398 and hMCT4 D379). Because the functional characteristic of hMCT2-mediated l-lactate transport has not been reported, we built a hMCT2-expressing system using Xenopus laevis oocytes. The transport activity of hMCT2 was enhanced by co-expression with embigin, an ancillary protein, and kinetic analysis of hMCT2-mediated l-lactate uptake revealed that the apparent Km value (0.32 ± 0.02 mM) was lower than that mediated by hMCT1 and 4. Finally, we investigated the conserved aspartic acids of hMCT2 and 4, and revealed that these residues were essential for l-lactate transport. These findings suggested that the extracellular aspartic acids conserved in hMCT1, 2, and 4 played important roles in transport activity and pH dependency, and can function as a first step of substrate and H+ recognition and transport from the extracellular to the intracellular region. These findings contributed to enhance our understanding of the transport process of hMCT1, 2, and 4.

Association between N-desmethylclozapine and clozapine-induced sialorrhea: Involvement of increased nocturnal salivary secretion via muscarinic receptors by N-desmethylclozapine.
Shuhei Ishikawa, Masaki Kobayashi, Naoki Hashimoto, Hideaki Mikami, Akihiko Tanimura, Katsuya Narumi, Ayako Furugen, Ichiro Kusumi, Ken Iseki
The Journal of pharmacology and experimental therapeutics, 375, 2, 376, 384, 2020年08月29日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Clozapine-induced sialorrhea (CIS) is a common side effect of clozapine. There is no established standard treatment for CIS since the underlying mechanism remains unknown. This study aimed to elucidate the mechanisms involved in CIS. In our clinical study, a prospective observational study evaluated the association between serum and saliva concentrations of clozapine or its metabolites and drooling severity and frequency scale (DSFS) score. In our in vivo study, we first developed a new CIS animal model; subsequently, we measured salivary secretion and concentrations of clozapine or its metabolites in the animal model. In our in vitro study, we measured the calcium ion (Ca2+) response to evaluate the effect of clozapine or its metabolites on human salivary gland cell line (HSY cells), and then examined whether their effect was inhibited by atropine. In our clinical study, serum and saliva N-desmethylclozapine concentrations were significantly correlated with nocturnal DSFS score. In our in vivo study, daily single oral administration of 100 mg/kg clozapine for 7 days significantly increased salivary secretion in rats. Furthermore, N-desmethylclozapine concentrations in serum and submandibular glands of rats were higher than clozapine concentrations. In our in vitro study, N-desmethylclozapine only elicited an increase in the intracellular Ca2+ in HSY cells. N-desmethylclozapine-induced Ca2+ responses were inhibited by atropine. These results suggest that N-desmethylclozapine is implicated in CIS by increasing nocturnal salivation via the muscarinic receptors. Moreover, our developed animal model that reflects CIS in clinical condition plays a key role as a bridge between basic and clinical research. Significance Statement Clozapine-induced sialorrhea (CIS) is a severe and frequent adverse reaction. Although the mechanism underlying CIS is less well understood. This paper reports that N-desmethylclozapine, a metabolite of clozapine, is implicated in CIS by increasing nocturnal salivation via the muscarinic receptors, and that oral administration of clozapine at 100 mg/kg once daily for 7 days to rat is the optimum method for establishing the new animal model reflecting the clinical scenario of CIS.

Different mechanisms of cisplatin resistance development in human lung cancer cells.
Keisuke Okamoto, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Ken Iseki, Masaki Kobayashi
Biochemical and biophysical research communications, 530, 4, 745, 750, 2020年08月08日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Cisplatin (CDDP) is a highly potent and important anticancer drug in lung cancer treatment. Long-term use of an anticancer agent causes resistance in cancer cells, and CDDP resistance involves multiple mechanisms. As the mechanism of resistance development differs depending on the cancer cell types, we aimed to evaluate the detailed mechanism of resistance to CDDP in two types of lung cancer cells: SBC-3 and A549 cells. The CDDP-resistant SBC-3/DDP and A549/DDP cells were established through continuous treatment with a gradually increasing dose of CDDP. The viability of SBC-3/DDP and A549/DDP cells treated with CDDP was 3.68 and 2.08 times higher than that of the respective parental cells. Moreover, SBC-3/DDP cells showed significantly increased cystine/glutamate transporter (xCT) mRNA level, and A549/DDP cells showed markedly increased sex determining region Y-box 2 (SOX2) mRNA level. Moreover, the uptake of cystine, a substrate of xCT, was higher in SBC-3/DDP cells than in SBC-3 cells, and cystine uptake in A549/DDP cells was not different from that in A549 cells. In addition, co-treatment with CDDP and sulfasalazine, an xCT inhibitor, showed lower the concentration of 50% inhibition for cell viability than CDDP alone in SBC-3 and SBC-3/DDP cells, but not in A549 and A549/DDP cells. Furthermore, SBC-3 cells transiently overexpressing xCT were resistant to CDDP, and xCT knockdown in A549/DDP cells did not significantly change the level of SOX2 mRNA and viability of cells upon CDDP treatment. In conclusion, the two lung cancer cell lines showed different mechanisms of resistance to CDDP.

Validation of the usefulness of artificial neural networks for risk prediction of adverse drug reactions used for individual patients in clinical practice
Shungo Imai, Yoh Takekuma, Hitoshi Kashiwagi, Takayuki Miyai, Masaki Kobayashi, Ken Iseki, Mitsuru Sugawara
PLOS ONE, 15, 7, e0236789, e0236789, Public Library of Science (PLoS), 2020年07月29日, ［査読有り］
研究論文（学術雑誌）

Comparison of the nephroprotective effects of non-steroidal anti-inflammatory drugs on cisplatin-induced nephrotoxicity in vitro and in vivo.
Keisuke Okamoto, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Ken Iseki, Masaki Kobayashi
European journal of pharmacology, 884, 173339, 173339, 2020年07月26日, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Cisplatin (CDDP) is an anticancer drug, often used in the treatment of several types of cancers. CDDP-induced nephrotoxicity (CIN) is one of the most severe adverse events associated with the use of CDDP. It has been suggested that the co-administration of non-steroidal anti-inflammatory drugs (NSAIDs) is a risk factor for CIN. However, the specific NSAIDs that affect CIN and the precise mechanisms underlying this interaction remain unclear. Hence, we aimed to evaluate the effect of NSAIDs on CDDP-induced cytotoxicity in vitro and confirmed the results in vivo. Using the epithelioid clone of the normal rat kidney cells (NRK-52E cells), we assessed the effects of 17 NSAIDs on CDDP-induced cytotoxicity all at once using the MTT assay. Furthermore, we evaluated two NSAIDs, which significantly attenuated or enhanced CDDP-induced cytotoxicity, in vivo. Wistar rats were treated with CDDP (5 mg/kg, i.p., day 1) and NSAIDs (p.o., day 1-4), and the kidneys were excised on day 5. Our results demonstrated that several NSAIDs attenuated, while others enhanced CDDP-induced cytotoxicity. Celecoxib significantly attenuated and flurbiprofen markedly enhanced cell dysfunction by CDDP. These results were reproduced in vivo as celecoxib decreased and flurbiprofen increased the expression of kidney injury molecule 1 (Kim-1) mRNA, a sensitive kidney injury marker, compared to the CDDP group. Moreover, celecoxib increased the antioxidant and autophagy markers quantified by qPCR in vitro and prevented a decrease in body weight induced by CDDP in vivo. In conclusion, we revealed that celecoxib significantly attenuated CIN in vitro and in vivo.

Evaluation of possible pharmacokinetic interaction between methotrexate and proton pump inhibitors in rats.
Hinata Ueda, Katsuya Narumi, Yu Sato, Ayako Furugen, Masaki Kobayashi, Ken Iseki
Pharmacological reports : PR, 72, 5, 1426, 1432, 2020年07月15日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: Methotrexate (MTX), an antifolate agent, is primarily eliminated by the kidney. Organic anion transporter 3 (OAT3) contributes to renal MTX clearance. Several studies have shown an association between co-administration of proton pump inhibitors (PPIs) and delayed elimination of MTX, but the findings are conflicting. In this study, we aimed to evaluate whether the differential inhibitory effects of PPIs on the OAT3-mediated transport of MTX are associated with the risks of delayed MTX elimination. METHODS: We investigated the effects of PPIs on rat (r) OAT3-mediated MTX uptake using HEK293T cells expressing rOAT3. To examine whether PPIs could affect the pharmacokinetics of MTX, changes in plasma concentration-time profiles were assessed when MTX (50 mg/kg, ip) and a range of PPIs (2 mg/kg, iv) were administered to rats. RESULTS: In vitro studies demonstrated that PPIs inhibited rOAT3-mediated uptake of MTX, with estimated IC50 values of 2.1-5.2 μM, and a rank order of esomeprazole ≈ lansoprazole ≈ omeprazole > rabeprazole. When MTX and esomeprazole were co-administered to rats, the plasma concentration of MTX 6 h after administration and the t1/2 were significantly higher than those in the vehicle group. The effect of lansoprazole was not significant, but showed a tendency to prolong plasma MTX levels. Famotidine, a histamine H2-receptor antagonist, showed a weak inhibitory effect on rOAT3-mediated MTX uptake, although it did not affect plasma concentration-time profile of MTX in vivo. CONCLUSION: Esomeprazole increases the t1/2 of MTX in rats, which may be partially attributed to the inhibition of rOAT3.

Effect of palonosetron and dexamethasone administration on the prevention of gastrointestinal symptoms in hepatic arterial chemoembolization with epirubicin.
Tatsuhiko Sakamoto, Yoshitaka Saito, Masaki Kobayashi, Takehiro Yamada, Yoh Takekuma, Masato Nakai, Koji Ogawa, Ken Iseki, Mitsuru Sugawara
Supportive care in cancer : official journal of the Multinational Association of Supportive Care in Cancer, 28, 7, 3251, 3257, 2020年07月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: There are several studies on premedication to prevent postembolization syndromes which occurs after transcatheter arterial chemoembolization (TACE), but the medication to be used is still not established. This study aimed to examine the effect of palonosetron and dexamethasone on the prevention of gastrointestinal symptoms induced by TACE. METHODS: Patients with hepatocellular carcinoma who were treated with TACE with epirubicin were retrospectively evaluated. The complete response rate of antiemetic drugs and incidence and severity of gastrointestinal symptoms were compared between the antiemetic group (AE group), which includes 51 patients prophylactically administered with palonosetron 0.75 mg and dexamethasone 9.9 mg intravenously before TACE on day 1 and dexamethasone 6.6 mg intravenously on days 2 and 3, and control group with 101 patients without antiemetic premedication. RESULTS: Complete response rate in the entire evaluation period was significantly higher in the AE group compared with that in the control group. In the acute phase, the incidence and severity of nausea, vomiting, and anorexia significantly decreased in the AE group, but only anorexia improved in the delay phase. Additionally, postembolization syndromes, such as abdominal pain and fever, were significantly attenuated in the AE group; however, constipation worsened in this group. CONCLUSIONS: Premedication of palonosetron and dexamethasone significantly prevents the incidence and reduces the severity of gastrointestinal symptoms especially in the acute phase. Further studies will be needed to determine the most recommended 5-HT3 antagonist or dosage of dexamethasone in establishing the optimal antiemetic regimen.

Evaluation of daptomycin-induced cellular membrane injury in skeletal muscle.
Takehiro Yamada, Shuhei Ishikawa, Nobuhisa Ishiguro, Masaki Kobayashi, Ken Iseki
Biological & pharmaceutical bulletin, 43, 9, 1338, 1345, 2020年06月23日, ［査読有り］, ［国内誌］
英語, 研究論文（学術雑誌）, Daptomycin, a cyclic lipopeptide antibiotic, has bactericidal activity against gram-positive organisms and is especially effective against methicillin-resistant Staphylococcus aureus. Although daptomycin causes unique adverse drug reactions such as elevation of creatine phosphokinase or rhabdomyolysis, the detailed mechanisms underlying these adverse drug reactions in skeletal muscle are unclear. This study aimed to elucidate whether daptomycin causes direct skeletal muscle cell toxicity and investigate the relationship between daptomycin exposure and musculoskeletal toxicity. First, we evaluated the relationship between daptomycin exposure and skeletal muscle toxicity. Of the 38 patients who received daptomycin intravenously, an elevation in creatine phosphokinase levels was observed in five. The median plasma trough concentration of daptomycin in patients with elevated creatine phosphokinase levels was significantly higher than that in patients whose creatine phosphokinase levels were within the normal range, suggesting that increased exposure to daptomycin is related to elevation in creatine phosphokinase levels. In an in vitro study using human rhabdomyosarcoma cells, daptomycin reduced cell viability and increased membrane damage. These effects were more marked under hypoxic conditions. A necroptotic pathway seemed to be involved because phosphorylated mixed lineage kinase domain-like protein expression was enhanced following daptomycin exposure, which was significantly enhanced under hypoxic conditions. These findings indicate that daptomycin elicits cytotoxic effects against skeletal muscle cells via the necroptotic pathway, and the extent of toxicity is enhanced under hypoxic conditions.

Cellular uptake properties of lamotrigine in human placental cell lines: Investigation of involvement of organic cation transporters (SLC22A1-5).
Nami Hasegawa, Ayako Furugen, Kanako Ono, Mai Koishikawa, Yuki Miyazawa, Ayako Nishimura, Takeshi Umazume, Katsuya Narumi, Masaki Kobayashi, Ken Iseki
Drug metabolism and pharmacokinetics, 35, 3, 266, 273, 2020年06月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Lamotrigine (LTG) is an important antiepileptic drug for the treatment of seizures in pregnant women with epilepsy. However, it is not known if the transport of LTG into placental cells occurs via a carrier-mediated pathway. The aim of this study was to investigate the uptake properties of LTG into placental cell lines (BeWo and JEG-3), and to determine the involvement of organic cation transporters (OCTs, SLC22A1-3) and organic cation/carnitine transporter (OCTNs, SLC22A4-5) in the uptake process. The uptake of LTG at 37 °C was higher than that at 4 °C. OCT1 and OCTNs were detected in both cell lines. The uptake of LTG was not greatly affected by the extracellular pH, Na+-free conditions, or the presence of l-carnitine, suggesting that OCTNs were not involved. Although several potent inhibitors of OCTs (chloroquine, imipramine, quinidine, and verapamil) inhibited LTG uptake, other typical inhibitors had no effect. In addition, siRNA targeted to OCT1 had no significant effect on LTG uptake. The mRNA expression in human term placenta followed the order OCTN2 > OCT3 > OCTN1 > OCT1 ≈ OCT2. These observations suggested that LTG uptake into placental cells was carrier-mediated, but that OCTs and OCTNs were not responsible for the placental transport process.

Effects of single and repetitive valproic acid administration on the gene expression of placental transporters in pregnant rats: An analysis by gestational period.
Naoko Jinno, Ayako Furugen, Yuko Kurosawa, Yuki Kanno, Katsuya Narumi, Masaki Kobayashi, Ken Iseki
Reproductive toxicology (Elmsford, N.Y.), 96, 47, 56, 2020年05月11日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, The use of valproic acid (VPA), an antiepileptic drug, during pregnancy, is known to increase various fetal risks. Since VPA has been known to inhibit histone deacetylases (HDACs); its administration could alter gene transcription levels. However, in vivo effects of VPA administration on placental transporters have not been fully elucidated. The purpose of the present study was to comprehensively evaluate the effects of single and repetitive VPA administration on the expression of placental transporters and analyze them by gestational day. We investigated 18 transporters (8 ATP-binding cassette (ABC) and 10 solute carrier (SLC) transporters) in the placentas of pregnant rats that were orally administered 400 mg/kg/day VPA for one or four days, during mid- or late gestation. In the control rats, 4 ABC transporter genes (Abcb1a, 1b, Abcc2, Abcc4) were upregulated, 3 (Abcc3, Abcc5, Abcg2) downregulated through gestation, whereas 1 (Abcc1) was not changed. Regarding SLC transporters, 6 genes (Slc7a5, Slc16a3, Slc22a3, Slc22a4, Slco2b1, Slco4a1) were increased, 1 (Slc29a1) decreased through gestation, whereas 3 (Slc7a8, Slc22a5, Slco2a1) showed no significant change. Single VPA administration altered the expression of 9 transporters and repetitive administration, 13 transporters. In particular, VPA remarkably decreased Abcc4 and Slc22a4 in late gestation and increased Abcc5 during mid-gestation. Our findings indicated that VPA administration changed transporter expression levels in rat placenta, and suggested that sensitivity to VPA differs across gestational stages.

Non-steroidal Anti-inflammatory Drugs Are a Risk Factor for Cisplatin-induced Nephrotoxicity: A Meta-analysis of Retrospective Studies.
Keisuke Okamoto, Yoshitaka Saito, Katsuya Narumi, Ayako Furugen, Ken Iseki, Masaki Kobayashi
Anticancer research, 40, 3, 1747, 1751, 2020年03月, ［査読有り］, ［最終著者, 責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND/AIM: Previous reports have demonstrated that non-steroidal anti-inflammatory drugs (NSAIDs) are a risk factor for cisplatin-induced nephrotoxicity (CIN). Here, the results of these previous studies were comprehensively assessed via a meta-analysis. MATERIALS AND METHODS: After a database search to select eligible studies, a meta-analysis was performed using a forest plot, followed by an assessment of the heterogeneity and publication bias and a subgroup analysis. RESULTS: Seven studies were extracted as candidates. All were retrospective studies and evaluated the effect of NSAIDs on CIN as a secondary endpoint. According to the meta-analysis, total odds ratio was 1.88 (95% confidence interval=1.44-2.45). Further, high heterogeneity and publication bias were not observed. A subgroup analysis of the chemotherapy evaluation period revealed that CIN tended to be enhanced in the first course group (evaluation in only 1 course) and was significantly enhanced in the total course group (evaluation in 1 or more courses) by NSAIDs co-administration. CONCLUSION: NSAIDs co-administration could be a risk factor for CIN.

Molecular characterization of the orphan transporter SLC16A9, an extracellular pH- and Na+-sensitive creatine transporter.
Yuya Futagi, Katsuya Narumi, Ayako Furugen, Masaki Kobayashi, Ken Iseki
Biochemical and biophysical research communications, 522, 2, 539, 544, 2020年02月05日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporters (hMCTs) mediate the transport of monocarboxylates across plasma membranes. One such transporter, hMCT9, has been shown to be related to serum uric acid levels and the risk of renal overload gout. However, the functional characteristics of hMCT9 remain unknown. The aim of this study was to investigate the expression and localization of hMCT9 using a Xenopus laevis oocyte heterologous expression system and characterize its transport properties. Kinetic analysis of hMCT9-mediated creatine uptake revealed that uptake consisted of two components, with apparent Km values of 237 mm (low-affinity) and 23.7 mm (high-affinity), respectively. The transport activity of hMCT9 was dependent on the extracellular pH and activity sharply increased with increasing pH. Under Na+-free conditions, hMCT9-mediated creatine uptake was reduced by one-half, indicating that hMCT9 is a Na+-sensitive transporter. Moreover, carbonyl cyanide 3-chlorophenylhydrazone (a protonophore) inhibited hMCT9 activity, whereas valinomycin (a K+-ionophore) did not inhibit the transporter. These results suggest that hMCT9 is susceptible to changes in H+ gradients. A cis-inhibition assay of hMCT9-and hMCT12-mediated creatine transport revealed that cyclocreatine, creatine, guanidineacetate, and 3-guanidinopropionate are recognized by the transporter, and 4-guanidinobutyrate and guanidinoethyl sulfonate selectively inhibited hMCT9 activity. These findings demonstrate that hMCT9 is an extracellular pH- and Na+-sensitive creatine transporter.

Relationships between plasma lactate, plasma alanine, genetic variations in lactate transporters and type 2 diabetes in the Japanese population.
Issei Higuchi, Yuki Kimura, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Hideaki Miyoshi, Akinobu Nakamura, Takehiro Yamada, Tatsuya Atsumi, Ken Iseki
Drug metabolism and pharmacokinetics, 35, 1, 131, 138, 2020年02月, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, The present study aimed to characterize the relationships between plasma lactate, plasma alanine, monocarboxylate transporter (MCT) polymorphisms, and indices of diabetes in patients with type 2 diabetes (T2D) in Japan. Eighty-three patients with T2D were prospectively enrolled. The gluconeogenesis and glycogenolysis are enhanced and uptake of glucose is decreased in the T2D liver. Since the liver plays an important role in maintaining glucose metabolism, we examined the relationships between liver enzymes and indices of diabetes. Some studies have reported that MCT1 (SLC16A1) polymorphism causes metabolic diseases. In addition, a high frequency of MCT1 polymorphism was reported in a healthy Japanese population. However, little is known about the relationships between T2D and MCT polymorphisms. Plasma l-lactate concentration positively correlated with indices of diabetes (fasting plasma glucose [FPG] and hemoglobin A1c [HbA1c]) and with the liver enzymes alanine aminotransferase (ALT) and gamma-glutamyl transpeptidase (γ-GTP). MCT1 polymorphisms were associated with all of these markers. We identified no significant correlations between d-lactate or alanine concentrations and any of these markers, but a significant association was observed between l-lactate, a marker of oxidative capacity, and indices of diabetes. We conclude that plasma l-lactate concentration may represent a predictor of the progression or severity of T2D.

Extracellular lysine 38 plays a crucial role in pH-dependent transport via human monocarboxylate transporter 1.
Atsushi Yamaguchi, Yuya Futagi, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Biochimica et biophysica acta. Biomembranes, 1862, 2, 183068, 183068, 2020年02月01日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporters (hMCTs) are expressed in many tissues and mediate the transport of various substrates across the plasma membrane. Among hMCTs, hMCT1-4 cotransport H+ with monocarboxylates such as pyruvate and l-lactate, implying that these proteins recognize both substrate and H+. However, the mechanism of translocation, and particularly that of hMCT1 pH-dependent transport, remains largely unknown. This study aimed at identifying residues involved in the pH dependence of hMCT1 using a combination of amino acid-modifying reagents, site-directed mutagenesis in a Xenopus laevis oocyte expression system, and homology modeling. We showed that diethyl pyrocarbonate (DEPC), phenylglyoxal (PGO), and 4,4'-diisothiocyanato-2,2'-stilbenedisulfonic acid disodium salt (DIDS), which react with histidine, arginine, and lysine residues respectively, all inhibited hMCT1 activity. Since DEPC, PGO, and DIDS are membrane impermeable reagents, we mutated to other residues individual histidine, arginine, and lysine residues located within the extracellular regions of hMCT1. Analyses of these mutants demonstrated that except for K38, the extracellular basic residues of hMCT1 were not involved in its transport activity and pH dependence. Moreover, analyses of various mutants in which K38 was substituted for another residue and of an hMCT1 homology model focusing on the location of K38 in the three-dimensional structure delineated the mechanism of hMCT1 pH dependence. Collectively, our data indicate that K38 plays an essential role in hMCT1 transport activity. We would like to propose a mechanism whereby K38 is positioned within a hydrophobic and narrow cavity that is part of the transport pathway, and regulates pH-dependent gating of hMCT1.

Efficacy of additional dexamethasone administration for the attenuation of paclitaxel-associated acute pain syndrome.
Yoshitaka Saito, Masaki Kobayashi, Takehiro Yamada, Jun Sakakibara-Konishi, Naofumi Shinagawa, Ichiro Kinoshita, Hirotoshi Dosaka-Akita, Ken Iseki
Supportive care in cancer : official journal of the Multinational Association of Supportive Care in Cancer, 28, 1, 221, 227, 2020年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: Paclitaxel-associated acute pain syndrome (P-APS) affects 80% of patients undergoing therapy. Although it has been shown that prednisone administration for 5 days relieves P-APS, detailed results have not been reported thus far. Therefore, in this study, we evaluated the preventive effect of dexamethasone (DEX) administration against P-APS. METHODS: A total of 60 patients who received carboplatin (area under the curve; AUC = 5-6) plus paclitaxel (200 mg/m2) (plus bevacizumab 15 mg/kg, if non-squamous carcinoma of lung) were enrolled. Eight milligrams of DEX was orally administered on days 2 and 3 to the DEX group patients, and the frequency, severity, duration of P-APS, and other adverse effects in the first cycle were retrospectively evaluated and compared to those observed in control group patients, who were not administered DEX on days 2 and 3. RESULTS: No difference in terms of patient characteristics, except for type of cancer, was observed between groups. The results showed that the frequency of all grade P-APS was approximately 70% and there was no difference between groups. Frequency of ≥ G2 P-APS was 40% in the control group and 14% in the DEX group, demonstrating a significant reduction. Duration of P-APS was 5.8 days in the control group and 4.3 days in the DEX group, which tended to become shorter following additional DEX administration, although this was not significant. Adverse effects other than P-APS induced by chemotherapy were similar between the two groups. CONCLUSION: Additional DEX administration is safe and useful for the attenuation of the severity of P-APS.

Homology modeling and site-directed mutagenesis identify amino acid residues underlying the substrate selection mechanism of human monocarboxylate transporters 1 (hMCT1) and 4 (hMCT4).
Yuya Futagi, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Cellular and molecular life sciences : CMLS, 76, 24, 4905, 4921, 2019年12月, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporters (hMCTs/SLC16As) mediate the transport of monocarboxylic compounds across plasma membranes. Among the hMCTs, hMCT1 and hMCT4 are expressed in various tissues, and transport substrates involved in energy metabolism. Both transporters mediate L-lactate transport, but, although hMCT1 also transports L-5-oxoproline (L-OPro), this compound is minimally transported by hMCT4. Thus, we were interested in the molecular mechanism responsible for the difference in substrate specificity between hMCT1 and hMCT4. Therefore, we generated 3D structure models of hMCT1 and hMCT4 to identify amino acid residues involved in the substrate specificity of these transporters. We found that the substrate specificity of hMCT1 was regulated by residues involved in turnover number (M69) and substrate affinity (F367), and these residues were responsible for recognizing (directly or indirectly) the -NH- moiety of L-OPro. Furthermore, our homology model of hMCT1 predicted that M69 and F367 participate in hydrophobic interactions with another region of hMCT1, emphasizing its potentially important role in the binding and translocation cycle of L-OPro. Mutagenesis experiments supported this model, showing that efficient L-OPro transport required a hydrophobic, long linear structure at position 69 and a hydrophobic, γ-branched structure at position 367. Our work demonstrated that the amino acid residues, M69 and F367, are key molecular elements for the transport of L-OPro by hMCT1. These two residues may be involved in substrate recognition and/or substrate-induced conformational changes.

Construction of a flow chart-like risk prediction model of ganciclovir-induced neutropaenia including severity grade: A data mining approach using decision tree.
Shungo Imai, Takehiro Yamada, Kumiko Kasashi, Nobuhisa Ishiguro, Masaki Kobayashi, Ken Iseki
Journal of clinical pharmacy and therapeutics, 44, 5, 726, 734, 2019年10月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, WHAT IS KNOWN AND OBJECTIVE: Haematological toxicities such as neutropaenia are a common side effect of ganciclovir (GCV); however, risk factors for GCV-induced neutropaenia have not been well established. Decision tree (DT) analysis is a typical technique of data mining consisting of a flow chart-like framework that shows various outcomes from a series of decisions. By following the flow chart, users can estimate combinations of risk factors that may increase the probability of certain events. In our previous study, we demonstrated the usefulness of this approach in the evaluation of adverse drug reactions. Therefore, we aimed to construct a risk prediction model of GCV-induced neutropaenia including severity grade. METHODS: We performed a retrospective study at the Hokkaido University Hospital and enrolled patients who received GCV between April 2008 and March 2018. Neutropaenia was defined as an absolute neutrophil count (ANC) <1500 cells/mm3 and a decrease to <75% relative to baseline. We classified the patients who developed neutropaenia in three groups (Grades 2-4) based on the National Cancer Institute-Common Terminology Criteria for Adverse Events. Data collection was achieved through the retrieval of medical records. We employed a chi-squared automatic interaction detection algorithm to construct the DT model and compared the accuracies to the logistic regression model (a conventional statistical method) to evaluate the established model. RESULTS AND DISCUSSION: In total, 396 adult patients were included in the study; 61 (15.4%) developed neutropaenia. Three predictive factors (hematopoietic stem cell transplantation, baseline ANC <3854 cells/mm3 and duration of therapy ≥15 days) were extracted using the DT analysis to produce five subgroups, the incidence of neutropaenia ranged between 1.7% and 52.8%. In each subgroup, patients who developed neutropaenia were categorized based on the severity. The accuracies of each model were the same (84.6%), which indicated precision. WHAT IS NEW AND CONCLUSION: We successfully built a risk prediction model of GCV-induced neutropaenia including severity grade. This model is expected to assist decision-making in the clinical setting.

Black tea extract and theaflavin derivatives affect the pharmacokinetics of rosuvastatin by modulating organic anion transporting polypeptide (OATP) 2B1 activity.
Ayuko Kondo, Katsuya Narumi, Keisuke Okuhara, Yuka Takahashi, Ayako Furugen, Masaki Kobayashi, Ken Iseki
Biopharmaceutics & drug disposition, 40, 8, 302, 306, 2019年09月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Theaflavins (TFs) are derived from black tea, an important source of dietary polyphenols. Although the potential interactions between dietary polyphenols and drugs have been demonstrated through in vitro and in vivo studies, little information is available concerning the influence of TFs on drug disposition. Organic anion transporting polypeptide 2B1 (OATP2B1) is expressed in human enterocytes and plays a role in the intestinal absorption of numerous drugs. The current study evaluated the effects of black tea extracts on the pharmacokinetics of rosuvastatin in rats, and investigated the effect of four major TFs (theaflavin, theaflavin-3-gallate, theaflavin-3'-gallate and theaflavin-3,3'-digallate) on the transport activity of OATP2B1. Black tea extracts significantly decreased the maximum plasma concentration (Cmax ) and area under the plasma concentration-time curve (AUC0 -8 ) of rosuvastatin by 48% and 37%, respectively (p < 0.001 and p < 0.01, respectively). Moreover, OATP2B1-mediated rosuvastatin and estrone-3-sulfate uptake was significantly reduced in the presence of TFs. A kinetic study revealed that the uptake efficiency (in terms of Vmax /Km ) of rosuvastatin was decreased following treatment with TFs. Black tea extracts also reduced OATP2B1-mediated rosuvastatin uptake. These results suggest that black tea reduces the plasma concentrations of rosuvastatin by inhibiting the intestinal OATP2B1-mediated transport of rosuvastatin.

リネゾリド投与患者における嘔吐発現の因果関係の検証
堤 竹蔵, 今井 俊吾, 山田 勝久, 山田 武宏, 笠師 久美子, 小林 正紀, 井関 健
薬学雑誌, 139, 7, 1055, 1061, 公益社団法人 日本薬学会, 2019年07月01日, ［査読有り］
日本語, Linezolid (LZD), an antimicrobial agent against methicillin-resistant Staphylococcus aureus, demonstrates good bone and joint penetration, and is used for prosthetic bone and joint infections. Recently, we observed vomiting in several patients administered LZD. However, there are few reports on the incidence rate of, and risk factors for, LZD-induced nausea and vomiting. In this study, we aimed to verify the relationship between LZD administration and vomiting. Patients administered LZD at the Department of Orthopedic Surgery of Hokkaido University Hospital between November 2008 and December 2017 were enrolled in the study. The primary endpoint was the comparison of the vomiting rate between patients administered LZD (LZD group) and those administered other antibiotics (non-LZD group). For the secondary endpoint, to verify the risk factors of vomiting, a univariate logistic regression analysis was performed. In total, 130 patients were included in this study; 77 patients in the LZD group, and 53 in the non-LZD group. Vomiting occurred in 18 patients in the LZD group and 4 patients in the non-LZD group (23.4% and 7.5%, respectively); this was significantly higher in the LZD group. In the univariate logistic regression analysis, LZD administration, gender (female), age ≥65 years, renal impairment (creatinine clearance <60 mL/min) and concomitant use of rifampicin were extracted as potential risk factors of vomiting. The results of this study reveal a possible relationship between LZD administration and vomiting.

Quantification of eight benzodiazepines in human breastmilk and plasma by liquid-liquid extraction and liquid-chromatography tandem mass spectrometry: Application to evaluation of alprazolam transfer into breastmilk.
Ayako Furugen, Ayako Nishimura, Masaki Kobayashi, Takeshi Umazume, Katsuya Narumi, Ken Iseki
Journal of pharmaceutical and biomedical analysis, 168, 83, 93, 2019年05月10日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Breastfeeding is strongly encouraged for infant and maternal health. Benzodiazepines (BZDs) are widely prescribed drugs for symptoms, such as anxiety and insomnia, which many women could experience during the postpartum period. However, limited information is currently available to evaluate the transfer of different BZDs into breastmilk. In order to assess the proprieties of this medication during breastfeeding, robust and sensitive analytical methods to quantify BZDs are required. For this purpose, we developed a method for quantification of BZDs, including alprazolam, bromazepam, clonazepam, clotiazepam, etizolam, flunitrazepam, lorazepam, and CM7116 (a metabolite of ethyl loflazepate), in human breastmilk and plasma using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Sample preparation was performed by a simple liquid-liquid extraction (LLE) with ethyl acetate. For sample preparation of CM7116, the pretreatment process to completely obtain the metabolite was added before the LLE step. The BZDs were separated by a C18 column using a gradient elution of acetonitrile in aqueous ammonium acetate solution, and were detected in the positive ion electrospray mode with multiple reaction monitoring (MRM). Lower limits of quantification (LLOQs) in breastmilk ranged from 0.25 to 0.5 ng/mL, and those in plasma ranged from 0.5 to 1.0 ng/mL. The intra-day and inter-day precision, and accuracy of data were assessed and found to be acceptable. The developed method was successfully applied to measure the concentration of alprazolam in breastmilk and plasma, which were donated by a lactating woman who had been regularly treated with alprazolam. Milk to plasma (M/P) ratios were calculated as 0.52 (before oral administration) and 0.49 (2 h after administration) 3 days after delivery. The M/P ratio 1 month after delivery was calculated as 0.41 (2 h after administration). We estimated that the relative infant dose (RID) values of alprazolam ranged from 3.11 to 4.61%.

下顎埋伏智歯抜歯術におけるセフカペンピボキシルとアモキシシリンの手術部位感染予防効果の比較　　　　　　　　　　　　　　　
山神 彰, 山田 武宏, 北川 善政, 大廣 洋一, 佐藤 淳, 石黒 信久, 今井 俊吾, 小林 正紀, 井関 健
医療薬学, 45, 5, 254, 261, (一社)日本医療薬学会, 2019年05月, ［査読有り］
日本語, 下顎埋伏智歯抜歯術においてセフカペンピボキシル(CFPN-PI)またはアモキシシリン(AMPC)の予防投与を受けた患者の手術部位感染(SSI)発生率を比較した。解析対象となったのはCFPN-PI群129例(男性50例、女性79例、中央値24.0歳)、AMPC群164例(男性75例、女性89例、中央値25.0歳)であった。SSI発生率はCFPN-PI群において11.6%(15/129)、AMPC群において2.4%(4/164)であり、CFPN-PI群のSSI発生率は有意に高かった。次いで、SSI発生に影響する要因分析を行った。単変量解析において抗菌薬(CFPN-PIの使用)、性別(女性)、術後入院、手術難易度、反対側下顎埋伏智歯抜歯の有無、Pell-Gregory分類がP≦0.2となり、これらを多変量解析における解析対象因子として抽出した。解析対象因子のvariance inflation factorはいずれも4未満であり多重共線性はみられなかった。多変量解析の結果、抗菌薬(CFPN-PIの使用)および術後入院がそれぞれオッズ比5.61、4.66を示し、SSIに影響する独立因子として抽出された。

Construction of a risk prediction model of vancomycin-associated nephrotoxicity to be used at the time of initial therapeutic drug monitoring: A data mining analysis using a decision tree model.
Shungo Imai, Takehiro Yamada, Kumiko Kasashi, Yusuke Niinuma, Masaki Kobayashi, Ken Iseki
Journal of evaluation in clinical practice, 25, 1, 163, 170, 2019年02月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, OBJECTIVES: In our previous study, we built a risk prediction model of vancomycin (VCM)-associated nephrotoxicity using decision tree (DT) analysis. However, this has several limitations in clinical applications. Our objective here was to construct a clinically applicable risk prediction model to be used at the time of initial therapeutic drug monitoring (TDM), in patients with uncomplicated infections. METHOD: A retrospective study was conducted at Hokkaido University Hospital. Subjects that had received VCM were extracted between November 2011 and April 2017. Nephrotoxicity was defined as an increase in serum creatinine of 0.5 mg/dL or 50% or higher from baseline. The additional inclusion criteria in this study were as follows: (1) the target trough level of VCM was set to 10 to 15 mg/L, and (2) the duration of therapy was 7 to 14 days. Patients were assumed to have uncomplicated infections. Risk factors for nephrotoxicity were evaluated, which could be extracted at the initial TDM. In the DT analysis, a chi-squared automatic interaction detection algorithm was constructed. RESULTS: A total of 402 patients were enrolled, and 56 (13.9%) patients developed nephrotoxicity. In the DT analysis, concomitant medications (furosemide, piperacillin-tazobactam, and vasopressor drugs) and an initial VCM trough concentration ≥ 15.0 mg/L were extracted as predictive variables by which patients were divided into six subgroups. The incidence of nephrotoxicity was 5.2% to 70.0%, with subgroups classified as low to high risk of nephrotoxicity. The accuracy of DT model was favourable (87.1%). CONCLUSION: We propose that the DT model built in this study is applicable to clinical practice.

Mutual role of ecto-5'-nucleotidase/CD73 and concentrative nucleoside transporter 3 in the intestinal uptake of dAMP.
Katsuya Narumi, Tsukika Ohata, Yuichi Horiuchi, Hiroshi Satoh, Ayako Furugen, Masaki Kobayashi, Ken Iseki
PloS one, 14, 10, e0223892, 2019年, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, 2'-Deoxyadenosine 5'-monophosphate (dAMP), a deoxyribonucleotide found in DNA, affects intestinal cell growth. The molecular mechanisms underlying gastrointestinal absorption of foreign DNA ingested along with food has hardly been investigated. The aim of this study was to investigate the mechanism underlying intestinal absorption of dAMP. The uptake of [3H]dAMP by Caco-2 cells was Na+- and pH-dependent and was inhibited by various nucleosides. In contrast, nitrobenzylthioinosine (NMBPR), an equilibrative nucleoside transporter inhibitor, showed little inhibitory effects on [3H]dAMP uptake. Additionally, human concentrative nucleoside transporter (CNT) 3, transiently expressed in COS-7 cells, mediated the uptake of [3H]dAMP. A kinetic study revealed that the Km value of CNT3-mediated uptake of dAMP (59.6 μM) was close to that of 2'-deoxyadenosine (dAdo) (56.3 μM), whereas the dAMP Vmax (15.6 pmol·mg protein-1min-1) was 500-fold lesser than the dAdo Vmax (7782 pmol·mg protein-1min-1). Further, [3H]dAMP uptake was greater in COS-7 cells expressing ecto-5'-nucleotidase/CD73 with CNT3 than in those expressing CNT3 alone. These data suggest that, although dAMP is a substrate of CNT3, it is dephosphorylated to dAdo by CD73 and is efficiently absorbed as dAdo from the intestinal lumen.

[Paclitaxel-associated Acute Pain Syndrome Similarly Occurs in the Patients with or without Previously Administered Non-steroidal Anti-inflammatory Drugs Prior to Paclitaxel Administration].
Saito Y, Yamada T, Kobayashi M, Sakakibara-Konishi J, Shinagawa N, Kinoshita I, Dosaka-Akita H, Iseki K
Yakugaku zasshi : Journal of the Pharmaceutical Society of Japan, 139, 12, 1601, 1608, 2019年, ［査読有り］

Analysis of the effects of polyunsaturated fatty acids on transporter expressions using a PCR array: Induction of xCT/SLC7A11 in human placental BeWo cells.
Kanako Ono, Ayako Furugen, Yuko Kurosawa, Naoko Jinno, Katsuya Narumi, Masaki Kobayashi, Ken Iseki
Placenta, 75, 34, 41, 2019年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, OBJECTIVE: Polyunsaturated fatty acids (PUFAs), including arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), are essential for adequate fetal growth. The aim of the present study was to elucidate the effects of PUFAs on the expression and function of placental transporters, which play important roles in placental functions including the supply of nutrients to the fetus, excretion of metabolites, and protection of the fetus from xenobiotics. METHODS: Human placental choriocarcinoma BeWo cells were used as a trophoblast model. PUFA-induced alteration in the gene expression of 84 transporters was investigated by a commercially available PCR array. Protein levels and the activity of transporters were assessed by western blotting and uptake experiments, respectively. The placental expression of the transporters was analyzed using pregnant Wistar rats. RESULTS: PUFAs (AA, EPA, and DHA) increased cystine/glutamate transporter xCT/SLC7A11, which mediates the cellular uptake of cystine coupled with the efflux of glutamate in human placental choriocarcinoma BeWo cells. These PUFAs also increased [14C]-cystine uptake in BeWo cells. PUFA-induced xCT/SLC7A11 mRNA expression was not blocked by nuclear factor-erythroid 2-related factor-2 (NRF2) knockdown. Reverse transcription (RT)-PCR analysis indicated that xCT/Slc7a11 mRNA was detected in rat placenta and the expression level at gestational day (GD) 12 was higher than that at GD 20. CONCLUSION: These results indicate that PUFAs promoted cystine uptake in placental cells by inducing xCT/SLC7A11 expression and NRF2 did not contribute to upregulation of xCT/SLC7A11 by PUFAs. Furthermore, xCT expression in rat placenta may change during pregnancy.

薬学実務実習前後における薬学生のコミュニケーション分析～Roter method of interaction process analysis（RIAS）を用いて～
武隈 洋, 森 綾子, 小林 正紀, 山田 勇磨, 佐藤 夕紀, 鳴海 克哉, 古堅 彩子, 菅原 満
薬学雑誌, 138, 12, 1579, 1586, 公益社団法人 日本薬学会, 2018年12月01日
日本語, Communication education is now necessary for pharmaceutical education since the role of pharmacists has expanded from “medicine-based” to “person-based”. However, a standard for assessing the effectiveness of a communication education program has not been established. Hence, the aim of this study was to determine the effectiveness of clinical training in pharmacy for enhancing the ability of pharmacy students to communicate. Role playing with simulated patients was performed by pharmacy students before and after clinical practice for pharmacy, and the effects of learning were analyzed by Roter method of interaction process analysis (RIAS). Analysis by RIAS enabled quantification and objective evaluation of communication by pharmacy students. The results showed improvement of interactive communication, decrease of “Question asking” and “Others” including “Transition words”, and increase of “Partnership behaviors” and “Counsel behaviors”. The pharmacy students became skillful in communication without showing hesitation. The results therefore showed that clinical training contributes to improvement in the ability of pharmacy students to communicate.

Valproic acid transport in the choriocarcinoma placenta cell line JEG-3 proceeds independently of the proton-dependent transporters MCT1 and MCT4.
Yuri Ishiguro, Ayako Furugen, Katsuya Narumi, Ayako Nishimura, Takeshi Hirano, Masaki Kobayashi, Ken Iseki
Drug metabolism and pharmacokinetics, 33, 6, 270, 274, 2018年12月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Medication therapy is the first line of treatment in the management of epilepsy. Fetal exposure to valproic acid (VPA), an antiepileptic drug, poses an elevated risk of teratogenicity in early pregnancy. Some studies have reported that monocarboxylate transporters (MCTs) may be involved in the placental transport of VPA. However, it has not been determined which MCTs contribute to VPA transport into the placenta. Therefore, the aim of this study was to determine how MCTs contribute to VPA transport into the placenta using the human placenta choriocarcinoma cell line JEG-3. VPA uptake was investigated using JEG-3 cells and radiolabeled VPA. MCT expression in JEG-3 cells was detected using RT-PCR and western blotting. Knockdown of MCTs was carried out using siRNAs. VPA uptake into JEG-3 cells was pH- and concentration-dependent, and described by using the Michaelis-Menten equation (Km = 0.95 ± 0.17 mM; Vmax = 19.3 ± 1.21 nmol/mg protein/15 s). MCT1 and MCT4 expression was found in JEG-3 cells, and typical MCT inhibitors significantly inhibited VPA uptake into JEG-3 cells. However, knockdown of MCT1 and MCT4 did not alter VPA uptake. In conclusion, VPA transport is mediated by a proton-dependent transporter in JEG-3 cells, but not by MCT1 and MCT4.

Valproate sensitizes human glioblastoma cells to 3-bromopyruvate-induced cytotoxicity.
Yuri Ishiguro, Masaki Kobayashi, Masaya Ideno, Katsuya Narumi, Ayako Furugen, Ken Iseki
International journal of pharmaceutics, 551, 1-2, 97, 102, 2018年11月15日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Glioblastoma (GBM) is the most common brain tumor; however, no effective treatment for it is available yet. Monocarboxylate transporters, which are highly expressed in GBM, play a role in transporting antitumor agents, such as 3-bromopyruvate (3-BrPA). Valproate, primarily used to treat epilepsy, has been considered a possible treatment option for malignant GBM. In this study, we aimed to investigate the combined effects of 3-BrPA and valproate on GBM cell growth and elucidate the underlying mechanisms. Valproate enhanced 3-BrPA-induced cell death in T98G cells, used as a GBM model. Multidrug resistance-associated protein 2 (MRP2) and breast cancer resistance protein (BCRP) mRNA levels significantly increased after valproate treatment. 3-BrPA-induced cell death, which was enhanced by valproate, was inhibited in the presence of MK571, a MRP inhibitor, or Ko143, a BCRP inhibitor. In addition, treatment with 3-BrPA and valproate for 48 h reduced cellular ATP levels compared to those in the 3-BrPA alone treatment group. However, cellular ATP levels were recovered in the presence of MK571 or Ko143, compared to those in the 3-BrPA and valproate treatment groups. In conclusion, we suggested that valproate enhanced 3-BrPA-induced cell death. This might be attributable to the increase in cellular ATP consumption owing to valproate-induced MRP2 or BCRP expression.

外来化学療法におけるがん専門薬剤師による長期継続的な薬学的患者管理の有用性の検討
齋藤 佳敬, 原田 幸子, 小林 正紀, 植田 孝介, 山﨑 浩二郎, 熊井 正貴, 宮本 剛典, 笠師 久美子, 山田 武宏, 小松 嘉人, 井関 健
薬学雑誌, 138, 11, 1409, 1416, 公益社団法人 日本薬学会, 2018年11月01日, ［査読有り］
日本語, It is important that pharmacists ensure safe chemotherapy implementation. In addition to inspecting chemotherapeutic prescriptions according to patient condition and drug-drug interactions, the management of chemotherapy-induced adverse effects and associated pharmaceutical intervention is one of the most important responsibilities of pharmacists in medical care teams. In May 2016, an oncology pharmacist was set responsible for the specialized, long-term, and successive pharmaceutical care, including instructions about appropriate use of medication at an outpatient chemotherapy center. We evaluated the effectiveness of the continuous pharmaceutical care. The number of medication counseling and associated pharmaceutical interventions increased with time. Specifically, the number of pharmaceutical interventions (prescription questions and pharmaceutical proposals) was 745 (459 and 286, respectively) in the surveillance period, which significantly increased compared to that observed within the same duration before posting an oncology pharmacist. The adoption rate was approximately 70% for prescription questions and 98% for pharmaceutical proposals. We also found that approximately 70% of the proposals attenuated the painful symptoms. Furthermore, approximately 60% of all pharmaceutical interventions were established after the third visit; in particular, approximately 20% of the pharmaceutical proposals were suggested after the sixth visit, indicating that continuous medication counseling results in an increase in pharmaceutical proposals. In conclusion, long-term and successive pharmaceutical care by oncology pharmacy specialists in outpatient chemotherapy contributes to a safe and less onerous chemotherapy implementation, as it has been highly adopted, is effective in many cases, and has been proven to be important for risk management in chemotherapy.

血液培養陽性患者におけるde-escalationの実施状況とその有用性調査　　　　　　　　　　　　　　　
新沼 悠介, 今井 俊吾, 冨山 直樹, 鏡 圭介, 山神 彰, 山田 武宏, 小林 正紀, 井関 健, 石黒 信久, 福元 達也
北海道病院薬剤師会誌, 95, 9, 12, (一社)北海道病院薬剤師会, 2018年11月, ［査読有り］
日本語, 当院における血液培養陽性患者へのde-escalationの実施状況を調査し、その有用性(薬剤コスト削減効果)を明らかにした。菌種別のde-escalation実施率は、メチシリン感受性黄色ブドウ球菌(MSSA)が最も高く、次いで腸球菌属(Enterococcus spp.)が高かった。抗MRSA薬からの切り替えが最も多く、これはMSSAやEnterococcus spp.のde-escalation実施率の高さに反映されていると考えられた。de-escalation実施群と非実施群における治療失敗率の比較では、両群において有意差は認められなかった。治療失敗例は各群に1例ずつ認められ、その内訳はいずれも培養結果判明後30日以内の死亡であり、感染症の重篤度との関連が疑われた症例である。菌種はそれぞれMSSAとEnterococcus spp.であった。菌の持続的検出・菌血症の再発に関しては両群で認められなかった。治療期間と治療コストの比較については、de-escalation実施群に関して多剤で治療を行っていた症例が有意に多かったため、単剤で治療を行った症例を抽出し比較を行った。治療期間に関しては両群で差は認められなかった。一方、治療コストに関しては実施群で有意に低いという結果となった。

Genetic variations in the monocarboxylate transporter genes (SLC16A1, SLC16A3, and SLC16A11) in the Japanese population.
Yuki Kimura, Masaki Kobayashi, Masaru Asari, Issei Higuchi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Drug metabolism and pharmacokinetics, 33, 5, 215, 218, 2018年10月, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, MCT1 (SLC16A1), MCT4 (SLC16A3), and MCT11 (SLC16A11) are members of the monocarboxylate transporter (MCT) family. MCT1 and MCT4 transport pH-related monocarboxylates, such as lactate and pyruvate. MCT11 may also be a proton-coupled monocarboxylate transporter. Although alterations of these substrates are involved in the pathology of cancer and diabetes, little is known about MCT polymorphisms. In this study, genetic variation was evaluated in SLC16A1, SLC16A3, and SLC16A11 in the Japanese population (healthy volunteers, n = 92). Polymorphisms in the coding regions of the SLC16A1, SLC16A3, and SLC16A11 genes were screened by DNA sequencing. A single polymorphism that caused a change in the amino acid sequence was found in SLC16A1 (rs1049434 (T1470A, D490E)) and in SLC16A3 (rs368788465 (C641T, S214F)). Five polymorphisms were detected in the SLC16A11 gene (rs117767867 (G337A, V113I), rs13342692 (A380G, D127G), rs13342232 (T561C, silent), rs75418188 (G1018A, G340S), and rs75493593 (C1327A, P443T)). This information for a healthy population provides a comparison for further studies of patients with various diseases such as cancer and diabetes.

Evaluation of the effects of antiepileptic drugs on folic acid uptake by human placental choriocarcinoma cells.
Yuko Kurosawa, Ayako Furugen, Ayako Nishimura, Katsuya Narumi, Masaki Kobayashi, Ken Iseki
Toxicology in vitro : an international journal published in association with BIBRA, 48, 104, 110, 2018年04月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Folate status during pregnancy is important for fetal development and health. The placenta plays an important role in supplying the fetus with folate. Most women with epilepsy continue their medication during pregnancy. In the present study, we aimed to evaluate the effects of 16 antiepileptic drugs, clinically used for treatment of epilepsy, on folic acid uptake in two in vitro placental models, BeWo and JEG-3 cells. Short-term exposure to antiepileptic drugs had no effects on [3H]-folic acid uptake by BeWo cells. However, long-term exposure (24h) to valproic acid (VPA) increased [3H]-folic acid uptake by BeWo and JEG-3 cells. VPA treatment for 24h increased folate receptor-α (FRα) and proton-coupled folate transporter (PCFT) mRNA expression; however, it did not affect reduced folate carrier expression. These results suggested that the increase in folic acid uptake after exposure to VPA can be attributed to the induction of FRα and PCFT expression. Furthermore, the present study showed that exposure to clinical concentrations of oxcarbazepine and stiripentol reduced the viability of BeWo cells. Therefore, the findings of the present study may contribute to better understanding of the mechanisms of toxicity of antiepileptic drugs, and estimation of their potential risk to fetus.

Reishi mushroom Ganoderma lucidum Modulates IgA production and alpha-defensin expression in the rat small intestine.
Atsuhito Kubota, Masaki Kobayashi, Sota Sarashina, Reiko Takeno, Keisuke Okamoto, Katsuya Narumi, Ayako Furugen, Yuji Suzuki, Natsuko Takahashi, Ken Iseki
Journal of ethnopharmacology, 214, 240, 243, 2018年03月25日, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, ETHNOPHARMACOLOGICAL RELEVANCE: Immunoglobulin A (IgA) secretion and alpha-defensins play a role in the innate immune system to protect against infection. Ganoderma lucidum (W.Curt.: Fr.) P. Karst. (Reishi) is a well-known mushroom in traditional Chinese medicine. This study aimed to determine the effects of Reishi on IgA secretion from Peyer's patch (PP) cells and alpha-defensin-5 (RD-5) and RD-6 expression in the rat small intestine. MATERIALS AND METHODS: The rats received an oral injection of 0.5-5mg/kg of Reishi powder (1mL/kg) by sonde. All animals were euthanized 24h after Reishi administration. We examined RD-5, RD-6, and Toll-like receptor (TLR) 4 mRNA levels in the jejunum, ileum, and in Peyer's patches (PP) through quantitative real-time PCR analysis. IgA secretion from PP was measured through enzyme-linked immunosorbent assay of the supernatant after primary culture. RESULTS: Reishi increased IgA secretion in the presence of lipopolysaccharide (LPS) and increased TLR4 mRNA levels, but had no effect on the viability of PP cells. Moreover, Reishi increased RD-5, RD-6, and TLR4 mRNA levels significantly in the ileum in a concentration-dependent manner. CONCLUSIONS: Reishi can induce IgA secretion and increase the mRNA levels of RD-5 and RD-6 in the rat small intestine, through a TLR4-dependent pathway. The present results indicate that Reishi might reduce the risk of intestinal infection.

Identification of a selective inhibitor of human monocarboxylate transporter 4.
Yuya Futagi, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Biochemical and biophysical research communications, 495, 1, 427, 432, 2018年01月01日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, The human monocarboxylate transporters (hMCTs/SLC16As) mediate the uptake of various monocarboxylates. Several isoforms of hMCTs are expressed in cancerous tissue as well as in normal tissue. In cancerous tissue, hypoxia induces the expression of hMCT4, which transports the energetic metabolite l-lactate across the plasma membrane. Since hMCT4 is involved in pH regulation and the transport of l-lactate in cancer cells, an hMCT4 inhibitor could function as an anticancer agent. Although several non specific hMCT inhibitors have been developed, a selective hMCT4 inhibitor has not yet been identified. The aim of this study was therefore to identify a selective hMCT4 inhibitor for use as a pharmacological tool for studying hMCT4. The heterologous expression system of the Xenopus oocyte was used to assess the effects of test compounds on hMCT4, whereupon isobutyrate derivatives, fibrates, and bindarit (2-[ (1-benzyl-1H-indazol-3-yl)methoxy]-2-methylpropanoic acid) were demonstrated to exhibit selective inhibitory effects against this transporter. It is suggested that the structure formed from the joining of an isobutyrate moiety and two aromatic rings by appropriate linkers is important for acquiring the selective hMCT4-inhibiting activity. These findings provide novel insights into the ligand recognition of hMCT4, and contribute to the development of novel anticancer agents.

Involvement of monocarboxylate transporter 1 (SLC16A1) in the uptake of l-lactate in human astrocytes.
Masaya Ideno, Masaki Kobayashi, Shotaro Sasaki, Yuya Futagi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Life sciences, 192, 110, 114, 2018年01月01日, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, PURPOSE: Astrocytes, the most abundant glial cells in the central nervous system (CNS), help neurons survive. Monocarboxylate transporters (MCTs) are reported to transport l-lactate, which is important for CNS physiology and cognitive function. However, it remains unclear which MCT isoform is functionally expressed by human astrocytes. The aim of this study was to establish the contribution of each MCT isoform to l-lactate transport in human astrocytes. METHODS: The function of l-lactate transport was studied using NHA cells as a human astrocyte model and radiolabeled l-lactate. The expression of MCT in human astrocytes was detected by immunohistochemistry staining. RESULTS: The cellular uptake of l-lactate was found to be pH- and concentration-dependent with a Km value for l-lactate uptake of 0.64mM. This Km was similar to what has been previously established for MCT1-mediated l-lactate uptake. α-Cyano-4- hydroxycinnamate (CHC) and 5-oxoproline, which are both MCT1 inhibitors, were found to significantly inhibit the uptake of l-lactate, suggesting MCT1 is primarily responsible for l-lactate transport. Moreover, MCT1 protein was expressed in human astrocytes. CONCLUSION: pH-dependent l-lactate transport is mediated by MCT1 in human astrocytes.

Gamma-Aminobutyric Acid (GABA) Attenuates Ischemia Reperfusion-Induced Alterations in Intestinal Immunity.
Atsuhito Kubota, Masaki Kobayashi, Sota Sarashina, Reiko Takeno, Genki Yasuda, Katsuya Narumi, Ayako Furugen, Natsuko Takahashi-Suzuki, Ken Iseki
Biological & pharmaceutical bulletin, 41, 12, 1874, 1878, 2018年, ［査読有り］, ［国内誌］
英語, 研究論文（学術雑誌）, The aims of this study were to determine the effects of gamma-aminobutyric acid (GABA) on immunoglobulin A (IgA) secretion from Peyer's patch (PP) cells; to assess rat alpha-defensin-5 (RD-5) expression in the rat small intestine; and to determine the effect of GABA on intestinal ischemia reperfusion (I/R) injury-induced intestinal innate immunity. We found that GABA caused an increase in IgA secretion in the presence and absence of lipopolysaccharide (LPS). Moreover, GABA also significantly increased the mRNA levels of RD-5 and superoxide dismutase (Sod) 1, 3. Intestinal I/R was induced by a 30-min occlusion of the superior mesenteric artery followed by a reperfusion for 60-min. This led to a significant decrease in IgA secretion, and mRNA levels of RD-5 and Sod 1-3 in the ileum. On the other hand, administration of GABA before I/R induction had a significant protective effect against oxidative injury and attenuated the effects on intestinal immunity.

Effects of proton pump inhibitors and famotidine on elimination of plasma methotrexate: Evaluation of drug-drug interactions mediated by organic anion transporter 3
Katsuya Narumi, Yu Sato, Masaki Kobayashi, Ayako Furugen, Kumiko Kasashi, Takehiro Yamada, Takanori Teshima, Ken Iseki
BIOPHARMACEUTICS & DRUG DISPOSITION, 38, 9, 501, 508, WILEY, 2017年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Methotrexate (MTX) is an antifolate agent used in the treatment of numerous types of cancer, and eliminated by active tubular secretion via organic anion transporter 3 (OAT3). Gastric antisecretory drugs, such as proton pump inhibitors (PPIs) and histamine H-2 receptor antagonists, are widely used among patients with cancer in clinical practice. The aim of the present study was to analyse the potential drug-drug interactions between MTX and gastric antisecretory drugs in high-dose MTX (HD-MTX) therapy. The impact of PPIs on the plasma MTX concentration on 73cycles of HD-MTX therapy was analysed retrospectively in 43 patients. Also investigated was the involvement of OAT3 in PPI-MTX drug interaction in an in vitro study using human OAT3 expressing HEK293 cells. In a retrospective study, patients who received a PPI had significantly higher MTX levels at 48h (0.38 vs. 0.15mol l(-1), respectively, p=0.000018) and 72h (0.13 vs. 0.05mol l(-1), respectively, p=0.0002) compared with patients who did not receive a PPI (but received famotidine). Moreover, in vitro experiments demonstrated that PPIs (esomeprazole, lansoprazole, omeprazole and rabeprazole) inhibited hOAT3-mediated uptake of MTX in a concentration-dependent manner (IC50 values of 0.40-5.5 m), with a rank order of lansoprazole > esomeprazole > rabeprazole > omeprazole. In contrast to PPIs, famotidine showed little inhibitory effect on hOAT3-mediated MTX uptake. These results demonstrated that co-administration of PPI, but not famotidine, could result in a pharmacokinetic interaction that increases the plasma MTX levels, at least in part, via hOAT3 inhibition.

Magnesium co-administration decreases cisplatin-induced nephrotoxicity in the multiple cisplatin administration
Yoshitaka Saito, Keisuke Okamoto, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Takehiro Yamada, Ken Iseki
LIFE SCIENCES, 189, 18, 22, PERGAMON-ELSEVIER SCIENCE LTD, 2017年11月, ［査読有り］, ［責任著者］
英語, 研究論文（学術雑誌）, Purpose: Pretreatment with magnesium (Mg) has been reported to attenuate cisplatin (CDDP)-induced nephrotoxicity (CIN). This attenuation involves modulation of the expression of renal transporters, resulting in reduced renal platinum accumulation after a single round of CDDP treatment. In this study, we investigated whether Mg co-administration ameliorates CIN after multiple doses of CDDP as effectively as after a single dose.
Methods: Rats were divided into control, Mg alone, CDDP alone, and CDDP with Mg groups. Rats received CDDP (2.5 mg/kg), MgSO4 (40 mg/kg), or saline once per week for three weeks. Seven days after the third round of treatment, the kidneys were excised, and the expression of renal transporters and renal platinum accumulation were analyzed.
Results: CDDP significantly elevated serum creatinine levels, which were significantly reduced by Mg co-administration. Renal platinum accumulation was significantly lower in the CDDP-Mg group than in the CDDP group. Expression of renal organic cation transporter 2 (rOct2) and multidrug and toxin extrusion protein 1 (rMate1), which are involved in CDDP transport, did not differ between the groups. However, the expression of copper transporter 1 (rCtr1) was significantly downregulated after Mg co-administration.
Conclusion: Mg co-administration significantly attenuated CIN by reducing renal platinum accumulation even after multiple rounds of treatment with CDDP as effectively as in a model of a single CDDP administration. However, the specific underlying mechanism was different between single and multiple administrations, further studies will be needed to identify what contributes to this difference and to elucidate how Mg regulates the expression of renal transporters.

The flexible cytoplasmic loop 3 contributes to the substrate affinity of human monocarboxylate transporters.
Yuya Futagi, Shotaro Sasaki, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
Biochimica et biophysica acta. Biomembranes, 1859, 10, 1790, 1795, 2017年10月, ［査読有り］, ［責任著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Human monocarboxylate transporters (hMCTs/SLC16As) mediate the transport of small molecular weight monocarboxylates. Among hMCTs, hMCT1 exhibits high-affinity l-lactate transport and broad substrate recognition, whereas hMCT4 shows highly specific substrate recognition and low-affinity l-lactate transport, indicating that hMCT1 and hMCT4 have different roles in the body. However, the molecular mechanism of transporter-mediated substrate transport remains unknown. The aim of this study is to identify the domain, which determines the substrate selectivity and affinity of hMCT1 and hMCT4. We constructed a chimera, hMCT4/1, in which the cytoplasmic loop 3 (TM6/7loop) region of hMCT4 was replaced by the corresponding region of hMCT1. Xenopus laevis oocyte heterologous expression system was used to characterize functional features of the chimera. We have demonstrated that the substrate affinity of hMCT1 and hMCT4 depends on the TM6/7loop. Non-conserved His237 residue in the TM6/7loop functions as a regulatory moiety of the substrate affinity. In contrast, the substrate selectivity of the transporters did not depend on the TM6/7loop, suggesting that the domain is not directly involved in substrate recognition. Our study provides important insights into the structures and functions of hMCT1 and hMCT4 transporters. These findings contribute to the development of novel hMCT1 and/or hMCT4 inhibitors as anticancer agents.

Magnesium attenuates cisplatin-induced nephrotoxicity by regulating the expression of renal transporters
Yoshitaka Saito, Keisuke Okamoto, Masaki Kobayashi, Katsuya Narumi, Takehiro Yamada, Ken Iseki
EUROPEAN JOURNAL OF PHARMACOLOGY, 811, 191, 198, ELSEVIER SCIENCE BV, 2017年09月, ［査読有り］, ［責任著者］
英語, 研究論文（学術雑誌）, Cisplatin ( CDDP)-induced nephrotoxicity (CIN) is one of the most serious toxicities caused by this potent antitumor agent. It has been reported that Mg premedication attenuates CIN in clinical trials; however, the mechanism underlying its nephroprotection is not fully understood. Therefore, the aim of this study was to determine whether Mg administration affects CDDP accumulation by regulating the expression level of renal transporters. Rats were divided into control, Mg (40 mg/kg) alone, 2.5 mg/kg CDDP with (20 and 40 mg/kg) and without Mg, 5 mg/kg CDDP groups. These substances were administered on the same day and 7 days later their kidneys were removed. The expression levels of renal transporters and platinum (Pt) accumulation were analyzed. The serum creatinine level was significantly increased by CDDP administration and treatment with Mg significantly ameliorated such elevation. The expressions of the renal organic cation transporter 2 (rOct2) and renal multidrug and toxin extrusion protein 1 (rMate1) were downregulated and upregulated, respectively following co-administration with Mg, which significantly reduced the renal Pt accumulation in the 2.5 mg/kg CDDP-treated group. Moreover, Mg dose-dependently downregulated rOct2, not affecting rMate expression, resulting in the attenuation of CIN. Mg co-administration protected the downregulation of the transient receptor potential subfamily Melastatin 6 (rTrpm6), but not the epidermal growth factor (rEgf), as a result, Mg co-injection attenuated CDDP-induced hypomagnesemia. In conclusion, Mg co-administration reduced Pt accumulation by regulating the expression of the renal transporters, rOct2 and rMate1 and, thereby, attenuated CIN.

Insulin stimulates transport of organic anion compounds mediated by organic anion transporting polypeptide 2B1 in the human intestinal cell line Caco-2
Taku Kobayashi, Takahiro Koizumi, Masaki Kobayashi, Jiro Ogura, Yuichi Horiuchi, Yuki Kimura, Ayuko Kondo, Ayako Furugen, Katsuya Narumi, Natsuko Takahashi, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 32, 2, 157, 163, JAPANESE SOC STUDY XENOBIOTICS, 2017年04月, ［査読有り］, ［責任著者］
英語, 研究論文（学術雑誌）, Organic anion transporting polypeptide 2B1 (OATP2B1) is the major uptake transporter in the intestine, and transports various clinically used therapeutic agents. Insulin acts through the insulin receptor in targeted cells, and Rab8A is one of the insulin signaling pathways. The small intestine in humans also expresses insulin receptor and Rab8A. It has been reported that insulin stimulates peptide transporter 1 (PEPT1) expression at the apical membrane and increases uptake of PEPT1 substrates in small intestine epithelial model cells (Caco-2 cells). However, the effect of insulin on OATP2B1 in the small intestine has not been fully investigated. We found that Rab8A was associated with OATP2B1-mediated estrone-3-sulfate (E3S) uptake. Insulin stimulated the uptake of E3S by Caco-2 cells and the enhancement was sustained for 120 min. The Vmax value of E3S uptake significantly increased upon insulin exposure. Caco-2 cells treated with insulin showed increased OATP2B1 expression at the cell surface. The apical-to-basal transport of E3S was also increased by insulin. The increase of E3S transport was inhibited by the cold condition (4 degrees C) or the OATP2B1 inhibitor, taurocholate. These results indicate that insulin acts on the small intestine to increase OATP2B1-mediated absorption. (C) 2016 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Organic anion-transporting polypeptide (OATP) 2B1 contributes to the cellular uptake of theaflavin
Ayuko Kondo, Katsuya Narumi, Jiro Ogura, Ai Sasaki, Keisuke Yabe, Taku Kobayashi, Ayako Furugen, Masaki Kobayashi, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 32, 2, 145, 150, JAPANESE SOC STUDY XENOBIOTICS, 2017年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Organic anion-transporting polypeptide (OATP) 2B1 has been reported in the apical membranes of the human small intestinal epithelium, where it contributes to the intestinal absorption of pharmacologically active drugs. To investigate the potential for OATP2B1-mediated drug-food interactions, the effects of several polyphenolic compounds on OATP2B1-mediated estrone-3-sulfate (E3S) transport were studied by using OATP2B1-expressing HEK293 cells. Our results showed that some compounds, especially theaflavin, were strong inhibitors of OATP2B1-mediated E3S uptake. Theaflavin showed a significantly higher uptake into the OATP2B1-expressing HEK293 cells than the control cells. The concentration dependence of the uptake of theaflavinwas determined over a range of concentrations (0.5-100 mu M) and the kinetic parameters (K-m and V-max) of theaflavin uptake were found to be 5.12 +/- 0.67 mu M and 41.6 +/- 1.3 pmol/mg protein/min, respectively. The OATP2B1-mediated theaflavin uptake was inhibited by known OATP2B1 substrates such as E3S, bromsulphthalein (BSP), dehydroepiandrosterone-3-sulfate (DHEAS), and fluvastatin. Our results indicate that theaflavin is a novel substrate of OATP2B1. The results of this study might be helpful to predict the potential OATP2B1-mediated drug-theaflavin interactions and to avoid undesirable clinical consequences. (C) 2016 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Fructose suppresses uric acid excretion to the intestinal lumen as a result of the induction of oxidative stress by NADPH oxidase activation.
Chihiro Kaneko, Jiro Ogura, Shunichi Sasaki, Keisuke Okamoto, Masaki Kobayashi, Kaori Kuwayama, Katsuya Narumi, Ken Iseki
Biochimica et biophysica acta. General subjects, 1861, 3, 559, 566, 2017年03月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, BACKGROUND: A high intake of fructose increases the risk for hyperuricemia. It has been reported that long-term fructose consumption suppressed renal uric acid excretion and increased serum uric acid level. However, the effect of single administration of fructose on excretion of uric acid has not been clarified. METHODS: We used male Wistar rats, which were orally administered fructose (5g/kg). Those rats were used in each experiment at 12h after administration. RESULTS: Single administration of fructose suppressed the function of ileal uric acid excretion and had no effect on the function of renal uric acid excretion. Breast cancer resistance protein (BCRP) predominantly contributes to intestinal excretion of uric acid as an active homodimer. Single administration of fructose decreased BCRP homodimer level in the ileum. Moreover, diphenyleneiodonium (DPI), an inhibitor of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox), recovered the suppression of the function of ileal uric acid excretion and the Bcrp homodimer level in the ileum of rats that received single administration of fructose. CONCLUSIONS: Single administration of fructose decreases in BCRP homodimer level, resulting in the suppression the function of ileal uric acid excretion. The suppression of the function of ileal uric acid excretion by single administration of fructose is caused by the activation of Nox. The results of our study provide a new insight into the mechanism of fructose-induced hyperuricemia.

Premedication with intravenous magnesium has a protective effect against cisplatin-induced nephrotoxicity
Yoshitaka Saito, Masaki Kobayashi, Takehiro Yamada, Kumiko Kasashi, Rio Honma, Satoshi Takeuchi, Yasushi Shimizu, Ichiro Kinoshita, Hirotoshi Dosaka-Akita, Ken Iseki
SUPPORTIVE CARE IN CANCER, 25, 2, 481, 487, SPRINGER, 2017年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Magnesium supplementation is an effective protective method against cisplatin-induced nephrotoxicity (CIN); however, there are few reports regarding the mechanism of its nephroprotective effect. The aim of this study was to determine whether premedication with intravenous magnesium prevents CIN and to determine the relationship between its nephroprotective effect and serum magnesium level.Fifty-eight patients with head and neck cancer who received cisplatin, docetaxel, and 5-fluorouracil (DCF) were retrospectively investigated. Grade 2 or more serum creatinine elevation was defined as CIN. The incidence of CIN was compared between a magnesium sulfate (20 mEq, 2.46 g) premedication group and a non-magnesium group during the first cycle and in all cycles.CIN did not occur in any patients receiving magnesium premedication but did occur in 5 of 29 patients during the first cycle and in 6 patients during all subsequent cycles in patients who did not receive magnesium premedication. Furthermore, the variation of creatinine clearance was significantly worse in the non-magnesium group than in the magnesium premedication group from baseline. There was no difference in adverse effects or response rate between the two groups. Univariate analysis suggested that magnesium premedication significantly reduced the risk of CIN. On the other hand, serum magnesium depletion was seen in both groups to equal degrees despite supplementation.Intravenous magnesium premedication has a protective effect on cisplatin-induced nephrotoxicity without the influence on the serum magnesium level. Magnesium premedication is a simple nephroprotective method that does not influence other adverse effects or rate of response to chemotherapy.

Validating the Effectiveness of Switching the Vancomycin TDM Analysis Software Based on the Predictive Accuracy.
Shungo Imai, Takehiro Yamada, Nobuhisa Ishiguro, Takenori Miyamoto, Keisuke Kagami, Naoki Tomiyama, Yusuke Niinuma, Daisuke Nagasaki, Koji Suzuki, Akira Yamagami, Kumiko Kasashi, Masaki Kobayashi, Ken Iseki
Yakugaku zasshi : Journal of the Pharmaceutical Society of Japan, 137, 9, 1185, 1192, 2017年, ［査読有り］, ［国内誌］
日本語, 研究論文（学術雑誌）, Based on the predictive performance in our previous study, we switched the therapeutic drug monitoring (TDM) analysis software for dose setting of vancomycin (VCM) from "Vancomycin MEEK TDM analysis software Ver2.0" (MEEK) to "SHIONOGI-VCM-TDM ver.2009" (VCM-TDM) in January 2015. In the present study, our aim was to validate the effectiveness of the changing VCM TDM analysis software in initial dose setting of VCM. The enrolled patients were divided into two groups, each having 162 patients in total, who received VCM with the initial dose set using MEEK (MEEK group) or VCM-TDM (VCM-TDM group). We compared the rates of attaining the therapeutic range (trough value; 10-20 μg/mL) of serum VCM concentration between the groups. Multivariate logistic regression analysis was performed to confirm that changing the VCM TDM analysis software was an independent factor related to attaining the therapeutic range. Switching the VCM TDM analysis software from MEEK to VCM-TDM improved the rate of attaining the therapeutic range by 21.6% (MEEK group: 42.6% vs. VCM-TDM group: 64.2%, p<0.01). Patient age ≥65 years, concomitant medication (furosemide) and the TDM analysis software used VCM-TDM were considered to be independent factors for attaining the therapeutic range. These results demonstrated the effectiveness of switching the VCM TDM analysis software from MEEK to VCM-TDM for initial dose setting of VCM.

Involvement of L-type amino acid transporter 1 in the transport of gabapentin into human placental choriocarcinoma cells
Ayako Furugen, Yuri Ishiguro, Masaki Kobayashi, Katsuya Narumi, Ayako Nishimura, Takeshi Hirano, Ken Iseki
REPRODUCTIVE TOXICOLOGY, 67, 48, 55, PERGAMON-ELSEVIER SCIENCE LTD, 2017年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Gabapentin (GBP) is a widely used antiepileptic drug, with potential for use in the treatment of epilepsy in pregnant women. Although studies have examined GBP transport mechanisms across the blood-brain barrier, kidney, and intestine, the mechanism in the placenta has not been fully elucidated. We previously reported that GBP accumulates at high concentrations in human placental choriocarcinoma BeWo cells. The purpose of this study was to examine the transport mechanism of GBP in placental choriocarcinoma cells (BeWo and JEG-3), and to identify the carrier involved. High concentrations of intracellular GBP accumulations were also found in JEG-3 cells. A kinetic analysis showed that a single carrier system was involved in the uptake of GBP. Furthermore, substrates for L-type amino acid transporter (LAT) and siRNAs targeted to LAT1 significantly decreased GBP uptake. Our observations from this study suggest that LAT1 is the main contributor to GBP transport in placental choriocarcinoma cells. (C) 2016 Published by Elsevier Inc.

Characterization of loxoprofen transport in Caco-2 cells: the involvement of a proton-dependent transport system in the intestinal transport of loxoprofen
Katsuya Narumi, Masaki Kobayashi, Ayuko Kondo, Ayako Furugen, Takehiro Yamada, Natsuko Takahashi, Ken Iseki
BIOPHARMACEUTICS & DRUG DISPOSITION, 37, 8, 447, 455, WILEY-BLACKWELL, 2016年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Loxoprofen, a propionate non-steroidal anti-inflammatory drug (NSAID), is used widely in East Asian countries. However, little is known about the transport mechanisms contributing to its intestinal absorption. The objectives of this study were to characterize the intestinal transport of loxoprofen using the human intestinal Caco-2 cell model. The transport of loxoprofen was investigated in cellular uptake studies. The uptake of loxoprofen into Caco-2 cells was pH- and concentration-dependent, and was described by a Michaelis-Menten equation with passive diffusion (K-m: 4.8mm, V-max: 142nmol/mg protein/30s, and K-d: 2.2l/mg protein/30s). Moreover, the uptake of loxoprofen was inhibited by a typical monocarboxylate transporter (MCT) inhibitor as well as by various monocarboxylates. The uptake of [C-14] l-lactic acid, a typical MCT substrate, in Caco-2 cells was saturable with relatively high affinity for MCT. Because loxoprofen inhibited the uptake of [C-14] l-lactic acid in a noncompetitive manner, it was unlikely that loxoprofen uptake was mediated by high-affinity MCT(s). Our results suggest that transport of loxoprofen in Caco-2 cells is, at least in part, mediated by a proton-dependent transport system. Copyright (c) 2016 John Wiley & Sons, Ltd.

Interaction of atorvastatin with the human glial transporter SLC16A1
Shotaro Sasaki, Yuya Futagi, Masaya Ideno, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
EUROPEAN JOURNAL OF PHARMACOLOGY, 788, 248, 254, ELSEVIER SCIENCE BV, 2016年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Solute carrier (SLC) 16A1 is a pH-dependent carrier of 5-oxoproline, a derivative of the amino acid. SLC16A1 interacts with carboxylate group-containing substrates, which are also present in atorvastatin, and might be the reason for its ability to interact with atorvastatin. Does atorvastatin interact with the carrier? Does it also interact with the carrier via the substrate recognition site? This study was carried out to answer these questions. Polymerase chain reaction was used to determine the expression of SLC16A1 in normal human astrocytes. We induced SLC16A1 expression in a mammalian cell line and in Xenopus laevis oocytes. We used [H-3] 5-oxoproline for direct measurement of SLC16A1-specific transport activity. SLC16A1 was clearly observed in normal human astrocytes. 3-Hydroxy-3-methyl-glutaryl-CoA reductase inhibitors inhibited the SLC16A1-specific transport of 5-oxoproline. Atorvastatin was the most potent inhibitor, with an inhibition constant of 40 mu M. The drug was a non-competitive inhibitor of SLC16A1. In the present study, we showed non-competitive inhibition of SLC16A1-specific transport activity by atorvastatin. However, the affinity between the drug and the carrier was extremely low. Therefore, the interaction of atorvastatin with SLC16A1 is unlikely to be a problem in clinical practice. (C) 2016 Published by Elsevier B.V.

Effect of diclofenac on SLC16A3/MCT4 by the Caco-2 cell line
Shotaro Sasaki, Yuya Futagi, Masaya Ideno, Masaki Kobayashi, Katsuya Narumi, Ayako Furugen, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 31, 3, 218, 223, JAPANESE SOC STUDY XENOBIOTICS, 2016年06月, ［査読有り］
英語, 研究論文（学術雑誌）, In the present study, we demonstrated that monocarboxylate transporter 4 (MCT4) is functionally expressed in Caco-2 cells. We studied the effects of 4 nonsteroidal anti-inflammatory drugs on the uptake of L-lactate as a good substrate of MCT4 by the cells. The monocarboxylate drugs inhibited the uptake of L-lactate into the cells. Diclofenac, as a member of the aryl-acetic acid group of nonsteroidal anti-inflammatory drugs, was the most potent inhibitor, with an inhibition constant of 20 mu M. In the next study, we determined the type of inhibition for diclofenac. An L-lactate carrier is non-competitively inhibitable by the drug. We also demonstrated, in Xenopus oocyte expression system, potential of diclofenac for MCT4 inhibitor. The present results could provide a useful tool to discover MCT4-specific inhibitors. (C) 2016 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Involvement of Monocarboxylate Transporter 4 Expression in Statin-Induced Cytotoxicity
Yurika Kikutani, Masaki Kobayashi, Toru Konishi, Shotaro Sasaki, Katsuya Narumi, Ayako Furugen, Natsuko Takahashi, Ken Iseki
JOURNAL OF PHARMACEUTICAL SCIENCES, 105, 4, 1544, 1549, WILEY-BLACKWELL, 2016年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Statins, 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors, are the most widely used cholesterol-lowering agents for prevention of obstructive cardiovascular events. However, statins can cause a variety of skeletal muscle problems, and exercise leads to an increase in statin-induced muscle injury. Exercise induces the protein content of monocarboxylate transporter 4 (MCT4), which is expressed strongly in skeletal muscle and is thought to play a major role in the transport of metabolically important monocarboxylates such as L-lactate. We previously reported that alpha-cyano-4-hydroxycinnamate, an MCT4 inhibitor, increased the inhibition of growth of RD cells, a prototypic embryonal rhabdomyosarcoma cell line (an RD cell line), as a model of in vitro skeletal muscle, induced by a statin. However, it is unclear whether statin-induced RD cell cytotoxicity is associated with MCT4 expression. We, therefore, examined the relationship between statin-induced cytotoxicity and MCT4 expression in RD cells. Atorvastatin reduced the number of viable cells and upregulated MCT4, but not MCT1, mRNA level in a concentration-dependent manner. MCT4 knockdown suppressed atorvastatin-, simvastatin-, and fluvastatin-induced reduction of cell viability and apoptosis compared with negative controletreated cells. In this study, we demonstrated that MCT4 expression is associated with statin-induced cytotoxicity. (C) 2016 American Pharmacists Association(R). Published by Elsevier Inc. All rights reserved.

Influence of high glucose state on bromopyruvate-induced cytotoxity by human colon cancer cell lines
Masaya Ideno, Shotaro Sasaki, Masaki Kobayashi, Yuya Futagi, Katsuya Narumi, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 31, 1, 67, 72, JAPANESE SOC STUDY XENOBIOTICS, 2016年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Background: Attention must be paid to chemotherapy for cancer patients in a hyperglycemia state. It is difficult for chemotherapy to cure cancer in patients in a hyperglycemia state. This study was carried out to determine the change in cell viability after treatment with bromopyruvate, which is an alkylating drug with anti-tumor activity, in a high glucose condition.
Methods: The function of L-lactate and bromopyruvate transport was studied using human colon cancer cell lines (LoVo and HT-29) and radiolabeled L-lactate and bromopyruvate. Cell viability was monitored by the trypan blue exclusion assay. The expression level of human monocarboxylate transporter 1 (hMCT1) was evaluated by Western blot analysis.
Results: Bromopyruvate-induced cell death was suppressed by a high glucose condition. L-Lactate and bromopyruvate uptake were suppressed by a high glucose condition. hMCT1 as a bromopyruvate carrier was functionally expressed in the cells. However, the expression of hMCT1 was suppressed by a high glucose state.
Conclusions: Down-regulation of hMCT1 by a high glucose state is one of the possibilities of the bromopyruvate resistance. We should pay scrupulous attention to cancer chemotherapy for patients who have developed diabetes. Copyright (C) 2015, Published by Elsevier Ltd on behalf of The Japanese Society for the Study of Xenobiotics.

Quantification of new antiepileptic drugs by liquid chromatography/electrospray ionization tandem mass spectrometry and its application to cellular uptake experiment using human placental choriocarcinoma BeWo cells
Ayako Furugen, Masaki Kobayashi, Ayako Nishimura, Shigeo Takamura, Katsuya Narumi, Takehiro Yamada, Ken Iseki
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 1002, 228, 233, ELSEVIER SCIENCE BV, 2015年10月, ［査読有り］
英語, 研究論文（学術雑誌）, A method for quantification of new antiepileptic drugs, including lamotrigine (LTG), levetiracetam (LEV), gabapentin (GBP), and topiramate (TPM), in cellular samples, using liquid chromatography/electrospray ionization tandem mass spectrometry was developed to better understand the membrane transport mechanisms of these drugs. Cell lysate was deproteinized by methanol containing LEV-d(3) as an internal standard (IS). Chromatographic separation was performed on a C18 column using gradient elution with methanol-water-formic acid (10:90:0.1, v/v/v) and methanol-formic acid (100:0.1, v/v). Analytes were detected in positive ion electrospray mode with selected reaction monitoring (SRM). This method was applicable for a linear range of 5 to 500 pmol for LTG; 5 to 1000 pmol for LEV; 10 to 10,000 pmol for GBP; and 5 to 5000 pmol for TPM. The intra-day precision, inter-day precision, and accuracy data were assessed and found to be acceptable. This developed and validated method was then successfully applied to the investigation of uptake of the new antiepileptic drugs in placental choriocarcinoma BeWo cells. The intracellular concentration of these drugs in BeWo cells, accumulating over 30 min at 37 degrees C was in the order of GBP > LTG > LEV approximate to TPM. Furthermore, the uptake of GBP at 4 degrees C was much lower than that at 37 degrees C. The uptake of GBP was saturated at high concentrations. The kinetic parameters calculated for GBP uptake in BeWo cells were determined as K-m of 105.4 +/- 6.4 mu M and Vmax at 8153 +/- 348 pmol/mg protein/min. The novel method described here should enable investigators to elucidate the transport mechanisms of these antiepileptic drugs in BeWo cells. (C) 2015 Elsevier B.V. All rights reserved.

Reactive oxygen species derived from xanthine oxidase interrupt dimerization of breast cancer resistance protein, resulting in suppression of uric acid excretion to the intestinal lumen
Jiro Ogura, Kaori Kuwayama, Shunichi Sasaki, Chihiro Kaneko, Takahiro Koizumi, Keisuke Yabe, Takashi Tsujimoto, Reiko Takeno, Atsushi Takaya, Masaki Kobayashi, Hiroaki Yamaguchi, Ken Iseki
BIOCHEMICAL PHARMACOLOGY, 97, 1, 89, 98, PERGAMON-ELSEVIER SCIENCE LTD, 2015年09月, ［査読有り］
英語, 研究論文（学術雑誌）, The prevalence of hyperuricemia/gout increases with aging. However, the effect of aging on function for excretion of uric acid to out of the body has not been clarified. We found that ileal uric acid clearance in middle-aged rats (11-12 months) was decreased compared with that in young rats (2 months). In middle-aged rats, xanthine oxidase (XO) activity in the ileum was significantly higher than that in young rats. Inosine-induced reactive oxygen species (ROS), which are derived from XO, also decreased ileal uric acid clearance. ROS derived from XO decreased the active homodimer level of breast cancer resistance protein (BCRP), which is a uric acid efflux transporter, in the ileum. Pre-administration of allopurinol recovered the BCRP homodimer level, resulting in the recovering ileal uric acid clearance. Moreover, we investigated the effects of ROS derived from XO on BCRP homodimer level directly in Caco-2 cells using hypoxanthine. Treatment with hypoxanthine decreased BCRP homodimer level. Treatment with hypoxanthine induced mitochondrial dysfunction, suggesting that the decreasing BCRP homodimer level might be caused by mitochondrial dysfunction. In conclusion, ROS derived from XO decrease BCRP homodimer level, resulting in suppression of function for uric acid excretion to the ileal lumen. ROS derived from XO may cause the suppression of function of the ileum for the excretion of uric acid with aging. The results of our study provide a new insight into the causes of increasing hyperuricemia/gout prevalence with aging. (C) 2015 Elsevier Inc. All rights reserved.

Pharmacokinetics and the optimal regimen for levofloxacin in critically ill patients receiving continuous hemodiafiltration
Takeshi Wada, Masaki Kobayashi, Yuichi Ono, Asumi Mizugaki, Kenichi Katabami, Kunihiko Maekawa, Daisuke Miyamoto, Yuichiro Yanagida, Mineji Hayakawa, Atsushi Sawamura, Ken Iseki, Satoshi Gando
Journal of Intensive Care, 3, 1, 22, BioMed Central Ltd., 2015年05月08日, ［査読有り］
英語, The aim of this study was to establish the pharmacokinetics of levofloxacin (LVFX) and determine the optimal dose of this drug in critically ill patients receiving continuous hemodiafiltration (CHDF). The results of in vivo and in vitro studies showed the pharmacokinetics of LVFX total clearance (CLtotal) according to the creatinine clearance (CLCre), dialysate flow (QD), and ultrafiltrate flow (QF), to be as follows: CLtotal (l/h) = 0.0836 × CLCre (ml/min) + 0.013 × body weight (kg) + 0.94(QD + QF) (l/h). The optimal dose of LVFX was expressed by the following formula: 50 × CLtotal. These results demonstrate that the usual dose of LVFX (500 mg) was sufficient for the patients evaluated in this study.

Regorafenib Is Transported by the Organic Anion Transporter 1B1 and the Multidrug Resistance Protein 2
Hiroki Ohya, Yoshihiko Shibayama, Jiro Ogura, Katsuya Narumi, Masaki Kobayashi, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 38, 4, 582, 586, PHARMACEUTICAL SOC JAPAN, 2015年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Regorafenib is a small molecule inhibitor of tyrosine kinases, and has been shown to improve the outcomes of patients with advanced colorectal cancer and advanced gastrointestinal stromal tumors. The transport profiles of regorafenib by various transporters were evaluated. HEK293/organic anion transporting polypeptide 1B1 (OATP1B1) cells exhibited increased drug sensitivity to regorafenib. Regorafenib inhibited the uptake of H-3-estrone sulfate by HEK293/OATP1B1 cells in a dose-dependent manner, but did not affect its elimination by P-glycoproteins. The concentration of regorafenib was significantly lower in LLC-PK1/multidrug resistance protein 2 (MRP2) cells than in LLC-PK1 cells treated with the MRP2 inhibitor, MK571. MK571 abolished the inhibitory effects of regorafenib on intracellular accumulation in LLC-PK1/MRP2 cells. The uptake of regorafenib was significantly higher in HEK293/0ATP1B1 cells than in OATP1B1-mock cells. Transport kinetics values were estimated to be K-m=15.9,um and V-max =1.24nmol/mg/min. No significant difference was observed in regorafenib concentrations between HEK293/OATP1B3 and OATP1B3-mock cells. These results indicated that regorafenib is a substrate for MRP2 and OATP1B1, and also suggest that the substrate preference of regorafenib may implicate the pharmacokinetic profiles of regorafenib.

Involvement of Histidine Residue His382 in pH Regulation of MCT4 Activity
Shotaro Sasaki, Masaki Kobayashi, Yuya Futagi, Jiro Ogura, Hiroaki Yamaguchi, Ken Iseki
PLOS ONE, 10, 4, e0122738, PUBLIC LIBRARY SCIENCE, 2015年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Monocarboxylate transporter 4 (MCT4) is a pH-dependent bi-directional lactate transporter. Transport of lactate via MCT4 is increased by extracellular acidification. We investigated the critical histidine residue involved in pH regulation of MCT4 function. Transport of lactate via MCT4 was measured by using a Xenopus laevis oocyte expression system. MCT4-mediated lactate transport was inhibited by Zn2+ in a pH physiological condition but not in an acidic condition. The histidine modifier DEPC (diethyl pyrocarbonate) reduced MCT4 activity but did not completely inactivate MCT4. After treatment with DEPC, pH regulation of MCT4 function was completely knocked out. Inhibitory effects of DEPC were reversed by hydroxylamine and suppressed in the presence of excess lactate and Zn2+. Therefore, we performed an experiment in which the extracellular histidine residue was replaced with alanine. Consequently, the pH regulation of MCT4-H382A function was also knocked out. Our findings demonstrate that the histidine residue His382 in the extracellular loop of the transporter is essential for pH regulation of MCT4-mediated substrate transport activity.

脂質異常症治療薬の副作用・薬物相互作用におけるモノカルボン酸トランスポータの役割
小林 正紀
YAKUGAKU ZASSHI, 135, 11, 1227, 1233, 公益社団法人 日本薬学会, 2015年, ［査読有り］, ［招待有り］, ［筆頭著者, 責任著者］
日本語, Although exercise and drug therapy are important to prevent progression of arteriosclerotic disease, exercise leads to an increase in muscular disorder induced by HMG-CoA reductase inhibitors (statins). Elucidation of this mechanism is needed to prevent the occurrence of muscular disorders. Since exercise induces expression of monocarboxylate transporter (MCT) 4, we focused on the association between MCT4 function and statin-induced muscle injury. First, we examined the transport of L-lactate via MCT4 using MCT4 cRNA-injected Xenopus laevis oocytes. L-lactate uptake by MCT4-expressing oocytes was markedly reduced by alkalizing the buffer pH and saturated at higher L-lactate concentrations. On the other hand, AMP-activated protein kinase (AMPK) and protein kinase C (PKC) are activated by exercise. We next examined whether AMPK and PKC activation affects the expression and function of MCT4 in rat skeletal muscle and RD cells as an in vitro skeletal muscle model. AMPK and PKC activation increased MCT4 expression level and lactate efflux by MCT4. Finally, we examined the association between MCT4 function and statin-induced cytotoxicity. Statins inhibited transport of L-lactate via MCT4 in a concentration-dependent manner. Statin-induced cytotoxicity was associated with intracellular acidification and caspase-3/7 activation. On the other hand, bicarbonate suppressed statin-induced pH alteration, caspase activation, and morphological change. The results suggest that statin-induced muscle injury exacerbated by exercise is associated with intracellular acidification and that the effects of statins on L-lactate transport are mediated by MCT4.

Functional Characterization of 5-Oxoproline Transport via SLC16A1/MCT1
Shotaro Sasaki, Yuya Futagi, Masaki Kobayashi, Jiro Ogura, Ken Iseki
JOURNAL OF BIOLOGICAL CHEMISTRY, 290, 4, 2303, 2311, AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, 2015年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Thyrotropin-releasing hormone is a tripeptide that consists of 5-oxoproline, histidine, and proline. The peptide is rapidly metabolized by various enzymes. 5-Oxoproline is produced by enzymatic hydrolysis in a variety of peptides. Previous studies showed that 5-oxoproline could become a possible biomarker for autism spectrum disorders. Here we demonstrate the involvement of SLC16A1 in the transport of 5-oxoproline. An SLC16A1 polymorphism (rs1049434) was recently identified. However, there is no information about the effect of the polymorphism on SLC16A1 function. In this study, the polymorphism caused an observable change in 5-oxoproline and lactate transport via SLC16A1. The Michaelis constant (K-m) was increased in an SLC16A1 mutant compared with that in the wild type. In addition, the proton concentration required to produce half-maximal activation of transport activity (K-0.5, H(+)) was increased in the SLC16A1 mutant compared with that in the wild type. Furthermore, we examined the transport of 5-oxoproline in T98G cells as an astrocyte cell model. Despite the fact that 5-oxoproline is an amino acid derivative, Na+-dependent and amino acid transport systems scarcely contributed to 5-oxoproline transport. Based on our findings, we conclude that H+-coupled 5-oxoproline transport is mediated solely by SLC16A1 in the cells.

Quantification of intracellular and extracellular digoxin and ouabain by liquid chromatography/electrospray ionization tandem mass spectrometry
Hiroaki Yamaguchi, Kazuaki Miyamori, Toshihiro Sato, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 972, 73, 80, ELSEVIER SCIENCE BV, 2014年12月, ［査読有り］
英語, 研究論文（学術雑誌）, A liquid chromatography/tandem mass spectrometry method for the determination of intracellular accumulation in addition to transcellular transport of digoxin and ouabain in renal epithelial HK-2 cells was developed. The solid-phase extraction Bond Elut (R) C18 (100 mg/1 mL) cartridge was used for the extraction of digoxin and ouabain from extracellular (medium) and intracellular (cell lysate) matrices. Chromatographic separation was performed on a CAPCELL PAK C18 MGII column (2.0 mm x 150 mm, 5 mu m). This method covered a linear range of 0.5-1000 ng/mL of concentrations in medium and 0.5-1000 ng of concentrations in cell lysate for digoxin and ouabain. The intra-day precision and inter-day precision of analysis were less than 11.9%, and the accuracy was within +/- 11.6%. The total run time was 16 min. Our method was successfully applied to the transport experiments of digoxin and ouabain by HK-2 cell monolayers. (C) 2014 Elsevier B.V. All rights reserved.

Quantification of intracellular and extracellular eicosapentaenoic acid-derived 3-series prostanoids by liquid chromatography/electrospray ionization tandem mass spectrometry
Nobuaki Tanaka, Hiroaki Yamaguchi, Ayako Furugen, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
PROSTAGLANDINS LEUKOTRIENES AND ESSENTIAL FATTY ACIDS, 91, 3, 61, 71, ELSEVIER SCI LTD, 2014年09月, ［査読有り］
英語, 研究論文（学術雑誌）, 3-Series prostanoids are bioactive lipid mediators synthesized from eicosapentaenoic acid (EPA). Determination of intracellular and extracellular levels of prostanoids is needed to elucidate the mechanism of action, and we therefore developed a method for quantification of intracellular and extracellular levels of 3-series prostanoids (including prostaglandin E-3 (PGE(3)), PGD(3), PGF(3 alpha), thromboxane B-3 (TXB3), and Delta(17)-6-keto PGF(1 alpha),) by using liquid chromatography/electrospray ionization tandem mass spectrometry. The separation of prostanoids was performed with a CAPCELL PAK C18 MG II column (2.0 mm x 150 mm, 3 mu m) with an isocratic flow of acetonitrile/water/acetic acid (40:60:0.1, v/v/v). This method was validated for measurement of both extracellular and intracellular samples with high levels of precision and accuracy. We applied this method to human lung epithelial A549 cells stimulated with calcium ionophore A23187 under the condition of arachidonic acid or EPA treatment and we could measure PGE3 in both intracellular and extracellular samples. (C) 2014 Elsevier Ltd. All rights reserved.

Quercetin-3-rhamnoglucoside (rutin) stimulates transport of organic anion compounds mediated by organic anion transporting polypeptide 2B1
Jiro Ogura, Takahiro Koizumi, Masahiro Segawa, Keisuke Yabe, Kaori Kuwayama, Shunichi Sasaki, Chihiro Kaneko, Takashi Tsujimoto, Masaki Kobayashi, Hiroaki Yamaguchi, Ken Iseki
BIOPHARMACEUTICS & DRUG DISPOSITION, 35, 3, 173, 182, WILEY-BLACKWELL, 2014年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Quercetin-3-rhamnoglucoside (rutin) has a wide spectrum of biochemical and pharmacological activities. Rutin is absorbed mainly in its unmetabolized form. Organic anion transporting polypeptide (OATP) 2B1 is a major uptake transporter in the intestine. Thus, it is important for the prevention of adverse events to understand drug interactions mediated by OATP2B1 in the absorption process. This study assessed the effect of rutin on transport by OATP2B1. Rutin stimulated the uptake of estrone-3-sulfate (E-3-S), taurocholic acid (TCA), cholic acid (CA) and rosuvastatin by OATP2B1, but not p-coumaric acid or ferulic acid. The EC50 of rutin for transport by OATP2B1 was 2.32 mu m. The K-m value of E-3-S for OATP2B1 in the presence of rutin (9.21 mu m) was almost the same as that in the absence of rutin (8.53 mu m). On the other hand, the V-max of E-3-S transport by OATP2B1 in the presence of rutin (270 pmol/mg protein/min) was 1.2-fold higher than that in the absence of rutin (218 pmol/mg protein/min). Moreover, the expression level of OATP2B1 on the cell membrane was increased by treatment with rutin for 5 min without alteration of the total OATP2B1 expression level. Moreover, the increase in the localization of OATP2B1 at the cell surface was detected by the immunocytochemistry. The stimulatory effect of rutin is a little weak but may affect the absorption of OATP2B1 substrates, because rutin is taken daily in foods and its intestinal concentration would reach the stimulatory range of OATP2B1. Copyright (c) 2014 John Wiley & Sons, Ltd.

Immunoprotective effect of epigallocatechin-3-gallate on oral anticancer drug-induced α-defensin reduction in Caco-2 cells.
Takahashi N, Kobayashi M, Ogura J, Yamaguchi H, Satoh T, Watanabe K, Iseki K
Biological & pharmaceutical bulletin, 37, 3, 490, 492, PHARMACEUTICAL SOC JAPAN, 2014年03月, ［査読有り］
英語, 研究論文（学術雑誌）, The aim of this study was to determine the effect of interaction between tegafur (FT) and epigallocatechin-3-gallate (EGCG) on the expression of alpha-defensins (HD-5: human alpha-defensin 5, HD-6: human alpha-defensin 6) by using a Caco-2 cell line as a model of human intestinal epithelial cells. This is the first study in which the effect of interaction of an oral anticancer drug and functional food on the innate immune system was examined. alpha-Defensins are abundant constituents of mouse and human paneth cells and play a role in the innate immune system in intestine. We detected HD-5 and HD-6 mRNA in Caco-2 cells and evaluated the effects of FT and EGCG on these mRNA levels. HD-5 and HD-6 mRNA levels were decreased by exposure to FT. Production of reactive oxygen species (ROS) was induced by exposure to FT as well as H2O2 exposure, and EGCG suppressed FT-induced production of ROS. Furthermore, FT-induced decrease in HD-5 and HD-6 mRNA levels was almost completely suppressed by EGCG. These results indicate that EGCG restored the decrease of alpha-defensins induced by FT at the transcriptional level in Caco-2 cells, suggesting that EGCG can be used as adjunctive therapy in chemotherapy.

Alteration of Pharmacokinetics of Grepafloxacin in Type 2 Diabetic Rats
Meguho Watanabe, Masaki Kobayashi, Jiro Ogura, Natsuko Takahashi, Hiroaki Yamaguchi, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 17, 1, 25, 33, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2014年, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose. Patients with type 2 diabetes are generally treated with various pharmacological compounds and are exposed to a high risk of drug-drug interactions. However, alterations of pharmacokinetics in a type 2 diabetes model have been obscure. The present study was undertaken to investigate the effects of type 2 diabetes on the pharmacokinetics of the fluoroquinolone grepafloxacin (GPFX) and the expression level of P-glycoprotein (P-gp), one of the drug efflux transporters. Methods. We used Goto-Kakizaki (GK) rats, a lean model of type 2 diabetes. Plasma concentration and intestinal, renal, and biliary clearance of GPFX were measured after intravenous and intraintestinal administration in Wistar and GK rats. Real-time PCR and Western blotting were used to assess mRNA and protein expression levels. Results. We found a significant increase in the plasma concentrations of GPFX at 90, 120 and 240 minutes after intraintestinal administration in GK rats compared with the concentrations in Wistar rats but not after intravenous administration. The increase in plasma GPFX concentration was associated with reduction in jejunal clearance of GPFX caused by a decrease in secretory transport of GPFX. However, there was no correlation between the decrease in secretory transport of GPFX and P-gp expression level. Conclusion. Type 2 diabetic conditions alter P-gp function as well as expression level and correlate poorly with each other.

Intestinal P-glycoprotein Expression is Multimodally Regulated by Intestinal Ischemia-Reperfusion
Yusuke Terada, Jiro Ogura, Takashi Tsujimoto, Kaori Kuwayama, Takahiro Koizumi, Shunichi Sasaki, Hajime Maruyama, Masaki Kobayashi, Hiroaki Yamaguchi, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 17, 2, 266, 276, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2014年, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose. Reactive oxygen species (ROS) have multiple physiological effects that are amount-dependent. ROS are one of the causes of intestinal ischemia-reperfusion (I/R) injury. In this study, we investigated whether the amount of ROS and the degree of intestinal I/R injury affect the expression level of P-glycoprotein (P-gp). Methods. We used hydrogen peroxide (H2O2) as ROS in in vitro experiments. Intestinal I/R model rats, which were subjected 15-min ischemia (I/R-15), were used in in vivo experiments. Results. P-gp expression in Caco-2 cells was increased in response to 1 mu M of H2O2 but decreased upon exposure to 10 mM of H2O2. We previously reported that P-gp expression is decreased after intestinal I/R with 30-min ischemia (I/R-30), which time a large amount of ROS is generated. I/R-15 induced slightly less mucosal and oxidative injury than did I/R-30. P-gp expression in the jejunum was increased at 1 h after I/R-15, and ileal paracellular permeability was increased. The blood concentration of tacrolimus, a P-gp substrate, was lower during 0-20 min but was higher during 40-90 min post-administration compared with that in the sham-operated rats. P-gp expression in the ileum was decreased at 6 h after I/R-15, due to abnormal localization of P-gp, resulting in a high blood tacrolimus concentration in rats reperfused for 6 h. Conclusions. ROS multimodally regulate P-gp expression depending on its amount. This is important for understanding the pattern of P-gp expression after intestinal I/R.

Effect of oxidative stress on expression and function of human and rat organic anion transporting polypeptides in the liver
Takashi Tsujimoto, Jiro Ogura, Kaori Kuwayama, Takahiro Koizumi, Shunichi Sasaki, Yusuke Terada, Masaki Kobayashi, Hiroaki Yamaguchi, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 458, 2, 262, 271, ELSEVIER SCIENCE BV, 2013年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Reactive oxygen species (ROS) have physiological function and involve alteration of physical state. However, it is not clear effect of oxidative stress on pharmacokinetics. Organic anion transporting polypeptides (human: OATPs, rodent: Oatps) are important for uptake of endogenous and exogenous compounds into hepatocytes. Thus, alteration of OATPs/Oatps expression level may affect pharmacokinetics of various drugs. In this study, we investigated the alteration of OATPs/Oatps expression levels and function by oxidative stress, and the effect of alteration of those on pharmacokinetics of a typical OATPs/Oatps substrate pravastatin. OATPs/Oatps expression levels and function were altered by H2O2-induced oxidative stress in in vitro experiments. The alteration of Oatps expression by oxidative stress also occurred in in vivo experiments. Oatp1a1, Oatp1a4 and Oatp1b2 expression in the liver were decreased in rats fed powdery diet containing 2% inosine, which induces oxidative stress through activation of xanthine oxidase, for 1 day. The decrease in Oatps expression levels by oxidative stress caused the suppression of pravastatin uptake to the liver, and resulted in high plasma concentration of pravastatin and low biliary excretion. In conclusion, oxidative stress induces alteration of OATPs/Oatps expression and function in hepatocytes, resulting in alteration of pharmacokinetics of their substrates. (C) 2013 Elsevier B.V. All rights reserved.

Megalin Contributes to Kidney Accumulation and Nephrotoxicity of Colistin
Takahiro Suzuki, Hiroaki Yamaguchi, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Ken Iseki
ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, 57, 12, 6319, 6324, AMER SOC MICROBIOLOGY, 2013年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Interest has recently been shown again in colistin because of the increased prevalence of infections caused by multidrug-resistant Gram-negative bacteria. Although the potential for nephrotoxicity is a major dose-limiting factor in colistin use, little is known about the mechanisms that underlie colistin-induced nephrotoxicity. In this study, we focused on an endocytosis receptor, megalin, that is expressed in renal proximal tubules, with the aim of clarifying the role of megalin in the kidney accumulation and nephrotoxicity of colistin. We examined the binding of colistin to megalin by using a vesicle assay. The kidney accumulation, urinary excretion, and concentrations in plasma of colistin in megalin-shedding rats were also evaluated. Furthermore, we examined the effect of megalin ligands and a microtubule-depolymerizing agent on colistin-induced nephrotoxicity. We found that cytochrome c, a typical megalin ligand, inhibited the binding of colistin to megalin competitively. In megalin-shedding rats, renal proximal tubule colistin accumulation was decreased (13.5 +/- 1.6 and 21.3 +/- 2.6 mu g in megalin-shedding and control rats, respectively). Coadministration of colistin and cytochrome c or albumin fragments resulted in a significant decrease in urinary N-acetyl-beta-D-glucosaminidase (NAG) excretion, a marker of renal tubular damage (717.1 +/- 183.9 mU/day for colistin alone, 500.8 +/- 102.4 mU/ day for cytochrome c with colistin, and 406.7 +/- 156.7 mU/ day for albumin fragments with colistin). More-over, coadministration of colistin and colchicine, a microtubule-depolymerizing agent, resulted in a significant decrease in urinary NAG excretion. In conclusion, our results indicate that colistin acts as a megalin ligand and that megalin plays a key role in the accumulation in the kidney and nephrotoxicity of colistin. Megalin ligands may be new targets for the prevention of colistininduced nephrotoxicity.

Intestinal ischemia-reperfusion suppresses biliary excretion of hepatic organic anion transporting polypeptides substrate
Hajime Maruyama, Jiro Ogura, Asuka Fujikawa, Yusuke Terada, Takashi Tsujimoto, Takahiro Koizumi, Kaori Kuwayama, Masaki Kobayashi, Hiroaki Yamaguchi, Ken Iseki
Journal of Pharmacy and Pharmaceutical Sciences, 16, 5, 722, 731, 5, 2013年11月17日, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose. Intestinal ischemia-reperfusion (I/R) causes gut dysfunction and promotes multi-organ failure. The liver and kidney can be affected by multi-organ failure after intestinal I/R. Organic anion transporting polypeptides (OATPs) and organic anion transporters (OATs) are recognized in a broad spectrum from endogenous compounds to xenobiotics, including clinically important drugs. Therefore, it is important for understanding the pharmacokinetics to obtain evidence of alterations in OATPs and OATs expression and transport activities. In the present study, we investigated the expression of rat Oatps and Oats after intestinal I/R. Methods. We used intestinal ischemia-reperfusion (I/R) model rats. Real-time PCR and Western blotting were used to assess mRNA and protein expression levels. Plasma concentration and biliary excretion of sulfobromophthalein (BSP), which is used as a model compound of organic anion drugs, were measured after intravenous administration in intestinal I/R rats. Results. Although Oat1 and Oat3 mRNA levels were not altered in the kidney, Oatp1a1, Oatp1b2 and Oatp2b1 mRNA levels in the liver were significantly decreased at 1-6 h after intestinal I/R. Moreover, Oatp1a1 and Oatp2b1 protein expression levels were decreased at 1 h after intestinal I/R. Plasma concentration of BSP, which is a typical substrate of Oatps, in intestinal I/R rats reperfused 1 h was increased than that in sham-operated rats. Moreover, the area under the concentration-time curve (AUC0-90) in intestinal I/R rats reperfused 1 h was significantly increased than that in sham-operated rats. The total clearance (CLtot) and the biliary clearance (CLbile) in intestinal I/R rats reperfused 1 h were significantly decreased than those in sham-operated rats. Conclusions. Oatp1a1 and Oatp2b1 expression levels are decreased by intestinal I/R. The decreases in these transporters cause alteration of pharmacokinetics of organic anion compound. The newly found influence of intestinal I/R on the expression and function of Oatps may be a key to perform appropriate drug therapy.

Contribution of multidrug resistance-associated proteins (MRPs) to the release of prostanoids from A549 cells
Ayako Furugen, Hiroaki Yamaguchi, Nobuaki Tanaka, Narumi Shiida, Jiro Ogura, Masaki Kobayashi, Ken Iseki
PROSTAGLANDINS & OTHER LIPID MEDIATORS, 106, 37, 44, ELSEVIER SCIENCE INC, 2013年10月, ［査読有り］
英語, 研究論文（学術雑誌）, Previous studies indicated that several members of the multidrug resistance-associated protein (MRP) family mediate the transport of prostanoids. However, theimportance of MRPs in the release process of prostanoids has not been fully elucidated. In this study, we investigated the contribution of MRPs, including MRP1, MRP2, and MRP4, to the release process of the prostanoids from human lung adenocarcinoma epithelial A549 cells. The extracellular levels of PGE(2), PGF(2 alpha), and TXB2 (a metabolite of TXA(2)) were decreased by treatment with MRP inhibitors (dipyridamole, MK571, and probenecid). The studies using membrane vesicle suggest that the effects of the inhibitors were in part by inhibiting MRP4 function. The effects of knockdown of each MRP (MRP1, MRP2, and MRP4) were also investigated. The extracellular levels of PGE(2) and PGF(2 alpha) were significantly decreased after MRP4 knockdown. Our results suggest that MRPs including MRP4 contribute the release process of prostanoids in A549 cells. (C) 2013 Elsevier Inc. All rights reserved.

Regulation of the Expression and Activity of Glucose and Lactic Acid Metabolism-Related Genes by Protein Kinase C in Skeletal Muscle Cells
Sho Otake, Masaki Kobayashi, Katsuya Narumi, Shotaro Sasaki, Yurika Kikutani, Ayako Furugen, Meguho Watanabe, Natsuko Takahashi, Jiro Ogura, Hiroaki Yamaguchi, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 36, 9, 1435, 1439, PHARMACEUTICAL SOC JAPAN, 2013年09月, ［査読有り］
英語, 研究論文（学術雑誌）, Protein kinase C (PKC) modulators are very attractive therapeutic targets in cancer. Since most cancer cells display increased glycolysis, elucidations of the effects of PKC activation on glycolysis is necessary for the development of effective medicine. In the present study, to clarify the role of PKC in the regulation of glycolysis, we examined the effect of phorbol 12-myristate 13-acetate (PMA), a PKC activator, on the expression and activity of glucose and lactic acid metabolism-related genes in human rhabdomyosarcoma cells (RD cells). In parallel to increases in glucose uptake and mRNA levels of glucose transporters (GLUTs) induced by PMA treatment for 6h, the hexokinase (HK) mRNA level and activity were also significantly increased in RD cells. On the other hand, a significant increase in lactate dehydrogenase (LDH) mRNA level and activity was seen when the cells were incubated with PMA for 24h, but not for 6 or 12h, and was associated with lactic acid production. These effects by PMA treatment were markedly suppressed by Bisindolylmaleimide (BIM), a PKC inhibitor. Furthermore, chetomin, a hypoxia-inducible factor 1 (HIF-1) inhibitor, completely abrogated the increment of LDH mRNA level and activity as well as monocarboxylate transporter (MCT) 4, a lactic acid efflux transporter. In conclusion, we found that HK and LDH activity induced by PKC activation was associated with the glucose uptake and lactic acid level and that LDH and MCT4 are modulated by a common factor, HIF-1.

Regulation of multidrug resistance protein 2 (MRP2, ABCC2) expression by statins: Involvement of SREBP-mediated gene regulation
Masaki Kobayashi, Keisuke Gouda, Ikumi Chisaki, Koji Asada, Jiro Ogura, Natsuko Takahashi, Toru Konishi, Yusuke Koshida, Shotaro Sasaki, Hiroaki Yamaguchi, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 452, 1-2, 36, 41, ELSEVIER SCIENCE BV, 2013年08月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Multidrug resistance protein 2 (MRP2, ABCC2) is localized to the apical membrane of hepatocytes and played an important role in the biliary excretion of a broad range of endogenous and xenobiotic compounds and drugs, such as pravastatin. However, the effects of statins on MRP2 in the liver and the precise mechanisms of their actions have been obscure. The goal of this study was to determine the regulatory molecular mechanism for statin-induced MRP2 expression in hepatocytes. In vitro and in vivo studies suggested that pitavastatin increased MRP2 expression. Pitavastatin promoted liver X receptor (LXR) alpha/beta translocation from the cytosol to nuclei, resulting in LXR activation. Deletion and mutational analysis suggested that the potential sterol regulatory element (SRE) played a major role in the observed modulation of MRP2 expression by pitavastatin. Furthermore pitavastatin increased the protein-DNA complex, and when SRE was mutated, stimulation of the protein-DNA complex by pitavastatin was decreased. It was demonstrated that pitavastatin upregulated MRP2 expression by an SREBP regulatory pathway in hepatocytes and that the actions of statins may lead to improve the biliary excretion of MRP2 substrates. (C) 2013 Elsevier B.V. All rights reserved.

Crucial Residue Involved in L-Lactate Recognition by Human Monocarboxylate Transporter 4 (hMCT4)
Shotaro Sasaki, Masaki Kobayashi, Yuya Futagi, Jiro Ogura, Hiroaki Yamaguchi, Natsuko Takahashi, Ken Iseki
PLOS ONE, 8, 7, e67690, PUBLIC LIBRARY SCIENCE, 2013年07月, ［査読有り］
英語, 研究論文（学術雑誌）, Background: Monocarboxylate transporters (MCTs) transport monocarboxylates such as lactate, pyruvate and ketone bodies. These transporters are very attractive therapeutic targets in cancer. Elucidations of the functions and structures of MCTs is necessary for the development of effective medicine which targeting these proteins. However, in comparison with MCT1, there is little information on location of the function moiety of MCT4 and which constituent amino acids govern the transport function of MCT4. The aim of the present work was to determine the molecular mechanism of L-lactate transport via hMCT4.
Experimental approach: Transport of L-lactate via hMCT4 was determined by using hMCT4 cRNA-injected Xenopus laevis oocytes. hMCT4 mediated L-lactate uptake in oocytes was measured in the absence and presence of chemical modification agents and 4,4'-diisothiocyanostilbene-2,2'-disulphonate (DIDS). In addition, L-lactate uptake was measured by hMCT4 arginine mutants. Immunohistochemistry studies revealed the localization of hMCT4.
Results: In hMCT4-expressing oocytes, treatment with phenylglyoxal (PGO), a compound specific for arginine residues, completely abolished the transport activity of hMCT4, although this abolishment was prevented by the presence of L-lactate. On the other hand, chemical modifications except for PGO treatment had no effect on the transport activity of hMCT4. The transporter has six conserved arginine residues, two in the transmembrane-spanning domains (TMDs) and four in the intracellular loops. In hMCT4-R278 mutants, the uptake of L-lactate is void of any transport activity without the alteration of hMCT4 localization.
Conclusions: Our results suggest that Arg-278 in TMD8 is a critical residue involved in substrate, L-lactate recognition by hMCT4.

Rapid Stimulating Effect of the Antiarrhythmic Agent Amiodarone on Absorption of Organic Anion Compounds
Masahiro Segawa, Jiro Ogura, Satoru Seki, Shirou Itagaki, Natsuko Takahashi, Masaki Kobayashi, Takeshi Hirano, Hiroaki Yamaguchi, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 28, 3, 178, 186, JAPANESE SOC STUDY XENOBIOTICS, 2013年06月, ［査読有り］
英語, 研究論文（学術雑誌）, In a clinical setting, changes in pharmacokinetics due to drug-drug interactions can often directly affect the therapeutic safety and efficacy of drugs. Recently, interest has been shown in drug-drug interactions in the intestine. It is now recognized that changes in the functions of drug transporters substantially influence the absorption of administered drugs from the intestine. Amiodarone (AMD) is a potent drug used in the treatment of serious supraventricular and ventricular tachyarrhythmias. Despite its potent pharmacological effects, its wide clinical use is precluded by drug-drug interactions. In this study, we characterized the transporter function between AMD and various compounds in human intestinal model Caco-2 cells. AMD significantly and rapidly increased the uptake of [H-3]estrone-3-sulfate (E-3-S) for 5 min. The apical-to-basal transport of [H-3]E-3-S was significantly increased by AMD. The AMD-stimulated [H-3]E-3-S uptake was inhibited by organic anion transporting polypeptide (OATP) substrates. Caco-2 cells treated with AMD showed increased OATP2B1 expression on the cell surface. AMD also increased the absorption of sulfobromophthalein (ESP), which is a typical organic anion compound, and the expression level of Oatp2b1 at the membrane in in vivo experiments. The results indicate that AMD induces OATP2B1/Oatp2b1 expression at the membrane in the intestine and enhances absorption of organic anion compounds.

Quantitative determination of paclitaxel and its metabolites, 6α-hydroxypaclitaxel and p-3'-hydroxypaclitaxel, in human plasma using column-switching liquid chromatography/tandem mass spectrometry.
Yamaguchi H, Fujikawa A, Ito H, Tanaka N, Furugen A, Miyamori K, Takahashi N, Ogura J, Kobayashi M, Yamada T, Mano N, Iseki K
Biomedical chromatography : BMC, 27, 4, 539, 44, 2013年04月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, A column-switching liquid chromatography/electrospray ionization tandem mass spectrometry to determine paclitaxel and its metabolites, 6α-hydroxypaclitaxel and p-3'-hydroxypaclitaxel, in human plasma was developed. The analytical system had a Shim-Pack MAYI-ODS (10 × 4.6 mm i.d.) trapping column with deproteinization ability that concentrates analytes and removes water-soluble components. This method covered a linearity range of 5-5000 ng/mL of concentrations in plasma for paclitaxel, a range of 0.87-870 ng/mL for 6α-hydroxypaclitaxel and a range of 0.87-435 ng/mL for p-3'-hydroxypaclitaxel. The intra-day precision and inter-day precision of analysis were less than 11.1%, and the accuracy was within ±14.4% at concentrations of 5, 50, 500 and 5000 ng/mL for paclitaxel, 0.87, 8.7, 87 and 870 ng/mL for 6α-hydroxypaclitaxel, and 0.87, 8.7, 87 and 435 ng/mL for p-3'-hydroxypaclitaxel. The total run time was 30 min. Our method was successfully applied to clinical pharmacokinetic investigation.

Quantification of intact carboplatin in human plasma ultrafitrates using hydrophilic interaction liquid chromatography-tandem mass spectrometry and its application to a pharmacokinetic study
Hajime Ito, Hiroaki Yamaguchi, Asuka Fujikawa, Narumi Shiida, Nobuaki Tanaka, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 917, 18, 23, ELSEVIER SCIENCE BV, 2013年02月, ［査読有り］
英語, 研究論文（学術雑誌）, Carboplatin is a platinum agent that is used for treatment of non-small-cell lung cancer and ovarian cancer. A sensitive and selective analytical method for the quantification of carboplatin in human plasma ultrafiltrates using liquid chromatography-tandem mass spectrometry was developed. Human plasma ultrafiltrates were precipitated by acetonitrile containing carboplatin-d4 as an internal standard and were further diluted with acetonitrile. Chromatographic separation was performed on a Accucore HILIC (50 mm x 2.1 mm i.d., 2.6 mu m) column using mobile phase (acetonitrile-water-acetic acid = 90:10:0.1, v/v/v) at the flow rate of 0.2 mL/min. Detection was performed on electrospray ionization triple quadrupole tandem mass spectrometer using low-energy collision induced dissociation (CID-MS/MS) analysis operating in the selected reaction monitoring (SRM) scan mode. The lower limit of quantification for carboplatin was 0.025 mu g/mL. This method covered a linearity range of 0.025-50 mu g/mL. The intra-day precision and inter-day precision (R.S.D.) ranged from 1.5 to 4.3%, and the accuracy (R.E.) was within +/- 2.9%. The present method was applied to a clinical pharmacokinetic study of carboplatin in a cancer patient. (c) 2012 Elsevier B.V. All rights reserved.

Renal uptake of substrates for organic anion transporters Oat1 and Oat3 and organic cation transporters Oct1 and Oct2 is altered in rats with adenine-induced chronic renal failure
Hiroki Komazawa, Hiroaki Yamaguchi, Kazuhiro Hidaka, Jiro Ogura, Masaki Kobayashi, Ken Iseki
Journal of Pharmaceutical Sciences, 102, 3, 1086, 1094, John Wiley and Sons Inc., 2013年, ［査読有り］
英語, 研究論文（学術雑誌）, Chronic renal failure (CRF) leads to decreased drug renal clearance and glomerular filtration rate. However, little is known about renal tubular excretion and reabsorption in CRF. We examined transport activity of renal transporters using rats with adenine-induced CRF. We examined the effect of adenine-induced CRF on mRNA level, protein expression of transporters expressed in kidney by real-time polymerase chain reaction, and western blotting. In vivo kidney uptake clearances of benzylpenicillin and metformin, which are typical substrates for renal organic anion transporters Oat1 and Oat3 and organic cation transporters Oct1 and Oct2, respectively, were evaluated. Protein and mRNA expression levels of Oat1, Oat 3, Oct1, and Oct2 were significantly decreased in adenine-induced CRF rats. On the contrary, levels of P-glycoprotein and Mdr1b mRNA were significantly increased in adenine-induced CRF rats. The mRNA expression levels of Oatp4c1, Mate1, Urat1, Octn2, and Pept1 were significantly decreased. Kidney uptake clearance of benzylpenicillin and that of metformin were significantly decreased in adenine-induced CRF rats. Also, serum from CRF rats did not affect Oat1, Oat3, Oct1, and Oct2 function. In conclusion, our results indicate that adenine-induced CRF affects renal tubular handling of drugs, especially substrates of Oat1, Oat3, Oct1, and Oct2. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association.

A full validated hydrophilic interaction liquid chromatography-tandem mass spectrometric method for the quantification of oxaliplatin in human plasma ultrafiltrates
Hajime Ito, Hiroaki Yamaguchi, Asuka Fujikawa, Nobuaki Tanaka, Ayako Furugen, Kazuaki Miyamori, Natsuko Takahashi, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
JOURNAL OF PHARMACEUTICAL AND BIOMEDICAL ANALYSIS, 71, 99, 103, ELSEVIER SCIENCE BV, 2012年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Oxaliplatin is a platinum agent that is used for treatment of colorectal cancer. A sensitive and selective hydrophilic interaction liquid chromatography tandem mass spectrometric method for the quantification of oxaliplatin was developed. Human plasma ultrafiltrates were precipitated by acetonitrile containing carboplatin as an internal standard and further diluted with acetonitrile. Chromatographic separation of oxaliplatin and the internal standard was achieved with a column modified with phosphorylcholine and an isocratic mobile phase (acetonitrile/water/acetic acid = 90:10:0.1, v/v/v) at the flow rate of 0.2 mL/min. The lower limit of quantification for oxaliplatin was 25 ng/mL. The linearity range of the method was from 25 to 5000 ng/mL. The intra-day precision and inter-day precision (RSD) ranged from 0.8 to 6.1%, and the accuracy (RE) was within +/- 4.5%. The extraction recoveries from human plasma ultrafiltrates were 83.6-91.6%, and ion suppression caused by matrix components was 86.7-88.5% at three different levels, respectively. This method was applied to a clinical pharmacokinetic study of oxaliplatin in a cancer patient. The maximum concentration of colorectal cancer patient administered oxaliplatin was 1650 ng/mL. (C) 2012 Elsevier B.V. All rights reserved.

Regulation of human monocarboxylate transporter 4 in skeletal muscle cells: The role of protein kinase C (PKC)
Katsuya Narumi, Masaki Kobayashi, Sho Otake, Ayako Furugen, Natsuko Takahashi, Jiro Ogura, Shirou Itagaki, Takeshi Hirano, Hiroaki Yamaguchi, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 428, 1-2, 25, 32, ELSEVIER SCIENCE BV, 2012年05月, ［査読有り］
英語, 研究論文（学術雑誌）, In the present study, to clarify the role of protein kinase C (PKC) in the regulation of monocarboxylate transporter 4 (MCT4) expression, we examined the regulation mechanism of MCT4 expression in human rhabdomyosarcoma (RD) cells, an in vitro skeletal muscle model. Exposure of RD cells to PMA, a PKC activator, for 24 h resulted in a two-fold increase in the amount of lactic acid in the growth medium. In parallel to an increase in lactic acid release from RD cells, the level of MCT4 mRNA and protein were also significantly increased in RD cells. A PKC inhibitory study indicated that PMA-induced stimulation of MCT4 expression can be mediated through a novel PKC isoform, especially PKC delta. Moreover, rottlerin, a selective PKC delta inhibitor, decreased PMA-induced MCT4 promoter activity. Deletion and mutational analysis suggested that the potential hypoxia-response elements (HREs) played a major role in the observed modulation of MCT4 expression by PMA. Furthermore, we found that small interfering RNA (siRNA)-mediated knockdown of hypoxia-inducible factor 1 alpha (HIF-1 alpha) significantly inhibited PMA-induced MCT4 promoter activity. Our results show that the effects of PMA on MCT4 expression are mediated through an indirect pathway partially involving PKC delta and HIF-1 alpha transcription factor. (C) 2012 Published by Elsevier B.V.

Effects of Acidification and Alkalinization Agent on Statins-induced Muscle Toxicity
Masaki Kobayashi, Kazuhiro Hidaka, Ikumi Chisaki, Natsuko Takahashi, Jiro Ogura, Shirou Itagaki, Takeshi Hirano, Hiroaki Yamaguchi, Ken Iseki
YAKUGAKU ZASSHI-JOURNAL OF THE PHARMACEUTICAL SOCIETY OF JAPAN, 132, 5, 609, 615, PHARMACEUTICAL SOC JAPAN, 2012年05月, ［査読有り］, ［筆頭著者］
日本語, 研究論文（学術雑誌）, The aim of this study was to determine the effects of alpha-cyano-4-hydroxycinnamic acid (CHC), a lactate efflux inhibitor, and citrate, an alkaline reagent, on statin-induced muscle injury using a human prototypic embryonal rhabdomyosarcoma cell line (RD) as a model of in vitro skeletal muscle and on statin-induced muscle damage in an in vivo study. Statin-induced reduction of cell viability and apoptosis was measured by the 3-(4,5-dimethylthiazol-2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT) assay and caspase assay. In an in vivo study, plasma creatine phosphokinase (CPK) level was examined in cerivastatin-treated rats. CHC increased growth inhibition of RD cells induced by cerivastatin, a lipophilic statin, but not these induced by pravastatin, a hydrophilic statin. On the other hand, citrate suppressed cerivastatin-, simvastatin- and atorvastatin-induced reduction of cell viability and caspase activation in RD cells. Moreover, citrate prevented cerivastatin-induced increase in CPK concentration in a concentration-dependent manner. This is first study to evaluate CHC or citrate-induced exacerbation or improvement of statin-induced muscle damage.

A rapid and sensitive LC/ESI-MS/MS method for quantitative analysis of docetaxel in human plasma and its application to a pharmacokinetic study
Hiroaki Yamaguchi, Asuka Fujikawa, Hajime Ito, Nobuaki Tanak, Ayako Furugen, Kazuaki Miyamori, Natsuko Takahashi, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 893, 157, 161, ELSEVIER SCIENCE BV, 2012年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Docetaxel is a taxane family antineoplastic agent widely employed in cancer chemotherapy. We developed a liquid chromatography/tandem mass spectrometry method for the determination of docetaxel in human plasma. Plasma samples were deproteinized by acetonitrile containing internal standard paclitaxel. Chromatographic separation was performed on a TSKgel ODS-100V 3 mu m (50 mm x 2.0 mm i.d.) column using a mobile phase composed of acetonitrile-methanol-water-formic acid (50:5:45:0.1, v/v/v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer with multi pie reaction monitoring (MRM) mode via electrospray ionization (ESI) source. This method covered a linearity range of 5-5000 ng/mL with the lower limit of quantification of 5 ng/mL. The intra-day precision and inter-day precision (R.S.D.) of analysis were less than 6.7%, and the accuracy (R.E.) was within +/- 9.0% at the concentrations of 5, 20, 200, and 2000 ng/mL. The total run time was 5.0 min. This method was successfully applied for clinical pharmacokinetic investigation. (C) 2012 Elsevier B.V. All rights reserved.

Involvement of Cholesterol Membrane Transporter Niemann-Pick C1-Like 1 in the Intestinal Absorption of Lutein
Yuki Sato, Risa Suzuki, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Toshihiro Noda, Satoshi Mizuno, Mitsuru Sugawara, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 15, 2, 256, 264, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2012年, ［査読有り］
英語, 研究論文（学術雑誌）, PURPOSE. Lutein is a carotenoid mainly found in green leafy vegetables and is located in the macula lutea in the human eye. Since humans cannot synthesize lutein de novo, it must be digested as food. The physiological importance of an orally administered compound depends on its interaction with target tissues. It is therefore important to clarify the absorption mechanism in the intestine. Cholesterol membrane transporters Niemann-Pick C1 Like 1 (NPC1L1) and scavenger receptor class B type 1 (SR-B1) are involved in the intestinal absorption of highly lipophilic compounds including cholesterol. Ezetimibe, a selective inhibitor of intestinal NPC1L1, is the widespread lipid-lowering agent. It is important to investigate the possibility of food-drug interactions in order to prevent undesirable and harmful clinical consequences. The aim of this work was to determine whether NPC1L1, SR-B1 and other transporters are involved in absorption of lutein. METHODS. Caco-2 cells were used for accumulation and permeability study of lutein. Lutein concentration was determined by an HPLC system. The cDNA of transporters was isolated from total RNA of Caco-2 cells, and the expression of these transporters was confirmed by RT-PCR (reverse transcription - polymerase chain reaction). RESULTS. Ezetimibe inhibited up to 40% of lutein accumulation by Caco-2 cell monolayers. Block lipid transport 1 (BLT-1), a selective chemical inhibitor of SR-B1, also inhibited lutein accumulation by Caco-2 cells. On the other hand, ATP-depletion reagents (sodium fluoride and sodium azide or carbonyl cyanide m-chlorophenylhydrazone) did not influence the accumulation or permeation of lutein significantly. CONCLUSIONS. The results show that lutein absorption is, at least in part, mediated by influx transporters NPC1L1 and SR-B1 rather than mediated by efflux transporters such as ABC (ATP-binding cassette) transporters.

Intestinal Ischemia-Reperfusion Increases Efflux for Uric Acid Via Paracellular Route in the Intestine, but Decreases that Via Transcellular Route Mediated by BCRP
Jiro Ogura, Kaori Kuwayama, Atsushi Takaya, Yusuke Terada, Takashi Tsujimoto, Takahiro Koizumi, Hajime Maruyama, Asuka Fujikawa, Natsuko Takahashi, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Hiroaki Yamaguchi, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 15, 2, 295, 304, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2012年, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose. Uric acid is thought to be one of the most important antioxidants in human biological fluids. Intestinal ischemia-reperfusion (I/R) is an important factor associated with high rates of morbidity and mortality. Reactive oxygen species (ROS) are responsible for intestinal I/R injury. The aim of this study was to clarify the efflux for uric acid from the intestine after intestinal I/R. Methods. We used intestinal ischemia-reperfusion (I/R) model rats. Serosal to mucosal flux for [C-14]-uric acid was assessed by using Ussing-type diffusion chambers. BCRP/Bcrp expression was assessed by Western blot analysis. Caco-2 cells were used for a model of the intestinal epithelium, and rotenone was used as a mitochondrial dysfunction inducer. Results. Serosal to mucosal flux for uric acid was increased after intestinal I/R, and that for mannitol was also increased. Ko143, which is a BCRP inhibitor, did not affect the uric acid transport. The decreasing uric acid transport mediated by Bcrp was caused by decrease in the level of Bcrp homodimer, bridged by an S-S bond. The suppression of Bcrp S-S bond formation was associated with mitochondrial dysfunction. Moreover, BCRP S-S bond formation activity was decreased by rotenone in Caco-2 cells. Conclusions. Serosal to mucosal flux for uric acid is significantly increased via the paracelluler route, but that via the transcellular route mediated by Bcrp is decreased after intestinal I/R. The decreasing uric acid flux mediated by Bcrp is caused by suppression of Bcrp S-S bond formation. This suppression of Bcrp S-S bond formation may be related to mitochondrial dysfunction.

The Decrease in Farnesoid X Receptor, Pregnane X Receptor and Constitutive Androstane Receptor in the Liver after Intestinal Ischemia-Reperfusion
Jiro Ogura, Yusuke Terada, Takashi Tsujimoto, Takahiro Koizumi, Kaori Kuwayama, Hajime Maruyama, Asuka Fujikawa, Atsushi Takaya, Masaki Kobayashi, Shirou Itagaki, Natsuko Takahashi, Takeshi Hirano, Hiroaki Yamaguchi, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 15, 5, 616, 631, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2012年, ［査読有り］
英語, 研究論文（学術雑誌）, Purpose. Intestinal ischemia-reperfusion (I/R) damages remote organs, including the liver, and promotes multi-organ failure (MOF). However, the molecular mechanisms underlying acute liver injury after intestinal I/R have not been completely elucidated. Farnesoid X receptor (FXR), pregnane X receptor (PXR) and constitutive androstane receptor (CAR) regulate metabolizing enzymes and transporters, and coordinately prevent hepatotoxicity reflecting an inability of appropriate excretion of endogenous toxic compounds. In this study, we assessed FXR, PXR and CAR expression levels and their localization levels in nuclei in the liver after intestinal I/R. We also investigated the effect of IL-6 on FXR, PXR and CAR expression levels and their localization levels in nuclei in in vitro experiments. Methods. We used intestinal I/R model rats. Moreover, HepG2 cells were used in in vitro study. Real-time PCR and Western blotting were used to assess mRNA and protein expression levels. Nuclear receptor localization in nuclei was analyzed by Western blotting using nuclear extracts. Results. FXR and PXR expression levels began to be decreased at 3 h, and FXR, PXR and CAR expression levels were decreased at 6 h after intestinal I/R. The localization levels of FXR, PXR and CAR in nuclei began to be decreased at 3 h, and all of them were decreased at 6 h after intestinal I/R. In HepG2 cells, FXR, PXR and CAR expression levels were decreased by 0.5-1 ng/mL, 0.5-100 ng/mL and 100 ng/mL IL-6 treatment for 24 h, respectively. FXR, PXR and CAR localization levels in nuclei were suppressed by 0.5-10 ng/mL, 10-100 ng/mL and 10-100 ng/mL IL-6 treatment for 24 h, respectively. Conclusions. FXR, PXR and CAR expression levels are decreased in the liver after intestinal I/R. IL-6 is one of main causes the decreases in expressions of these receptors.

Quantification of intracellular and extracellular prostanoids stimulated by A23187 by liquid chromatography/electrospray ionization tandem mass spectrometry
Ayako Furugen, Hiroaki Yamaguchi, Nobuaki Tanaka, Hajime Ito, Kazuaki Miyamori, Asuka Fujikawa, Natsuko Takahashi, Jiro Ogura, Masaki Kobayashi, Takehiro Yamada, Nariyasu Mano, Ken Iseki
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 879, 30, 3378, 3385, ELSEVIER SCIENCE BV, 2011年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Prostanoids are bioactive substances that contribute to various biological and pathological processes. To evaluate both extracellular and intracellular levels of prostanoids at the same time, we developed methods for quantification of extracellular and intracellular levels of prostanoids, including prostaglandin E(2) (PGE(2)), PGD(2), PGD(2 alpha), 6-keto PGF(1 alpha), and TXB(2), in cultured cells using liquid chromatography/tandem mass spectrometry (LC/MS/MS), and we validated the LC/MS/MS methods. A solid-phase extraction cartridge was used for extraction of prostanoids. The prostanoids were separated by a C(18) column with an isocratic flow of acetonitrile/water/acetic acid (40:60:0.1, v/v/v). Calibration curves of extracellular measurement for the prostanoids were linear in the range from 0.1 to 100 ng/mL (r(2) > 0.999), and those of intracellular measurement were linear in the range from 0.05 to 50 ng (r(2) > 0.999). Validation assessment showed that both methods of extracellular and intracellular measurements were highly reliable with good accuracy and precision.
We also applied the methods to human airway epithelial Calu-3 cells and human lung adenocarcinoma epithelial A549 cells. (C) 2011 Elsevier B.V. All rights reserved.

Protective Effect of Soy Isoflavone Genistein on Ischemia-Reperfusion in the Rat Small Intestine
Yuki Sato, Shirou Itagaki, Setsu Oikawa, Jiro Ogura, Masaki Kobayashi, Takeshi Hirano, Mitsuru Sugawara, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 34, 9, 1448, 1454, PHARMACEUTICAL SOC JAPAN, 2011年09月, ［査読有り］
英語, 研究論文（学術雑誌）, Ischemia-reperfusion (I/R) injury of the intestine is an important factor associated with high rates of morbidity and mortality. Intestinal I/R is a common clinical problem in the settings of severe burns, circulatory shock and strangulation ileus. Intestinal I/R damages remote organs and promotes multi-organ failure. It has been shown that enteral feeding before ischemic insults is beneficial for reducing organ injury and improving survival after intestinal I/R. In that study, the authors used a standard complex enteral diet and they suggested that it is important to find new nutrient formulas. Since reactive oxygen species are responsible for intestinal I/R injury, we focused on a dietary polyphenol, the soy isoflavone genistein. Genistein has a wide spectrum of biochemical and pharmacological activities. However, the possibility of a protective effect of genistein as enteral nutrition on I/R injury has not been investigated. We therefore investigated the protective effect of genistein on oxidative injury using intestinal I/R model rats. We found that genistein, which has combined antioxidant activity from radical scavenging, xanthine oxidase inhibition and chain-breaking effects, exhibits a protective effect on intestinal I/R injury. The results suggest that genistein, a soy isoflavone, has the possibility as a new nutrient formula of enteral feeding.

Protective effect of lutein after ischemia-reperfusion in the small intestine
Yuki Sato, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Toshihiro Noda, Satoshi Mizuno, Mitsuru Sugawara, Ken Iseki
FOOD CHEMISTRY, 127, 3, 893, 898, ELSEVIER SCI LTD, 2011年08月, ［査読有り］
英語, 研究論文（学術雑誌）, Lutein is a carotenoid mainly found in green leafy vegetables and is located in the macula lutea in the human eye. Since humans cannot synthesise lutein de novo, it must be digested as food. The physiological importance of an orally administered compound depends on its interaction with target tissues. There is little information about the effects of intake of lutein in tissues other than the eyes. The aim of this study was to clarify the protective effect of lutein against oxidative injury using ischemia-reperfusion (I/R) model rats and to determine the relationship between pharmacokinetics and antioxidant activity of lutein. Intestinal I/R was induced by 30-min occlusion of the superior mesenteric artery followed by 60-min reperfusion. After 60 min of reperfusion, intestinal tissue was Used for analysis of Evans blue dye extravasation, lipid peroxidation and myeloperoxidase activity. Lutein administered before I/R had a significant protective effect against oxidative injury. (C) 2011 Elsevier Ltd. All rights reserved.

Regulation mechanism of ABCA1 expression by statins in hepatocytes
Masaki Kobayashi, Keisuke Gouda, Ikumi Chisaki, Manami Ochiai, Shirou Itagaki, Ken Iseki
EUROPEAN JOURNAL OF PHARMACOLOGY, 662, 1-3, 9, 14, ELSEVIER SCIENCE BV, 2011年07月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, ATP-binding cassette transporter A1 (ABCA1) is predicted to be involved in the control of apolipoprotein AI-mediated cholesterol efflux: biosynthesis of high-density lipoprotein (HDL). However, the effects of HMG-CoA reductase inhibitors (statins) on ABCA1 in the liver and the precise mechanisms of their actions have been obscure. The aims of this study were to determine whether statins (atorvastatin (Ato) and pitavastatin (Pit)) affect hepatic ABCA1 expression and to clarify the mechanisms of their actions using HepG2 cells and the rat liver. We examined alterations in mRNA and protein levels of ABCA1 and peroxisome proliferator-activated receptors (PPARs) by quantitative real-time polymerase chain reaction (PCR) and Western blot analysis, respectively. In vitro and in vivo studies suggested that Pit increases ABCA1 mRNA level, but not Ato. Pit greatly increased Abca1 mRNA level and also increased the amount of plasma HDL and the mRNA level of PPAR alpha. Clofibrate (PPAR alpha agonist) increased ABCA1 expression in HepG2 cells and rat primary hepatocytes more than did PPAR beta/delta and gamma agonists. Pit-induced ABCA1 expression alteration was blocked by GW6471 (PPAR alpha antagonist) and by PPAR alpha knockdown. In this study, we demonstrated that Pit affect ABCA1 expression via PPAR alpha in hepatocytes. The strategy to target a PPAR alpha agonist in the liver can lead to increases in ABCA1 expression and HDL level. (C) 2011 Elsevier B.V. All rights reserved.

Pharmacokinetic properties of lutein emulsion after oral administration to rats and effect of food intake on plasma concentration of lutein
Yuki Sato, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Toshihiro Noda, Satoshi Mizuno, Mitsuru Sugawara, Ken Iseki
BIOPHARMACEUTICS & DRUG DISPOSITION, 32, 3, 151, 158, WILEY-BLACKWELL, 2011年04月, ［査読有り］
英語, 研究論文（学術雑誌）, Lutein is a carotenoid found mainly in green leafy vegetables and is located in the macula lutea in the human eye. An intake of lutein as food is needed since humans cannot synthesize it de novo. Although lutein has received much attention recently due to its antioxidant activities, little information about the pharmacokinetic properties of lutein is available. Lutein emulsion formulation was used and the pharmacokinetics of lutein emulsion after oral administration to rats was investigated. The bioavailability of lutein using this formulation was calculated to be 5.20%. It was found that a large amount of lutein was accumulated in the intestinal mucosa. The absorption of orally administered compounds in the intestine can be enhanced by interaction with food or food components. Thus, the effect of food intake on the intestinal absorption of lutein was investigated. The plasma concentration of lutein after oral administration of the emulsion formulation was improved significantly by food intake. It is possible that the absorption of lutein in the intestine is improved significantly by some food components. Bile acids may also play important roles in the intestinal absorption of lutein since the absorption of lipophilic compounds such as cholesterol is related to bile acids. The results of these studies should contribute to an improvement of lutein absorption and provide important information for obtaining more effective pharmacological effects of lutein. Copyright (C) 2011 John Wiley & Sons, Ltd.

In vitro and in vivo antioxidant properties of chlorogenic acid and caffeic acid
Yuki Sato, Shirou Itagaki, Toshimitsu Kurokawa, Jiro Ogura, Masaki Kobayashi, Takeshi Hirano, Mitsuru Sugawara, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 403, 1-2, 136, 138, ELSEVIER SCIENCE BV, 2011年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Dietary polyphenols are thought to be beneficial for human health as antioxidants. Coffee beans contain a common polyphenol, chlorogenic acid. Chlorogenic acid is the ester of caffeic acid and quinic acid. Although these polyphenols have received much attention, there is little evidence indicating a relationship between the effect and the rate of absorption. In this study, we focused on the beneficial effects of chlorogenic acid and caffeic acid, a major metabolite of chlorogenic acid. We carried out in vitro and in vivo experiments. In the in vitro study, caffeic acid had stronger antioxidant activity than that of chlorogenic acid. The uptake of chlorogenic acid by Caco-2 cells was much less than that of caffeic acid. The physiological importance of an orally administered compound depends on its availability for intestinal absorption and subsequent interaction with target tissues. We then used an intestinal ischemia-reperfusion model to evaluate antioxidant activities in vivo. We found that both chlorogenic acid and caffeic acid had effects on intestinal ischemia-reperfusion injury. Since caffeic acid has a stronger antioxidant activity than that of chlorogenic acid and chlorogenic acid is hydrolyzed into caffeic acid in the intestine, it is possible that caffeic acid plays a major role in the protective effect of chlorogenic acid against ischemia-reperfusion injury. (C) 2010 Elsevier B.V. All rights reserved.

AMP-activated protein kinase regulates the expression of monocarboxylate transporter 4 in skeletal muscle
Ayako Furugen, Masaki Kobayashi, Katsuya Narumi, Meguho Watanabe, Sho Otake, Shirou Itagaki, Ken Iseki
LIFE SCIENCES, 88, 3-4, 163, 168, PERGAMON-ELSEVIER SCIENCE LTD, 2011年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Aims: The aim of this study was to determine the effect of 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), an AMP-activated protein kinase (AMPK) activator, on monocarboxylate transporter 4 (MCT4) expression in rat skeletal muscle and a prototypic embryonal rhabdomyosarcoma cell line (RD cells).
Main methods: We examined the alteration in Glucose transporter 4 (GLUT4) and MCT4 mRNA levels by quantitative real-time PCR. Alteration in GLUT4 and MCT4 protein levels was examined by Western blotting.
Key findings: In an in vivo study. AICAR increased MCT4 mRNA and protein levels in a fiber-type specific manner. In an in vitro study, AICAR increased MCT4 mRNA and protein levels. Moreover, AICAR-induced MCT4 expression was blocked by Compound C, an AMPK inhibitor.
Significance: In this study, we found that AMPK activation induced expression of MCT4 in RD cells and rat skeletal muscle in a fiber-type specific manner. These results indicate the possible involvement of an AMPK-mediated pathway associated with MCT4 expression in skeletal muscle. (C) 2010 Elsevier Inc. All rights reserved.

Regulation of Monocarboxylate Transporter 1 in Skeletal Muscle Cells by Intracellular Signaling Pathways
Katsuya Narumi, Ayako Furugen, Masaki Kobayashi, Sho Otake, Shirou Itagaki, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 33, 9, 1568, 1573, PHARMACEUTICAL SOC JAPAN, 2010年09月, ［査読有り］
英語, 研究論文（学術雑誌）, Skeletal muscle is the major producer of lactic acid in the body, but its oxidative fibers also use lactic acid as a respiratory fuel. Monocarboxylate transporter (MCT) 1 has been suggested to play a major role in influx of L-lactic acid for oxidation. The regulation mechanism of MCT1 was characterized utilizing rhabdomyosarcoma cells as an in vitro skeletal muscle model. The uptake of L-lactic acid via MCT1 was studied in the presence of various intracellular regulatory pathways, including pathways mediated by protein kinases A, C and G (PKA, PKC and PKG), protein tyrosine kinase (PTK), and Ca2+/calmodulin modulators. The results showed that PKG-, PTK-, and Ca2+/calmodulin-mediated regulatory pathways play no role in the regulation of L-lactic acid uptake, but a role for PKC- and PKA-mediated pathways was apparent. Uptake of L-lactic acid appeared to be stimulated by phorbol 12-myristate 13-acetate (PMA, a PKC activator) via an increase in V-max of transport processes with no alteration in K-m. In parallel, PMA treatment also resulted in an increase in the level of MCT1 expression. On the other hand, exposure to 8-Br-cAMP, a cAMP analog, and to forskolin, an adenylyl cyclase activator, resulted in a significant decrease in L-lactic acid uptake. Additionally, 8-Br-cAMP reduced V-max but not K-m values. Parallel to the decrease in V-max of L-lactic acid uptake, the level of MCT1 expression was decreased in response to incubation with 8-Br-cAMP. These results indicate the possible involvement of a PKC- and PKA-mediated pathway associated with expression of MCT1 and lactate transport.

Grapefruit juice enhance the uptake of coenzyme Q10 in the human intestinal cell-line Caco-2
Shirou Itagaki, Akiko Ochiai, Masaki Kobayashi, Mitsuru Sugawara, Takeshi Hirano, Ken Iseki
FOOD CHEMISTRY, 120, 2, 552, 555, ELSEVIER SCI LTD, 2010年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Coenzyme Q10 (CoQ10) is very widely consumed by humans as a food supplement. However, CoQ10 is taken up from the intestine into the circulation at a low rate. The absorption of compounds from the gastrointestinal tract is one of the important determinants for oral bioavailability. Secretory transport limits the Oral bioavailability of compounds. It has been reported that efflux transport of CoQ10 is mediated by P-glycoprotein (P-gp) in Caco-2 cells. We tried to improve intestinal absorption of CoQ10 by modulating P-gp. Since grapefruit juice (GFJ) is reported to inhibit P-gp function, we investigated the effect of GFJ on the transport of CoQ10 by Caco-2 cells. In the presence of GFJ, the basal-to-apical transport of CoQ10 was decreased and the uptake of CoQ10 was increased. These findings suggest that the combined administration of CoQ10 and GFJ could enhance CoQ10 absorption. (C) 2009 Elsevier Ltd. All rights reserved.

Pharmacokinetics of Oral and Intravenous Administration of Digoxin after Intestinal Ischemia-Reperfusion
Jiro Ogura, Hajime Maruyama, Masaki Kobayashi, Shirou Itagaki, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 33, 5, 922, 925, PHARMACEUTICAL SOC JAPAN, 2010年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Intestinal ischemia-reperfusion (I/R) causes gut dysfunction characterized by decreased basement membrane integrity and decreased barrier function. Indeed, it has been reported that the absorption of several drugs is altered after intestinal I/R. Intestinal I/R also promotes multi-organ failure (MOF). The liver and kidney can be affected by MOF after intestinal I/R. However, little is known about the alteration of pharmacokinetics after intravenous administration in intestinal I/R injury. In the present study, we investigated pharmacokinetics of digoxin after oral administration and intravenous administration in intestinal I/R injury. Plasma digoxin concentration in I/R rats after oral administration was not significantly altered at any time compared with that in sham-operated rats. Plasma digoxin concentration in rats reperfused for 1 h after intravenous administration was significantly higher than that in sham-operated rats. Plasma digoxin concentrations in rats reperfused for 6 and 24 h were the same as those in sham-operated rats. The area under the concentration time curve after intravenous administraion (AUC(i.s.)) and total clearance (CLtot) in rats reperfused for 1 h was 1.89- and 0.57-fold higher than that in sham-operated rats. However, elimination rate (k(e)) and half-life (t(1/2)) in rats reperfused for 1 h were not altered. Distribution volume (V-d) in rats reperfused for 1 h was decreased than that in sham-operated rats, but there was not statistical difference. These results suggest that intestinal I/R affected the V-d of digoxin, and plasma concentration of digoxin was increased. The present study suggests that understanding pharmacokinetics of drug after intravenous administration in intestinal I/R injury is important to provide valuable information for safe drug therapy for intestinal I/R patients.

In vitro 実験系による肺がん化学療法レジメンの抗腫瘍効果の評価
斎藤 由起子, 平野 剛, 沖 洋充, 笠師 久美子, 菅原 満, 小林 正紀, 高橋 夏子, 板垣 史郎, 井関 健
医療薬学, 36, 4, 220, 226, 一般社団法人日本医療薬学会, 2010年, ［査読有り］
日本語, Lung cancer is the leading cause of cancer mortality.There are 3 major types of lung cancer therapy-surgery,chemotherapy and radiotherapy.In most cases,chemotherapy is the first choice and cisplatin-based combination therapy plays a major role in lung cancer chemotherapy.
As the anticancer effects of lung cancer chemotherapy regimens have not been evaluated in vitro ,we evaluated the effects of combination therapy with cisplatin and irinotecan (IP therapy) and that with cisplatin and etoposide (EP therapy),using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in human lung adenocarcinoma A 549 epithelial cell line cells.In the MTT assay,drug solutions were added according to the same schedule as for the regimens.The exposure times of the drugs were their half-lives,and drug concentrations were the IC₅₀ values of the half-lives.Our results suggested that cisplatin had a much stronger antiproliferative effect than irinotecan and etoposide,and that all agents had a strong anticancer effect.
It has been shown that in these combination therapies,together,the drugs have a stronger anticancer effect than they do when given alone.For IP therapy,cell viability returned at day 28,while it returned at day 15 for EP therapy.These results suggested that the MTT assay can be used in vitro to evaluate cancer chemotherapy regimens as well as dosing schedules for IP therapy and EP therapy.

放射線治療に伴う口腔粘膜障害に対する予防·軽減効果が期待される物質の探索
板垣 史郎, 中田 千絵, 平野 剛, 鷹野 瑠美, 笠師 久美子, 菅原 満, 高橋 夏子, 小林 正紀, 井関 健
医療薬学, 36, 9, 696, 702, 一般社団法人日本医療薬学会, 2010年, ［査読有り］
日本語, In the clinical setting,the frequency of adverse effects is directly linked to the results of treatment.However,in the case of head and neck radiotherapy for cancer,adverse effects are unavoidable and among them,oral mucositis is considered to be the most significant and debilitating acute complication associated with such radiation therapy.In patients receiving head and neck radiotherapy,the prevalence of mucositis is 85-100% and mucositis-associated pain is the main type of cancer treatment-related pain.
A mouthwash or iceball containing sodium alginate,sodium azulene sulfonate and fibrinolysin deoxyribonuclease is widely used for the prevention and treatment of mucositis.In the USA,many strategies for the prevention of oral mucositis have been evaluated,but conflicting data has made the results inconclusive.Keratinocyte Growth Factor 1 (KGF citation=1)has achieved promising results in therapy and is the only medication currently approved by the FDA,though in Japan,it is not covered by the National Health Insurance scheme.Therefore,we feel it is important to establish useful strategies for the prevention and treatment of radiotherapy-induced mucositis based on significant scientific evidence.
As oxidative stress is associated with radiation-induced mucositis,in this study,we used ⁶⁰Co-γ ray-induced HO-1-N-1 cell injury to studying the characteristics of mucositis in this regard and found that two well-known antioxidants,N-AcetylL-cysteine and catechins,had preventive effects against γray-induced cytotoxicity.N-Acetyl-L-cysteine and catechins would therefore be good materials for the prevention of mucositis in patients undergoing cancer radiotherapy.

Protective effects of quercetin-3-rhamnoglucoside (rutin) on ischemia-reperfusion injury in rat small intestine
Shirou Itagaki, Setsu Oikawa, Jiro Ogura, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
FOOD CHEMISTRY, 118, 2, 426, 429, ELSEVIER SCI LTD, 2010年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Quercetin-3-rhamnoglucoside (rutin) has a wide spectrum of biochemical and pharmacological activities. Orally administered compounds are absorbed in the intestine. Although absorption of rutin from the jejunum of the rat was good, binding of rutin to the intestinal wall components may limit its absorption from the small intestine. The physiological importance of an orally administered compound depends on its interaction with target tissues. Since there is limited information on the importance of rutin in vivo, we focused on the protective effect of rutin on intestinal injury. The intestinal mucosa is extremely sensitive to reactive oxygen species. We used a rat mesenteric ischemia-reperfusion (I/R) injury model as a model of oxidative injury and investigated the antioxidant activities of rutin in vivo. We found that rutin, which has combined antioxidant activity from radical-scavenging, xanthine oxidase inhibition and chain-breaking effects, exhibits a protective effect on I/R injury in the rat small intestine. (C) 2009 Elsevier Ltd. All rights reserved.

Expression and Role of SNAT3 in the Placenta
C. Yoshioka, S. Yasuda, F. Kimura, M. Kobayashi, S. Itagaki, T. Hirano, K. Iseki
PLACENTA, 30, 12, 1071, 1077, W B SAUNDERS CO LTD, 2009年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Glutamine is the most versatile amino acid and its plasma concentration is the highest of all amino acid. Many transporters are therefore involved in glutamine uptake or efflux. Glutamine is actively released from the placenta into fetal circulation. In this study, we examined the alteration of transporters that transport glutamine into fetal circulation as gestation progresses. High expression levels of system A and y(+)L were found in the rat placenta in the late period of pregnancy and the expression levels of these transporters increased as gestation progressed (p < 0.05). On the other hand, the expression of SNAT3, the system N transporter, was detected in the early period of pregnancy and its expression level decreased as gestation progressed (p < 0.05). SNAT3 was also found to be expressed in isolated human primary cytotrophoblast cells and its expression level was decreased by their differentiation into syncytiotrophoblast cells (p < 0.05). Since this regulation is closely related to glutamine synthetase expression, SNAT3 may play a key role in providing glutamine corresponding to glutamine synthetase function in the early period of gestation. This is the first report on the expression of SNAT3 in the placenta in the early stage of pregnancy. (C) 2009 Elsevier Ltd. All rights reserved.

Liver X receptor regulates expression of MRP2 but not that of MDR1 and BCRP in the liver
Ikumi Chisaki, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES, 1788, 11, 2396, 2403, ELSEVIER SCIENCE BV, 2009年11月, ［査読有り］
英語, 研究論文（学術雑誌）, Liver X receptors (LXRs) belong to the nuclear hormone receptor superfamily. Multidrug resistance-associated protein 2 (MRP2), multidrug resistance 1 (MDR1) and breast cancer resistance protein (BCRP) play an important role in the efflux of a broad range of endogenous and xenobiotic compounds from hepatocytes. Since the effects of LXR activation on there transporters have been obscure, we investigated the effects of LXR agonists, TO901317 and 25-hydroxycholesterol, on MRP2, MDR1, BCRP expression in HepG2 cells and the rat liver. in an in vitro study, TO901317 increased ABCA1, an LXR target gene, and MRP2 mRNA and protein levels. On the other hand, TO901317 had little effect on MDR1 and BCRP mRNA levels. In an in vivo study, Abca1 and Mrp2 mRNA and protein levels were increased by TO901317. but TO901317 had no effect on Mdr1a and Bcrp mRNA levels in the rat liver. Moreover, TO901317-induced MRP2 mRNA expression was blocked by LXR alpha knockdown. In this study, we demonstrated that LXR activation induced expression of MRP2 but not that of MDR1 and BCRP in hepatocytes. The results suggest that agonists for LXR activate transcription of the MRP2 gene in order to promote excretion of endogenous and xenobiotic compounds from hepatocytes into bile. (C) 2009 Elsevier B.V. All rights reserved.

Response of the ABCG2 promoter in T47D cells and BeWo cells to sex hormone treatment
Satoru Yasuda, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
MOLECULAR BIOLOGY REPORTS, 36, 7, 1889, 1896, SPRINGER, 2009年09月, ［査読有り］
英語, 研究論文（学術雑誌）, The aim of this study was to elucidate the effects of sex hormones on activity of the ABCG2 promoter in different cell lines. T47D cells and BeWo cells were used as models for ABCG2-expressing cell lines, and luciferase assays using ABCG2 promoter-luciferase constructs were performed. It was shown that progesterone increased the response of the ABCG2 promoter in T47D cells but not in BeWo cells. On the other hand, estradiol had no effect on response of the ABCG2 promoter in either cell line. However, response of the ABCG2 promoter was enhanced by overexpression of ER alpha in both T47D cells and BeWo cells. T47D cells had higher sensitivity to ER alpha than did BeWo cells. Furthermore, it was shown that the inductive effect of progesterone on the ABCG2 promoter was inhibited by addition of RU486 or mithramycin A. Therefore, it was thought that the ABCG2 promoter responded to stimulation of the progesterone receptor (PR)-Sp1 pathway in T47D cells. Furthermore, progesterone suppressed the response of the ABCG2 promoter by changing the expression levels of PR-A and PR-B in BeWo cells. These findings suggested that there are differences between cell lines in the regulation mechanism of ABCG2 expression by sex hormone treatment.

Contribution of organic anion transporting polypeptide OATP2B1 to amiodarone accumulation in lung epithelial cells
Satoru Seki, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES, 1788, 5, 911, 917, ELSEVIER SCIENCE BV, 2009年05月, ［査読有り］
英語, 研究論文（学術雑誌）, The accumulation mechanisms of amiodarone (AMD) involving transporters in lung alveolar epithelial type 11 cells were studied. The uptake of AMD was examined using human alveolar epithelia I-derived cell line A549 as a model. AMD was transported by the carrier-mediated system, and the apparent K-m and V-max values were 66.8 +/- 30.3 mu M and 49.7 +/- 9.7 nmol/mg protein/5 min, respectively. The uptake of AMD by A549 cells was Na+-independent and was inhibited by substrates of human organic anion transporting polypeptide (OATP). The inhibition profiles were similar to the inhibitory effects of several compounds on OATP2B1 -mediated E-3-S transport, and RT-PCR analysis showed mRNA expression of OATP2B1 and 1B3 in A549 cells. SiRNAs targeted to the OATP2B1 gene decreased the OATP2B1 mRNA expression level in A549 cells up to about 50% and reduced the uptake of AMD up to about 40%. These results indicate that AMD uptake mediated by carriers, including OATP2B1, might lead to accumulation of AMD in the lung and AMD-induced pulmonary toxicity (AIPT). (C) 2009 Elsevier B.V. All rights reserved.

In vitro and in vivo antioxidant properties of ferulic acid: A comparative study with other natural oxidation inhibitors
Shirou Itagaki, Toshirnitsu Kurokawa, Chie Nakata, Yoshitaka Saito, Setsu Oikawa, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
FOOD CHEMISTRY, 114, 2, 466, 471, ELSEVIER SCI LTD, 2009年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Ferulic acid exhibits a wide range of therapeutic effects that are attributed to its potent antioxidant capacity. However, in vitro antioxidant properties of ferulic acid have not been elucidated in detail. Evidence that polyphenols, including ferulic acid, act as antioxidants in vivo is also limited. In order to elucidate in more detail the scientific background of antioxidant activities of ferulic acid, we carried out in vitro and in vivo experiments. We focused on superoxide anion scavenging activity, xanthine oxidase inhibition activity, and chain-breaking activity. The combined antioxidant activity from radical scavenging and xanthine oxidase inhibition of ferulic acid was much weaker than that of (-)-epigallocatechin gallate (EGCG) and ascorbic acid. On the other hand, EGCG, ascorbic acid and ferulic acid exhibited chain-breaking activity and prevented ischaemia-reperfusion-associated intestinal injury. Chain-breaking activity may play a contributory role in the protective effect of ferulic acid on oxidative injury in humans and in in vivo studies. (C) 2008 Elsevier Ltd. All rights reserved.

Placental folate transport during pregnancy
Satoru Yasuda, Satoko Hasui, Chiaki Yamamoto, Chihiro Yoshioka, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY, 72, 9, 2277, 2284, TAYLOR & FRANCIS LTD, 2008年09月, ［査読有り］
英語, 研究論文（学術雑誌）, The aim of this study was to elucidate the mechanism of folate transport in the placenta over the course of pregnancy. We found that folate receptor of (FR alpha) and reduced folate carrier (RFC) localized on the apical side of human placental villi. Since folate binding to placental brush-border membrane vesicles (BBMVs) was strongly inhibited by phosphatidylinositol-specific phospholipase C (PI-PLC) treatment, it is possible that FR alpha, a glycosyl phosphatidylinositol linked glycoprotein, is a candidate for folate uptake from maternal blood to the placenta. Moreover, additional inhibitory effects of thiamine pyrophosphate (TPP) and hemin on folate uptake after PI-PLC treatment suggested that not only FRa but also RFC and heme carrier protein I (HCPI) are involved in the folate transport mechanism in the human placenta. It was also found that accumulation of folate after intravenous injection increased with the progress of gestation in the rat placenta and the fetus. Furthermore, increases in the expression levels of mRNA of rFR alpha, rRFC, and rHCP1 in the rat placenta during pregnancy were observed. These findings suggest that FR alpha, RFC, and HCP1 are important carriers of folate in the placenta during pregnancy. The results of this study suggest that increases in the expression levels of FRa, REC, and HCP1 in the placenta play an important role in the response to increased need for folate for the placenta and fetus during development with the progress of gestation.

Interaction of coenzyme Q10 with the intestinal drug transporter P-glycoprotein
Shirou Itagaki, Akiko Ochiai, Masaki Kobayashi, Mitsuru Sugawara, Takeshi Hiran, Ken Iseki
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY, 56, 16, 6923, 6927, AMER CHEMICAL SOC, 2008年08月, ［査読有り］
英語, 研究論文（学術雑誌）, In clinical trials, patients usually take many kinds of drugs at the same time. Thus, drug-drug interactions can often directly affect the therapeutic safety and efficacy of many drugs. Oral delivery is the most desirable means of drug administration. Changes in the activity of drug transporters may substantially influence the absorption of administered drugs from the intestine. However, there have been a few studies on food-drug interactions involving transporters. It is important to be aware of the potential of food-drug interactions and to act in order to prevent undesirable and harmful clinical consequences. Coenzyme Q10 (CoQ10) is very widely consumed by humans as a food supplement because of its recognition by the public as an important nutrient in supporting human health. Since intestinal efflux transporter P-glycoprotein (P-gp) is one of the major factors in drug-drug interactions, we focused on this transporter. We report here for the first time that CoQ10, which is widely used as a food supplement, affects the transport activity of P-gp.

Amiodarone increases the accumulation of DEA in a human alveolar epithelium-derived cell line
Satoru Seki, Shirou Itagaki, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 31, 7, 1449, 1452, PHARMACEUTICAL SOC JAPAN, 2008年07月, ［査読有り］
英語, 研究論文（学術雑誌）, Amiodarone (AMD)-induced pulmonary toxicity (AIPT) is the most life-threatening side-effect of AMD treatment. N-Monodesethylamiodarone (DEA), an active metabolite of AMD, also exhibits cytotoxicity and tends to accumulate in the lung more intensively than AMD. In this study, we characterized the mechanism of DEA accumulation using A549 cells as a model of the alveolar epithelium. Typical ATP-depletion compounds caused an approximately 30% increase in the accumulation of DEA in A549 cells, although these effects were less than those in Caco-2 cells. Triiodothyronine (T3), which exhibited an inhibitory effect on DEA efflux in Caco-2 cells, did not affect the accumulation of DEA in A549 cells. On the other hand, 100uM AMD caused an approximately 200% increase in DEA content in A549 cells, although AMD accumulation was not affected by 100 um DEA. Since the reducing effect of AMD on cellular ATP levels and that of FCCP were similar, the mechanism by which DEA accumulation is increased by AMD might be different from the ATP-dependent DEA efflux mechanism. The decrease in cell viability by DEA in the presence of AMD (ICS value of DEA for A549 cell viability: 25.4 2.4 ELM) was more pronounced than that by DEA alone (IC5o value: 11.5 3.0 um). This further DEA accumulation by AMD might be a factor responsible for the greater accumulation of DEA than that of AMD in the lung in long-term AMD-treated patients.

Alteration of Mrp2 and P-gp expression, including expression in remote organs, after intestinal ischemia-reperfusion
Jiro Ogura, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
LIFE SCIENCES, 82, 25-26, 1242, 1248, PERGAMON-ELSEVIER SCIENCE LTD, 2008年06月, ［査読有り］
英語, 研究論文（学術雑誌）, The present study was carried out in order to identify the changes in expression of multidrug resistance-associated protein (Mrp) 2 and P-glycoprotein (P-gp) in the intestine and remote organs after intestinal ischemia-reperfusion (I/R). Mrp2 expression in the jejunum and liver was decreased at 6 h after I/R. This decrease in Mrp2 expression was associated with an increase in the serum level of IL-6. These results suggest that the decreased Mrp2 expression after intestinal I/R was regulated by IL-6. The expression level of mdr1a in the ileum, which encodes P-gp, was decreased at 6 and 24 h after I/R, and the expression level of mdr1b, also encodes P-gp, was not altered at any time. P-gp protein expression in the ileum was decreased at 6 h after I/R. In the liver, mdr1a expression was decreased at 6 h after I/R, but mdr1b expression was increased at 6 h after I/R. Pgp protein was not altered at any time. In the kidney, mdr1a expression was decreased at 24 h after I/R, but mdr1b expression was not altered at any time. P-gp protein expression in the kidney was decreased at 24 h after I/R, as was mdr1a expression. These results suggest that P-gp expression after intestinal I/R differs in each organ. This is the first report to provide evidence that expression levels of transporters in remote organs are altered intestinal after I/R. (c) 2008 Elsevier Inc. All rights reserved.

Post-transcriptional regulation of breast cancer resistance protein after intestinal ischemia-reperfusion
Jiro Ogura, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 31, 5, 1032, 1035, PHARMACEUTICAL SOC JAPAN, 2008年05月, ［査読有り］
英語, 研究論文（学術雑誌）, Breast cancer resistance protein (BCRP), the product of the ABCG2 gene, is a recently identified ATP binding cassette half-transporter. BCRP is expressed in a variety of tumor cells and many normal human tissues. In the small intestine, BCRP can limit the influx and facilitate the efflux to prevent intracellular accumulation of BCRP substrates. Ischemia-reperfusion (I/R) induces the release of reactive oxygen species, and organs are severely damaged by I/R. It has been shown that the expression of transporters was altered in the organ after I/R. The present study was undertaken to clarify the expression of BCRP after intestinal I/R. We showed that the expression level of Bcrp was significantly decreased at I h after I/R. Bcrp mRNA level was not altered at I h after I/R. These results suggest that Bcrp expression was regulated by a post-transcriptional regulation mechanism after intestinal I/R. Bcrp mRNA level was increased at 24 h after I/R, and the expression level of Bcrp protein was of the same level or slightly increased compared with sham operated-rats. Bcrp was slightly located at the intestinal membrane at 24 h after intestinal I/R. These results suggested that Bcrp was not translocated to the intestinal membrane after intestinal I/R. There is little information on post-transcriptional regulation compared with information on transcriptional regulation. In this study, it was shown that Bcrp expression is regulated by post-transcriptional regulation after intestinal I/R. These results of this study may provide important information for further studies aimed at revealing the biological function of Bcrp.

Association between risk of myopathy and cholesterol-lowering effect: A comparison of all statins
Masaki Kobayashi, Ikumi Chisaki, Katsuya Narumi, Kazuhiro Hidaka, Toshiki Kagawa, Shirou Itagaki, Takeshi Hirano, Ken Iseki
LIFE SCIENCES, 82, 17-18, 969, 975, PERGAMON-ELSEVIER SCIENCE LTD, 2008年04月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, In the present study, we examined the mechanisms underlying the cytotoxicity of pitavastatin, a new statin, and we compared the in vitro potencies of muscle cytotoxicity using a prototypic embryonal rhabdomyosarcoma cell line (RD cells), a typical side effect of statins and compared the cholesterol-lowering effects of statins using Hep G2 hepatoma cells. Pitavastatin reduced the number of viable cells and caused caspase-9 and -3/7 activation in a time- and concentration-dependent manner. The comparison of cytotoxities of statins showed that statins significantly reduced cell viability and markedly enhanced activity of caspase-3/7 in concentration-dependent manner. On the other hand, the effects of hydrophilic statins, pravastatin, rosuvastatin were very weak. The rank order of cytotoxicity was cerivastatin > sinavastatin acid> fluvastatin > atorvastatin > lovastatin acid > pitavastatin >> rosuvastatin, pravastatin. Statin-induced cytotoxicity is associated with these partition coefficients. On the other hand, the cholesterol-lowering effect of statins did not correlate with these partition coefficients and cytotoxicity. Thus, it is necessary to consider the association between risk of myopathy and cholesterol-lowering effect of a statin for precise use of statins. (c) 2008 Elsevier Inc. All rights reserved.

A new method for determination of both thalidomide enantiomers using HPLC systems
Kaname Sgmbongi, Masanori Tanaka, Kelsuke Sakurada, Masakl Kobayasfi, Shlrou Itagaki, Takeshi Hirano, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 31, 3, 497, 500, PHARMACEUTICAL SOC JAPAN, 2008年03月, ［査読有り］
英語, 研究論文（学術雑誌）, Although thalidomide was withdrawn due to teratogenicity and neuropathy, there is now growing clinical interest in this compound because of its immunomodulatory and anti-angiogenic properties. In 1998, thalidomide was approved by the U.S. Food and Drug Administration for the treatment of erythema nodosum leprosum (ENL), an inflammatory complication of Hansen's disease, through a restricted-use program. Thalidomide was approved for the treatment of relapsed or refractory multiple myeloma (MM) as an orphan drug in Japan. Direct deproteinization method was shown to be useful for quantitation of enantioselective thalidomide blood level. Stabilized blood was deproteinized with methanol and 2 m trichloroacetic acid. The supernatant was injected onto reverse-phase column (CHIRALPAK AD-RH). The mobile phase consisted of 10% acetonitrile, 70% methanol and 20% 0.025 M citrate buffer (pH 3.0), and the flow rate was 0.5 ml/min. Wavelength of detection was 220 nm. (-)-(S)-thalidomide and (+)-(R)-thalidomide were separated at 13.5 min and 17.6 min, respectively. The accuracy of this method was almost the same as that of the measurement technique with extraction and concentration. In clinical practice, MM patients usually take many kinds of drugs at the same time. Actually, this patient takes a lot of drugs with thalidomide. However, we found no interference of these drugs and thalidomide on the chromatogram. This simple and reliable HPLC determination method for both enantiomers of thalidomide is thought to be very useful for thalidomide studies.

The inhibitory effects of fluoroquinolones on L-carnitine transport in placental cell line BeWo
Takeshi Hirano, Satoru Yasuda, Yuki Osaka, Masaru Asari, Masaki Kobayashi, Shirou Itagaki, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 351, 1-2, 113, 118, ELSEVIER SCIENCE BV, 2008年03月, ［査読有り］
英語, 研究論文（学術雑誌）, L-Carnitine plays an important role in lipid metabolism by facilitating the transport of long-chain fatty acids across the mitochondrial inner membrane followed by fatty acid beta-oxidation. It is known that members of the OCTN family play an important role in L-carnitine transport in the placenta. Investigation of drug-drug or drug-nutrient interaction in the placenta is important for establishment of safety drug medication during pregnancy. The aim of this study was to determine the effects of fluoroquinolones, inhibitors of OCTN2, on L-carnitine transport in the placenta which is known to have a high expression level of OCTN2.
We investigated the inhibitory effect of five fluoroquinolones, ciprofloxacin (CPFX), gatifloxacin (GFLX), ofloxacin (OFLX), levofloxacin (LVFX) and grepafloxacin (GPFX), on L-carnitine transport mediated by OCTN2 in placental cell line BeWo cells. We found that all of the fluoroquinolones inhibited L-carnitine transport, GPFX being the strongest inhibitor. We also found that the inhibitory effects of LVFX and GPFX depended on their existence ratio of zwitterionic forms as, we reported previously.
Furthermore, we elucidated the LVFX transport mechanism in BeWo cells. LVFX was transported actively by transporters. However, we found that LVFX transport was Na+-independent and L-carnitine had no inhibitory effect on LVFX transport, suggesting that LVFX acts as inhibitor of OCTN2, not as a substrate for OCTN2. (C) 2007 Elsevier B.V. All rights reserved.

The mechanism of carrier-mediated transport of folates in BeWo cells: The involvement of heme carrier protein 1 in placental folate transport
Satoru Yasuda, Satoko Hasui, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY, 72, 2, 329, 334, TAYLOR & FRANCIS LTD, 2008年02月, ［査読有り］
英語, 研究論文（学術雑誌）, The aim of this study was to elucidate the mechanism of folate transport in the placenta. A study of folate was carried out to determine which carriers transport folates in the human choriocarcinoma cell line BeWo, a model cell line for the placenta. We investigated the effects of buffer pH and various compounds on folate uptake. In the first part of the study, the expression levels of the mRNA of the folate receptor alpha (FR alpha), the reduced folate carrier (RFC), and heme carrier protein 1 (HCP1) were determined in BeWo cells by RT-PCR analysis. Folate uptake into BeWo cells was greater under an acidic buffer condition than under a neutral one. Structure analogs of folates inhibited folate uptake under all buffer pH conditions, but anion drugs (e.g., pravastatin) inhibited folate uptake only under an acidic buffer condition. Although thiamine pyrophosphate (TPP), a substrate of RFC, had no effect on folate uptake, hemin (a weak inhibitor of folate uptake via HCPI) decreased folate uptake to about 80% of the control level under an acidic buffer condition. Furthermore, kinetic analysis showed that hemin inhibited the low-affinity phase of folate uptake under an acidic buffer condition. We conclude that pH-dependent folate uptake in BeWo cells is mediated by at least two carriers. RFC is not involved in folate uptake, but FR alpha (high affinity phase) and HCP1 (low affinity phase) transport folate in BeWo cells.

Functional characterization of human monocarboxylate transporter 9 (SLC16A9)
Kobayashi Masaki, Kagawa Toshiki, Narumi Katsuya, Hidaka Kazuhiro, Itagaki Shirou, Hirano Takeshi, Iseki Ken
YAKUGAKU ZASSHI-JOURNAL OF THE PHARMACEUTICAL SOCIETY OF JAPAN, 128, 47, 48, 2008年, ［査読有り］

Functional analysis of phenolsulfonphthalein transport system in Long-Evans Cinnamon rats.
Itagaki S, Chiba M, Kobayashi M, Sugawara M, Kobayashi M, Hirano T, Iseki K
Biochimica et biophysica acta, 1778, 1, 270, 275, 2008年01月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, It has been reported that the transport function for organic anions on the kidney is maintained in multidrug resistance-associated protein 2 (Mrp2)-deficient rats. Different from Mrp2-deficient rats, Long-Evans Cinnamon (LEC) rats have impaired urinary excretion of Mrp2-substrate, phenolsulfonphthalein (PSP). PSP is transported by the potential-sensitive urate transport system in rat brush-border membranes. We analyzed the function of PSP transport system in LEC rats. Unlike Long-Evans Agouti (LEA) rats, the initial uptake of PSP and urate into the renal brush-border membrane vesicles of LEC rats were not significantly enhanced in the presence of positive intravesicular potential, suggesting that the potential-sensitive urate transport system is impaired in LEC rats. LEC rats should be useful for elucidating the potential-sensitive urate transport system in rats at the molecular level.

Contribution of multidrug resistance-associated protein 2 to secretory intestinal transport of organic anions
Shirou Itagaki, Makoto Chiba, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 31, 1, 146, 148, PHARMACEUTICAL SOC JAPAN, 2008年01月, ［査読有り］
英語, 研究論文（学術雑誌）, Various mechanisms can influence the intestinal absorption and oral bioavailability of drugs. The barrier effects of efflux transporters may be one of the critical factors limiting the bioavailability of certain drugs. It has been reported that multidrug resistance-associated protein 2 (Mrp2) is expressed in the mucosal membrane of the epithelium of the small intestine and secretes various drugs into the jejunum lumen. However, it is possible that total intestinal secretion of Mrp2 substrates is accounted for the contribution of Mrp2 and other transporter(s) to the intestinal secretion of Mrp2 substrates. In this study, we found that phenolsulfonphthalein and pravastatin, both Mrp2 substrates, are transported by different transport systems in the intestine. These results suggest that contribution of transporters to the drug transport may be a critical factor affecting drug disposition and drug-drug interaction. In addition to evaluating the substrate specificity of a transporter, it is important to be aware of the contribution of a transporter to drug disposition.

Bicarbonate supplementation as a preventive way in statins-induced muscle damage
Masaki Kobayashi, Toshiki Kagawa, Katsuya Narumi, Shirou Itagaki, Takeshi Hirano, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 11, 1, 1, 8, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2008年, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Purpose. The aim of this study was to evaluate the bicarbonate-induced improvement of statins, cerivastatin, simvastatin acid and lovastatin acid-induced apoptosis using rat myoblast cell line (L6) as a model of in vitro skeletal muscle and of cerivastatin-induced muscle damage in vivo study. Methods. Statin-induced reduction of cell viability and apoptosis was measured by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) assay and caspase assay. In vivo, we evaluated plasma creatine phosphokinase (CPK) level in cerivastatin-treated rat. Results. Bicarbonate prevented cerivastatin-, simvastatin-acid and lovastatin acid-induced reduction of cell viability, morphological change and caspase activation in L6 cells. Moreover, in the in vivo study, bicarbonate prevented cerivastatin-induced increase in CPK concentrations. Conclusions. These results from in vitro and in vivo studies support that bicarbonate supplementation prevented statin-induced muscle damage.

Preventive effects of bicarbonate on cerivastatin-induced apoptosis
Masaki Kobayashi, Fumie Kaido, Toshiki Kagawa, Shirou Itagaki, Takeshi Hirano, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 341, 1-2, 181, 188, ELSEVIER SCIENCE BV, 2007年08月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Although HMG-CoA reductase inhibitors such as statins are the most widely used cholesterol-lowering agents, there is a risk of myopathy or rhabdmyolysis occurring in patients taking these drugs. It has been reported that a number of lipophilic statins cause apoptosis in various cells, but it is still not clear whether intracellular acidification is involved in statin-induced apoptosis. There have been few studies aimed at identifying compounds that suppress statin-induced myotoxicity. In the present study, we examined the relationship between cerivastatin-induced apoptosis and intracellular acidification and the effect of bicarbonate on cerivastatin-induced apoptosis using an RD cell line as a model of in vitro skeletal muscle. Cerivastatin reduced the number of viable cells and caused dramatic morphological changes and DNA fragmentation in a concentration-dependent manner. Moreover, cerivastatin-induced apoptosis was associated with intracellular acidification and caspase-9 and -3/7 activation. On the other hand, bicarbonate suppressed cerivastatin-induced pH alteration, caspase activation, morphological change and reduction of cell viability. Accordingly, bicarbonate suppressed statin-induced apoptosis. The strategy to combine statins with bicarbonate can lead to reduction in the chance of the severe adverse events including myopathy or rhabdmyolysis. (c) 2007 Elsevier B.V. All rights reserved.

Effect of medium pH on the cytotoxicity of hydrophilic statins
Masaki Kobayashi, Toshiki Kagawa, Rumi Takano, Shirou Itagaki, Takeshi Hirano, Ken Iseki
JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES, 10, 3, 332, 339, CANADIAN SOC PHARMACEUTICAL SCIENCES, 2007年05月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Purpose. The aim of this study was to examine the mechanism of pravastatin- and rosuvastatin-induced cytotoxicity and the relationship between pravastatin- and rosuvastatin-induced cytotoxicity and medium pH using human prototypic embryonal rhabdomyosarcoma cell line (RD) and rat myoblast cell line (L6) as a model of in vitro skeletal muscle. Methods. Statin-induced reduction of cell viability and apoptosis was measured by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) assay and caspase assay. Intracellular accumulation of statins was determined using an HPLC system. Results. Rosuvastatin cytotoxicity, reduction of cell viability, morphological changes and caspase activation at acidic pH (pH 6.8) were significantly greater than those at neutral pH (pH 7.4). Rosuvastatin accumulation at acidic pH was greater than that at pH 7.4. On the other hand, medium pH had no effect on pravastatin accumulation. Conclusions. Rosuvastatin cytotoxicity at acidic pH is associated with increasing intracellular accumulation of rosuvastatin. On the other hand, medium pH had no effect on cytotoxicity of pravastatin.

Improvement in intestinal coenzyme q10 absorption by food intake
Akiko Ochiai, Shirou Itagaki, Toshimitsu Kurokawa, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
YAKUGAKU ZASSHI-JOURNAL OF THE PHARMACEUTICAL SOCIETY OF JAPAN, 127, 8, 1251, 1254, PHARMACEUTICAL SOC JAPAN, 2007年, ［査読有り］
英語, 研究論文（学術雑誌）, Coenzyme Q10 (CoQ10) is widely consumed as a food supplement because of its recognition as an important nutrient in supporting human health. Absorption of compounds from the gastrointestinal tract is one of the important determinants of oral bioavailability. However, the absorption of dietary CoQ10 is slow and limited due to its hydrophobicity and large molecular weight. The absorption of orally applied compounds can be enhanced by interactions with food or food components. Thus, we investigated the effect of food intake on the absorption of CoQ10 after oral supplememation. In this study, we demonstrated that food intake enhanced the intestinal absorption of CoQ10. In order to improve intestinal absorption of CoQ10 after oral supplementation, we developed an emulsion formulation. Intestinal absorption of CoQ10 after administration of the emulsion formulation was also enhanced by food intake. Moreover, the peak concentration and the extent of absorption after administration of the emulsion formulation were greater than those after administration of a suspension formulation. It is possible that administration of CoQ10 in an emulsion formulation enhances the pharmacological effects of CoQ10.

Efflux transport of n-monodesethylamiodarone by the human intestinal cell-line caco-2 cells
Emi Kimoto, Satoru Seki, Shirou Itagaki, Maya Matsuura, Masaki Kobayashi, Takeshi Hirano, Yoshikazu Goto, Koji Tadano, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 22, 4, 307, 312, JAP SOC STUDY XENOBIOTICS, 2007年, ［査読有り］
英語, 研究論文（学術雑誌）, Amiodarone (AMD) is a benzofurane derivative with class III antiarrhythmic activity that is effective in controlling intractable cardiac arrhythmias. One of the most common and serious drug interactions in clinical practice is the interaction between digoxin and an antiarrhythmic agent. It has been reported that AMD and N-monodesethylamiodarone (DEA), the active metabolite of AMD, inhibit the P-glycoprotein (P-gp/MDR1)-mediated digoxin transport. However, the intestinal transport processes of AMD and DEA have not been fully revealed. In this study, we focused on the intestinal transport mechanism of DEA and characterized the intestinal transport of DEA using Caco-2 cells. Basal-to-apical transport of DEA by Caco-2 cells was greater than apical-to-basal transport. The relationship between concentration and basal-to-apical flux rate appeared to approach saturation. The uptake of DEA by Caco-2 cells was increased in the presence of typical ATP-depletion compounds and thyroid hormones. On the other hand, substrates for P-gp, multidrug resistance-associated proteins (MRPs/ABCCs) and breast cancer resistance protein (BCRP/ABCG2) had no effect on the efflux of DEA. These results suggest that an ATP-binding cassette (ABC) transporter, which is different from P-gp, MRPs and BCRP, mediates the efflux of DEA across the apical membrane in Caco-2 cells and that thyroid hormone inhibits this transporter.

Characterization of hepatobiliary organic anion transporters in Long-Evans Cinnamon rats
Makoto Chiba, Shirou Itagaki, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 22, 5, 387, 390, JAP SOC STUDY XENOBIOTICS, 2007年, ［査読有り］
英語, 研究論文（学術雑誌）, The liver plays important roles in the detoxification of xenobiotecs. Hepatobiliary transporters contribute to hepatic uptake and efflux processes of xenobiotecs. Expressions of these transporters may be modulated under the condition of hepatic failure. Long-Evans Cinnamon (LEC) rats provide a pertinent model for basic and clinical studies on hepatitis. However, only a few reports describing the properties of hepatobiliary transporters in LEC rats have appeared in the literature. We investigated the expression levels of hepatobiliary transporters in LEC rats by real-time RT-PCR. We found that hepatic expressions of three sinusoidal organic anion transporters, Ntcp, Oatp1a1 and Oatp1a4, were decreased in LEC rats. However, no significant difference of the expressions of Mrp2 and Bsep, organic anion transporters located on canalicular membrane, were found between Wistar rats and LEC rats.

Down-regulation of intestinal multidrug resistance-associated protein 2 in long-evans Cinnamon rats
Makoto Chiba, Shirou Itagaki, Masaki Kobayashi, Takeshi Hirano, Ken Iseki
DRUG METABOLISM AND PHARMACOKINETICS, 22, 6, 450, 455, JAP SOC STUDY XENOBIOTICS, 2007年, ［査読有り］
英語, 研究論文（学術雑誌）, Wilson's disease is an inherited, autosomal recessive disorder of copper accumulation and toxicity. Lifelong chelation therapy is essential in all Wilson's disease patients. Intestinal absorption of some compounds is limited partly because they are preferentially transported in the secretory direction. Several ATP-binding cassette (ABC) transporters are expressed in the apical membrane of the small intestine and secrete various drugs into the lumen. In this study, we investigated the characteristics of the intestinal efflux ABC transporters in LEC rats. We found that the expression of multidrug resistance-associated protein 2 (Mrp2) in the jejunum of Long-Evans Cinnamon (LEC) rats, an animal model for Wilson's disease, is decreased.

Mechanism of the inhibitory effect of zwitterionic drugs (levofloxacin and grepafloxacin) on carnitine transporter (OCTN2) in Caco-2 cells.
Hirano T, Yasuda S, Osaka Y, Kobayashi M, Itagaki S, Iseki K
Biochimica et biophysica acta, 1758, 11, 1743, 1750, 2006年11月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, L-Carnitine plays an important role in lipid metabolism by facilitating the transport of long-chain fatty acids across the mitochondrial inner membrane followed by fatty acid beta-oxidation. It is known that L-carnitine exists as a zwitterion and that member of the OCTN family play an important role in its transport. The aims of this study were to characterize L-carnitine transport in the intestine by using Caco-2 cells and to elucidate the effects of levofloxacin (LVFX) and grepafloxacin (GPFX), which are zwitterionic drugs, on L-carnitine uptake. Kinetic analysis showed that the half-saturation Na+ concentration, Hill coefficient and Km value of L-carnitine uptake in Caco-2 cells were 10.3 +/- 4.5 mM, 1.09 and 8.0 +/- 1.0 microM, respectively, suggesting that OCTN2 mainly transports L-carnitine. LVFX and GPFX have two pKa values and the existence ratio of their zwitterionic forms is higher under a neutral condition than under an acidic condition. Experiments on the inhibitory effect of LVFX and GPFX on L-carnitine uptake showed that LVFX and GPFX inhibited L-carnitine uptake more strongly at pH 7.4 than at pH 5.5. It was concluded that the zwitterionic form of drugs plays an important role in inhibition of OCTN2 function.

Inhibitory effects of statins on human monocarboxylate transporter 4
Masaki Kobayashi, Yukio Otsuka, Shirou Itagaki, Takeshi Hirano, Ken Iseki
INTERNATIONAL JOURNAL OF PHARMACEUTICS, 317, 1, 19, 25, ELSEVIER SCIENCE BV, 2006年07月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Human MCT4 (SLC16A3) is responsible for the efflux Of L-lactic acid from skeletal muscle cells and is essential for muscle homeostasis. However, the effects of monocarboxylate drugs, such as statins on the MCT4-mediated transport Of L-lactic acid have not been elucidated. Inhibition Of L-lactic acid transport mediated by MCT4 might to lead to collapse of muscle homeostasis. The aim of this study was to establish an MCT4 transfected cell line and to clarify the transport mechanism Of L-lactic acid and the effects of statins on this transport system. Results of Western blot analyses and immunohistochemistry studies indicated that the expression of CD147 and MCT4-FLAG protein were observed and was displayed clear plasma membrane localization in CD147 and MCT4-FLAG co-transfected cell line (cm cells). Uptake of L-lactic acid in cm cells was significantly greater than that in cells transfected with a vector alone. L-lactic acid uptake was concentration-dependent with a K. value of 28.43 +/- 3.87 mM. The results of a previous study showing a K-m value of 28.5 mM in hMCT4-expressed oocytes. Lipophilic statins significantly inhibited [C-14] L-lactic acid uptake in a concentration-dependent manner. In contrast, the inhibitory effects of hydrophilic statins were very weak. (c) 2006 Elsevier B.V. All rights reserved.

Purification by p-aminobenzoic acid (PABA)-affinity chromatography and the functional reconstitution of the nateglinide/H+ cotransport system in the rat intestinal brush-border membrane
Y Saito, S Itagaki, S Kubo, M Kobayashi, T Hirano, K Iseki
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 340, 3, 879, 886, ACADEMIC PRESS INC ELSEVIER SCIENCE, 2006年02月, ［査読有り］
英語, 研究論文（学術雑誌）, (-)-N-(trans-4-Isopropyleyclohexanecarbonyl)-D-phenylalanine (nateglinide) is a novel oral hypoglycemic agent possessing a peptide-type bond and a carboxyl group in its structure. Recently, we have shown that nateglinide transport occurs via the ceftibuten/H+ cotransport system, which is distinct from PepT1, and that the fluorescein/H+ cotransport system is involved in the Uptake of nateglinide. The aim Of this Study was to characterize the functional properties of the intestinal nateglinide transporter. In the first part of this Study, we demonstrated that the ceftibuten/H+ cotransport system is identical to the fluorescein/H+ cotransport system. We Succeeded in purification of the nateglinide transporter from brush-border membranes of the rat small intestine using p-aminobenzoic acid (PABA)-affinity chromatography. We then investigated the functional properties of the nateglinide transporter using proteoliposomes prepared from the PABA-affinity chromatography elute. We demonstrated that nateglinide, ceftibuten, and fluorescein are transported by the same transporter in the intestine. (c) 2005 Elsevier Inc. All rights reserved.

Transepithelial transport of telmisartan in Caco-2 monolayers
Y Goto, S Itagaki, S Umeda, M Kobayashi, T Hirano, K Iseki, K Tadano
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 28, 12, 2235, 2239, PHARMACEUTICAL SOC JAPAN, 2005年12月, ［査読有り］
英語, 研究論文（学術雑誌）, Telmisartan is the most recently marketed angiotensin 11 type I receptor antagonist. Drug-drug interactions involving transporters can directly affect the therapeutic safety and efficacy of many important drugs. In clinical practice, telmisartan is coadministered with many kinds of drugs. However, little is known about the contribution of transporters to the intestinal transport of telmisartan. The aim of this study was to determine the transport mechanism of telmisartan across intestinal epithelial cells. In the presence of an inwardly directed proton gradient, the apical-to-basal transport of telmisartan was greater than basal-to-apical transport. Thus, we focused on the uptake mechanism of telmisartan across brush-border membranes. The uptake of telmisartan by Caco-2 cells was shown to be energy- and proton-dependent. Although some monocarboxylates inhibited the uptake of telmisartan, L-lactic acid, which is a typical substrate of the monocarboxylate transporter (MCT) 1-MCT4, did nor affect the uptake of telmisartan. Preloading of acetic acid enhanced the uptake of telmisartan, showing a trans-stimulation effect. These results suggest that the carrier-mediated transport system is involved in the uptake of telmisartan by Caco-2 cells and that the apical-localized transport system is similar to MCTs, but not MCT1-MCT4. It is possible that telmisartan reduce the absorption of coadministered drugs by sharing the MCTs. Since MCTs have an important role in the intestinal absorption of pharmacologically active compounds, it is important to be aware of the potential of telmisartan-drug interactions involving MCTs and to act in order to prevent undesirable and harmful consequences.

Nateglinide uptake by a ceftibuten transporter in the rat kidney brush-border membrane.
Kobayashi M, Saito Y, Itagaki S, Hirano T, Iseki K
Biochimica et biophysica acta, 1715, 1, 19, 24, 2005年08月, ［査読有り］, ［筆頭著者］, ［国際誌］
英語, 研究論文（学術雑誌）, Nateglinide, a novel oral hypoglycemic agent, possesses a carbonyl group and a peptide-type bond in its structure. We previously reported that nateglinide transport occurs via a single system that may be identical to the ceftibuten/H(+) cotransport system by the rat small intestine. We speculated that the absorption system present on the intestinal epithelium may be similar to that found on the renal tubular epithelium. The aim of this study was to characterize the transporters on the apical side of the kidney that may contribute to the reabsorption of ceftibuten and nateglinide. The uptake of nateglinide by rat renal brush-border membranes is associated with an H(+)-coupled transport system. Ceftibuten competitively inhibited H(+)-dependent nateglinide uptake. In contrast, Gly-Sar, cephradine and cephalexin had no effect on nateglinide uptake. Nateglinide competitively inhibited H(+)-driven transporter-mediated ceftibuten uptake. We conclude that nateglinide transport occurs via a single system that is H(+)-dependent and may be identical to the ceftibuten/H(+) cotransport system.

Transport mechanism for L-lactic acid in human myocytes using human prototypic embryonal rhabdomyosarcoma cell line (RD cells)
M Kobayashi, Fujita, I, S Itagaki, T Hirano, K Iseki
BIOLOGICAL & PHARMACEUTICAL BULLETIN, 28, 7, 1197, 1201, PHARMACEUTICAL SOC JAPAN, 2005年07月, ［査読有り］, ［筆頭著者］
英語, 研究論文（学術雑誌）, Monocarboxylate transporter (MCT), which cotransport L-lactic acid and protons across cell membranes, are important for regulation of muscle pH. However, it has not been demonstrated in detail whether MCT isoform contribute to the transport Of L-lactic acid in skeletal muscle. The aim of this study was to characterize L-lactic acid transport using an human rhabdomyosarcoma (RD) cell line as a model of human skeletal muscle. mRNAs of MCT 1, 2 and 4 were found to be expressed in RD cells. The [C-14] L-lactic acid uptake was concentration-dependent with a K-m of 1.19 mm. This K-m value was comparable to its K-m values for MCTI or MCT2. MCT1 mRNA was found to be present markedly greater than that MCT2. Therefore, MCTI most probably acts on L-lactic acid uptake at RD cells. [C-14] L-Lactic acid efflux in RD cells was inhibited by alpha-cyano-4-hydroxycinnamate (CHC) but not by butyric acid, a substrate of MCT1. Accordingly, MCT2 or MCT4 is responsible for L-lactic acid efflux by RD cells. MCT4 mRNA was found to be present significantly greater than that MCT2. We conclude that MCTI is responsible for L-lactic acid uptake and L-lactic acid efflux is mediated by MCT4 in RD cells.

Substrate specificity of the nateglinide/H+ cotransport system for phenolic acids
Y Saito, S Itagaki, Y Otsuka, Y Kobayashi, H Okumura, M Kobayashi, T Hirano, K Iseki
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY, 53, 15, 6100, 6104, AMER CHEMICAL SOC, 2005年07月, ［査読有り］
英語, 研究論文（学術雑誌）, In clinical, patients usually take many kinds of drugs at the same time. Thus, drug-drug interactions involving transporters can often directly affect the therapeutic safety and efficacy of many important drugs. However, there have been few studies on food-drug interactions involving transporters. Dietary polyphenols have been widely assumed to be beneficial for human health. Polyphenols are found ubiquitously, and they are commercially prepared and used as functional foods. We have reported that ferulic acid, which is one of the most well-known polyphenols and is used as a functional food, affected the transport of nateglinide, an antidiabetic drug, by Caco-2 cells. In this study, we investigated the effects of other polyphenols on the nateglinide/H+ transport system. We report here that caffeic acid and p-coumaric acid have a different inhibitory manner on the uptake of nateglinide. The results of this study are useful to identify the substrate specificity of the nateglinide/H+ cotransporter.

Food-drug interaction between ferulic acid and nateglinide involving the fluorescein/H+ cotransport system
S Itagaki, Y Kobayashi, Y Otsuka, S Kubo, M Kobayashi, T Hirano, K Iseki
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY, 53, 7, 2499, 2502, AMER CHEMICAL SOC, 2005年04月, ［査読有り］
英語, 研究論文（学術雑誌）, In clinical, patients usually take many kinds of drugs at the same time. Thus, drug-drug interactions involving transporters can often directly affect the therapeutic safety and efficacy of many drugs. However, there have been few studies on food-drug interactions involving transporters. Dietary polyphenols have been widely assumed to be beneficial to human health. Polyphenols are commercially prepared and used as functional foods. We report here for the first time that ferulic acid, which is widely used as a functional food, affects the transport of clinical agents. It is important to be aware of the potential of food-drug interactions and to act in order to prevent undesirable and harmful clinical consequences.

Intestinal uptake of nateglinide by an intestinal fluorescein transporter.
Itagaki S, Otsuka Y, Kubo S, Okumura H, Saito Y, Kobayashi M, Hirano T, Iseki K
Biochimica et biophysica acta, 1668, 2, 190, 194, 2005年03月, ［査読有り］, ［国際誌］
英語, 研究論文（学術雑誌）, Nateglinide, a novel oral hypoglycemic agent, rapidly reaches its maximum serum concentration after oral administration, suggesting that it is rapidly absorbed in the intestine. However, nateglinide itself is not transported by MCT1 or PEPT1. The aim of this study was to characterize the transporters on the apical side of the small intestine that are responsible for the rapid absorption of nateglinide. It has been reported that the uptake of fluorescein by Caco-2 cells occurs via an H+-driven transporter and that the intestinal fluorescein transporter is probably not MCT1. We examined the contribution of the fluorescein transporter to the uptake of nateglinide by Caco-2 cells. Fluorescein competitively inhibited H+-dependent nateglinide uptake. All of fluorescein transporter inhibitors examined reduced the uptake of nateglinide. Furthermore, nateglinide inhibited fluorescein uptake. We conclude that the intestinal nateglinide/H+ cotransport system is identical to the intestinal fluorescein/H+ cotransport system.

H+-dependent transport mechanism of nateglinide in the brush-border membrane of the rat intestine
S Itagaki, Y Saito, S Kubo, Y Otsuka, Y Yamamoto, M Kobayashi, T Hirano, K Iseki
JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, 312, 1, 77, 82, AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS, 2005年01月, ［査読有り］
英語, 研究論文（学術雑誌）, (-)-N-(trans-4-Isopropylcyclohexanecarbonyl)-D-phenylalanine (nateglinide) is a novel oral hypoglycemic agent possessing a carboxyl group and a peptide-type bond in its structure. Although nateglinide quickly reaches the maximal serum concentration after oral administration, nateglinide itself is not transported by PepT1 or MCT1. The aim of this study was to characterize the transporters on the apical side of the small intestine that are responsible for the rapid absorption of nateglinide. The uptake of nateglinide by rat intestinal brush-border membrane vesicles is associated with a proton-coupled transport system. Ceftibuten competitively inhibited H+-dependent nateglinide uptake. Glycylsarcosine (Gly-Sar), cephradine, and cephalexin did not significantly inhibit the uptake of nateglinide. The combination of Gly-Sar and nateglinide greatly reduced the uptake of ceftibuten. The effect of the combined treatment was significantly greater than that of Gly-Sar alone. Furthermore, nateglinide competitively inhibited H+-driven ceftibuten transporter-mediated ceftibuten uptake. Ceftibuten transport occurs via at least two H+-dependent transport systems: one is PepT1, and the other is the ceftibuten/H+ cotransport system. On the other hand, we demonstrated that nateglinide transport occurs via a single system that is H+ dependent but is distinct from PepT1 and may be identical to the ceftibuten/ H+ cotransport system.

Mechanism of L-lactic acid transport in L6 skeletal muscle cells.
Kobayashi M, Itagaki S, Hirano T, Iseki K
Drug metabolism and pharmacokinetics, 19, 5, 363, 368, 2004年10月, ［査読有り］, ［筆頭著者］, ［国際誌］
英語, 研究論文（学術雑誌）, L-lactic acid transport plays an important role in the regulation of L-lactic acid circulation into and out of muscle. To clarify the transport mechanism of L-lactic acid in skeletal muscle, L-lactic acid uptake was investigated using a L6 cell line. mRNAs of monocarboxylate transporter (MCT) 1, 2 and 4 were found to be expressed in L6 cells. The [ (14)C] L-lactic acid uptake by L6 cells increased up to pH of 6.0. The [ (14)C] L-lactic acid uptake at pH 6.0 was concentration-dependent with a K(m) of 3.7 mM. This process was reduced by alpha-cyano-4-hydroxycinnamate, a typical MCT1, 2 and 4 inhibitor. These results suggest that an MCT participates in the uptake of L-lactic acid by L6 cells. [ (14)C] L-lactic acid uptake was markedly inhibited by monocarboxylic acids and monocarboxylate drugs but not by dicarboxylic acids and amino acids. Moreover, benzoic acid, a substrate for MCT1, competitively inhibited this process with K(i) of 1.7 mM. [ (14)C] L-lactic acid efflux in L6 cells was inhibited by alpha-cyano-4-hydroxycinnamate but not by benzoic acid. These results suggest that [ (14)C] L-lactic acid efflux in L6 cells is mediated by MCT other than MCT1.

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(公社)日本薬学会, 日本語

クロザピン誘発性流涎症のリスク因子の同定と発現メカニズムの解明　　　　　　　　　　　　　　　
石川修平, 小林正紀, 橋本直樹, 久住一郎, 第14回次世代を担う若手医療薬科学シンポジウム, 2020年11月

シスプラチンの抗腫瘍効果を増強させるNSAIDsの探索とシスプラチン起因性腎障害との関連　　　　　　　　　　　　　　　
岡本 敬介, 小林 正紀, 齋藤 佳敬, 古堅 彩子, 鳴海 克哉, 井関 健, 日本薬学会年会要旨集, 140年会, 27Y, am07S, 2020年03月
(公社)日本薬学会, 日本語

下顎埋伏智歯抜歯術におけるセフカペンピボキシルとアモキシシリンの手術部位感染予防効果の比較　　　　　　　　　　　　　　　
山神 彰, 小林 正紀, 山田 武宏, 北川 善政, 大廣 洋一, 佐藤 淳, 石黒 信久, 今井 俊吾, 武隈 洋, 菅原 満, 井関 健, 日本薬学会年会要旨集, 140年会, 28Z, am10S, 2020年03月
(公社)日本薬学会, 日本語

リバース・トランスレーショナルリサーチに基づいたシスプラチン起因性腎障害に 及ぼすNSAIDsの影響の解析　　　　　　　　　　　　　　　
岡本敬介, 齋藤佳敬, 小林正紀, 古堅彩子, 鳴海克哉, 井関健, 第29回日本医療薬学会年会講演要旨集, 2019年11月

データマイニング手法を用いた副作用発現リスクの定量的評価モデルの開発 ガンシンクロビル誘発性血小板減少症発現割合の推定モデルの構築
今井 俊吾, 山田 武宏, 笠師 久美子, 小林 正紀, 井関 健, 日本医薬品情報学会総会・学術大会講演要旨集, 22回, 83, 83, 2019年06月
(一社)日本医薬品情報学会, 日本語

下顎埋伏智歯抜歯術におけるセフカペンピボキシルとアモキシシリンの手術部位感染予防効果の比較
山神 彰, 山田 武宏, 北川 善政, 大廣 洋一, 佐藤 淳, 石黒 信久, 今井 俊吾, 小林 正紀, 井関 健, 医療薬学, 45, 5, 254, 261, 2019年05月
下顎埋伏智歯抜歯術においてセフカペンピボキシル(CFPN-PI)またはアモキシシリン(AMPC)の予防投与を受けた患者の手術部位感染(SSI)発生率を比較した。解析対象となったのはCFPN-PI群129例(男性50例、女性79例、中央値24.0歳)、AMPC群164例(男性75例、女性89例、中央値25.0歳)であった。SSI発生率はCFPN-PI群において11.6%(15/129)、AMPC群において2.4%(4/164)であり、CFPN-PI群のSSI発生率は有意に高かった。次いで、SSI発生に影響する要因分析を行った。単変量解析において抗菌薬(CFPN-PIの使用)、性別(女性)、術後入院、手術難易度、反対側下顎埋伏智歯抜歯の有無、Pell-Gregory分類がP≦0.2となり、これらを多変量解析における解析対象因子として抽出した。解析対象因子のvariance inflation factorはいずれも4未満であり多重共線性はみられなかった。多変量解析の結果、抗菌薬(CFPN-PIの使用)および術後入院がそれぞれオッズ比5.61、4.66を示し、SSIに影響する独立因子として抽出された。, (一社)日本医療薬学会, 日本語

ガンシクロビル誘発性血小板減少症の要因分析とDecision tree解析を用いたリスク推定モデルの構築
今井 俊吾, 山田 武宏, 笠師 久美子, 小林 正紀, 井関 健, 医薬品情報学, 21, 1, 9, 19, 2019年05月
ガンシクロビル(GCV)による血小板減少症の発現因子を従来の手法であるロジスティック回帰分析を用いて探索した。さらに、2種の決定木(DT)分析アルゴリズムを用いて血小板減少症発現割合の推定モデルを構築し、そのモデルの妥当性について検討した。GCVの点滴静注製剤の投与を受けた患者386例を対象とした。血小板減少症の発現は12.2%(386例中47例)に認められ、多変量解析において白血球数7000cells/mm3未満、血小板数101000cells/mm3未満、総ビリルビン値0.8mg/dL以上が血小板減少症発現に影響する独立因子として抽出された。CHAID法で構築したDTモデルでは、ツリーは3段階、六つのサブグループに分枝し、五つのリスク因子が抽出された。また、CRT法で構築したモデルでは、ツリーは3段階、七つのサブグループに分枝し、五つのリスク因子が抽出された。ロジスティック回帰モデルならびにCHAID法で構築されたDTモデルの精度は87.8%、CRT法で構築されたDTモデルの精度は88.3%であった。, (一社)日本医薬品情報学会, 日本語

病院・薬局薬剤師を対象としたターミナルケア態度尺度(FATCOD-Form B-J)を用いた意識調査
熊井 正貴, 加藤 信太郎, 小柳 遼, 久保田 康生, 古堅 彩子, 鳴海 克哉, 小林 正紀, 笠師 久美子, 土居 由有子, 郡 修徳, 井関 健, 日本緩和医療薬学雑誌 = Japanese journal of pharmaceutical palliative care and sciences, 12, 1, 1, 8, 2019年03月
日本緩和医療薬学会, 日本語

第2回領域別専門薬剤師によるclinical questionの解決手段と、薬物療法のエビデンス創出に向けて マグネシウム投与によるシスプラチン起因性腎障害の予防効果およびその機序の解明　　　　　　　　　　　　　　　
齋藤 佳敬, 小林 正紀, 岡本 敬介, 秋田 弘俊, 井関 健, 日本薬学会年会要旨集, 139年会, 1, 260, 260, 2019年03月
(公社)日本薬学会, 日本語

薬学実務実習前後における薬学生のコミュニケーション分析 RIAS(Roter method of interaction process analysis)を用いて　　　　　　　　　　　　　　　
武隈 洋, 森 綾子, 小林 正紀, 山田 勇磨, 佐藤 夕紀, 鳴海 克哉, 古堅 彩子, 菅原 満, 日本薬学会年会要旨集, 139年会, 4, 212, 212, 2019年03月
(公社)日本薬学会, 日本語

EFFECTS OF POLYUNSATURATED FATTY ACIDS ON THE EXPRESSION OF TRANSPORTERS IN HUMAN PLACENTAL CHORIOCARCINOMA CELLS
Kanako Ono, Ayako Furugen, Yuko Kurosawa, Naoko Jinno, Katsuya Narumi, Masaki Kobayashi, Ken Iseki, PLACENTA, 69, E87, E87, 2018年09月
W B SAUNDERS CO LTD, 英語, 研究発表ペーパー・要旨（国際会議）

後方視的臨床研究及びin vivoにおけるマグネシウムのシスプラチン起因性腎障害軽減効果の検証　　　　　　　　　　　　　　　
岡本敬介, 齋藤佳敬, 小林正紀, 井関健, 第12回次世代を担う若手医療薬科学シンポジウム要旨集, 2018年09月
研究発表ペーパー・要旨（全国大会，その他学術会議）

乳汁および血漿中におけるbenzodiazepine系薬剤のLC/MS/MS定量法の構築
古堅彩子, 西村あや子, 小林正紀, 馬詰武, 井関健, 井関健, 医療薬学フォーラム講演要旨集, 26th, 285, 2018年06月
日本語

GABA はラット小腸 I/R による腸管免疫変動を抑制する　　　　　　　　　　　　　　　
窪田 篤人, 小林 正紀, 更科 壮太, 保田 元気, 鳴海 克哉, 古堅 彩子, 鈴木 夏子, 井関 健, 第145回 日本薬学会北海道支部例会, 2018年05月

NSAIDsのCOX選択性がシスプラチン起因性細胞傷害に及ぼす影響
岡本敬介, 齋藤佳敬, 古堅彩子, 鳴海克哉, 小林正紀, 井関健, 井関健, 日本薬学会年会要旨集(CD-ROM), 138th, ROMBUNNO.27C‐am10S, 2018年
日本語

胎盤細胞への葉酸輸送に及ぼす抗てんかん薬の影響の評価
黒澤優子, 古堅彩子, 西村あや子, 鳴海克哉, 小林正紀, 井関健, 井関健, 日本薬学会年会要旨集(CD-ROM), 138th, ROMBUNNO.27T‐am10S, 2018年
日本語

2型糖尿病病態とMCT11遺伝子多型との関連性
木村有希, 樋口一世, 樋口一世, 小林正紀, 小林正紀, 鳴海克哉, 古堅彩子, 三好秀明, 中村昭伸, 渥美達也, 井関健, 井関健, 日本薬学会年会要旨集(CD-ROM), 138th, ROMBUNNO.27X‐pm02, 2018年
日本語

hMCT1,4の基質選択性に関わるアミノ酸残基の探索
二木悠哉, 二木悠哉, 古堅彩子, 鳴海克哉, 小林正紀, 井関健, 井関健, 日本薬学会年会要旨集(CD-ROM), 138th, ROMBUNNO.27U‐am06S, 2018年
日本語

リネゾリド投与患者における嘔吐発現の因果関係の検証
堤竹蔵, 今井俊吾, 堤佳那, 山田武宏, 笠師久美子, 小林正紀, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 28, 2018年

データマイニング手法を用いたバンコマイシンによる腎機能障害発現割合推定モデルの構築
今井俊吾, 山田武宏, 笠師久美子, 新沼悠介, 小林正紀, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 28, 2018年

病院・薬局薬剤師を対象としたターミナルケアに関する態度尺度(FATCOD-Form B-J)を用いた意識調査
熊井正貴, 加藤信太郎, 小柳遼, 久保田康生, 古堅彩子, 鳴海克哉, 小林正紀, 笠師久美子, 土居由有子, 郡修徳, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 28, 2018年

食品成分による記憶と腸管免疫の維持・改善に関する研究
小林正紀, 小林正紀, 高橋夏子, 井関健, 三島海雲記念財団年次報告書, 2017, 33, 2017年11月
日本語

エピルビシンを用いた肝動脈化学塞栓術（TACE ）施行時の制吐薬による消化器症状の予防効果の評価
坂本達彦, 齋藤佳敬, 小林正紀, 山田武宏, 笠師久美子, 中井正人, 小川浩司, 井関 健, 日本医療薬学会第27回年会講演要旨集, 27, 2017年11月
研究発表ペーパー・要旨（全国大会，その他学術会議）

医療薬学ブラッシュアップ講座 医薬品とトランスポーターに関する話題―4―
小林正紀, 道薬誌, 34, 7, 4‐8, 2017年07月01日
日本語

ビジュアル・アナログスケールを用いた「妊娠と薬相談外来」の評価
西村 あや子, 山田 崇弘, 小林 正紀, 笠師 久美子, 井関 健, 日本医薬品情報学会総会・学術大会講演要旨集, 20回, 121, 121, 2017年06月
(一社)日本医薬品情報学会, 日本語

シスプラチン起因性腎障害に対するマグネシウムの軽減効果の機序　　　　　　　　　　　　　　　
岡本敬介, 齋藤佳敬, 小林正紀, 清水康, 秋田弘俊, 井関健, 日本医療薬学会第1 回フレッシャーズ・カンファランス要旨集, 2017年06月
研究発表ペーパー・要旨（全国大会，その他学術会議）

医療薬学ブラッシュアップ講座 医薬品とトランスポーターに関する話題―3―
小林正紀, 道薬誌, 34, 5/6, 29‐33, 2017年05月01日
日本語

医療薬学ブラッシュアップ講座 医薬品とトランスポーターに関する話題―2―
小林正紀, 道薬誌, 34, 4, 4‐8, 2017年04月01日
日本語

実務実習前後における薬学生のRIASによるコミュニケーション分析　　　　　　　　　　　　　　　
森 綾子, 武隈 洋, 小林正紀, 山田勇磨, 佐藤夕紀, 鳴海克哉, 古堅彩子, 菅原 満, 総合技術研究会2017東京大学, 2017年03月08日
日本語, 研究発表ペーパー・要旨（全国大会，その他学術会議）

食品成分による腸管免疫の維持・改善に関する研究
窪田 篤人, 小林 正紀, 竹野 礼子, 更科 壮太, 高橋 夏子, 保田 元気, 古堅 彩子, 鳴海 克哉, 井関 健, 日本薬学会年会要旨集, 137年会, 4, 65, 65, 2017年03月
(公社)日本薬学会, 日本語

医療薬学ブラッシュアップ講座 医薬品とトランスポーターに関する話題―1―
小林正紀, 道薬誌, 34, 3, 4‐8, 2017年03月01日
日本語

Caco‐2細胞におけるOATP2B1基質輸送に及ぼすインスリンの影響
小林拓, 小林正紀, 鳴海克哉, 近藤安佑子, 古堅彩子, 井関健, 井関健, 日本薬剤学会年会講演要旨集(Web), 32nd, ROMBUNNO.11‐6‐18 (WEB ONLY), 2017年
日本語

Caco‐2細胞におけるロキソプロフェンの輸送特性
鳴海克哉, 小林正紀, 古堅彩子, 高橋夏子, 井関健, 井関健, 日本薬学会年会要旨集(CD-ROM), 137th, 4, ROMBUNNO.27PB‐am168, 177, 2017年
(公社)日本薬学会, 日本語

データマイニング手法を用いたガンシクロビル誘発性好中球減少症の要因分析
今井 俊吾, 山田 武宏, 小林 正紀, 笠師 久美子, 井関 健, 医療薬学, 43, 10, 552, 562, 2017年

Ganciclovir (GCV), an antiviral agent against cytomegalovirus infection, can cause hematological toxicity such as neutropenia. However, risk factors for neutropenia have not been well established. A decision tree (DT) model, which is a flowchart-like framework, is a data mining technique that is used to construct predictive models. In a previous study, we demonstrated its usefulness in defining prognostic factors in adverse drug reactions. Here, risk factors for GCV-induced neutropenia were analyzed using a DT model and a logistic regression model as a conventional method.

A single-center, retrospective study was conducted at Hokkaido University Hospital. A total of 369 patients administered GCV between April 2008 and March 2017 were enrolled. Neutropenia was defined as a decrease in absolute neutrophil count (ANC) of < 1,000 cells/mm³ and to a < 75％ decrease from the baseline. All data were retrospectively collected from the electronic medical records. In the DT analysis, a chi-squared automatic interaction detection algorithm was constructed. To evaluate the established algorithms, the accuracies of each model were calculated.

Forty (10.8％) patients developed neutropenia. Acquired immunodeficiency syndrome, hematopoietic stem cell transplantations, ANC < 6,679 cells/mm³, and duration of therapy ≥ 18 days were extracted to build the DT model, in which the patients were divided into five subgroups. Their incidence of neutropenia was 0.0 to 58.3％. The accuracies of each model were the same (89.7％), indicating that both were equally accurate.

The results show that a useful risk prediction model for GCV-induced neutropenia can be built using DT analysis.

, 一般社団法人日本医療薬学会, 日本語

データマイニング手法を用いたGanciclovir誘発性好中球減少症の要因分析
今井俊吾, 山田武宏, 小林正紀, 笠師久美子, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 27, 2017年

血液内科疾患の治療中に生じた発熱性好中球減少症に対するテイコプラニンのトラフ値と有効性の関連性
内山数貴, 田澤佑基, 今井俊吾, 鏡圭介, 杉田純一, 鳴海克哉, 山田武宏, 笠師久美子, 小林正紀, 豊嶋崇徳, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 27, 2017年

外来化学療法に対する疑義照会の新規観点別評価
川岸亨, 久保田康生, 原田幸子, 松浦泰寛, 菅原亮輔, 吉川佳那, 堤竹蔵, 笠師久美子, 山田武宏, 小林正紀, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 27, 2017年

プロクロルペラジンによるアカシジア発現および制吐効果に関する検討
熊谷卓博, 齋藤佳敬, 山田武宏, 小林正紀, 熊井正貴, 笠師久美子, 小松嘉人, 井関健, 日本医療薬学会年会講演要旨集(Web), 27, 2017年

α-Defensinが排出系トランスポータへ及ぼす影響
保田元気, 窪田篤人, 更科壮太, 小林正紀, 井関健, 井関健, 高橋夏子, 日本医療薬学会年会講演要旨集(Web), 27, 2017年

院外処方せんへのがん化学療法プロトコール名掲載の取り組み
久保田康生, 深井敏隆, 山田武宏, 小林正紀, 井関健, 北海道病院薬剤師会誌, 91, 5‐8, 8, 2016年11月25日
保険薬局が必要とする情報を収集し、分析・解析して、稼働後に近隣保険薬局と共同で実施した講習会の内容に反映させた。さらに講習会終了後に、運用上の問題点や評価を行うためのアンケート調査を実施した。院外処方せんへ化学療法プロトコール名の印字と、薬剤部ホームページを利用した標準治療プロトコールの公開を行った。注射抗がん薬の治療内容が含まれるプロトコールほど、その理解度は低いことが示された。化学療法に関して病院からの提供が望まれる情報については四つに分類することができ、1点目は患者への服薬指導に関する項目、2点目は病院における採用薬に関する項目、3点目はプロトコールの対象がん腫や、1コースの期間・休薬期間に関する項目、4点目は患者個人に関する項目であった。アンケート結果をもとに、講習会での情報の提供内容を決定した。, (一社)北海道病院薬剤師会, 日本語

モノカルボン酸トランスポータ4の役割に着目した疾患と副作用に関する研究
小林正紀, 医療薬学, 42, 7, 485‐491, 491, 2016年07月10日

Statins, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, are the most widely used cholesterol-lowering agents for prevention of obstructive cardiovascular events. However, statins can cause a variety of skeletal muscle problems including myopathy and rhabdomyolysis, which is characterized by an elevation of creatine kinase (CK). The occurrence of myopathy has been estimated to range from 1% to 5%. The prevalence of muscular symptoms is as high as 25% among statin users who exercise. However, the mechanism by which statins cause myopathy is not precisely known. Since exercise induces the expression of monocarboxylate transporter (MCT) 4, we focused on the association between MCT4 and statin-induced muscle injury.
This review summarizes our findings:
1) MCT4 is a pH-dependent transporter and contributes to lactate release in RD cells as a model of in vitro skeletal muscle. AMP-activated protein kinase (AMPK) activated by exercise, increased MCT4 expression level and lactate efflux by MCT4 in RD cells and rat skeletal muscle.
2) Statins reduce the number of viable cells and up-regulate MCT4 in RD cells. MCT4 knockdown suppresses statin-induced reduction of cell viability and apoptosis compared with negative control-treated RD cells.
The cytotoxicity of cerivastatin, one of the statins, is associated with intracellular acidification and caspase-3/7 activation.
3) Bicarbonate prevents cerivastatin-induced pH alteration, cytotoxicity in RD cells and muscle damage in an in vivo study.
These results suggest that both increased expression of MCT4 and disturbance of skeletal muscle cell pH homeostasis may play an important role in statin-induced cytotoxicity.

, 一般社団法人日本医療薬学会, 日本語

Decision tree modelの副作用発現リスク因子解析への応用可能性に関する研究
今井 俊吾, 山田 武宏, 笠師 久美子, 小林 正紀, 井関 健, 日本医薬品情報学会総会・学術大会講演要旨集, 19回, 83, 83, 2016年05月
(一社)日本医薬品情報学会, 日本語

Decision tree modelの医薬品副作用発現リスク因子解析への応用可能性に関する研究．　　　　　　　　　　　　　　　
今井俊吾, 山田武宏, 笠師久美子, 小林正紀, 井関健, 第63回北海道薬学大会 北海道病院薬剤師会会員研究発表会, 2016年05月

マグネシウムによるシスプラチン起因性腎障害の軽減効果の機序に関する研究　　　　　　　　　　　　　　　
岡本敬介, 齋藤佳敬, 小林正紀, 清水 康, 秋田弘俊, 井関 健, 日本薬学会北海道支部第143回例会要旨集, 2016年05月
研究発表ペーパー・要旨（全国大会，その他学術会議）

hMCT1,4を介した基質輸送における細胞内ループの役割
二木悠哉, 佐々木将太郎, 小林正紀, 井関健, 日本薬学会年会要旨集(CD-ROM), 136th, ROMBUNNO.27Q‐PM01S, 2016年
日本語

経口抗がん剤が腸管免疫に与える影響
高橋夏子, 保田元気, 窪田篤人, 小林正紀, 井関健, 井関健, 日本医療薬学会年会講演要旨集(Web), 26, 2016年

INVOLVEMENT OF NON-CONSERVED HISTIDINE RESIDUE HIS-382 IN PH REGULATION OF HUMAN SLC16A3 ACTIVITY
Shotaro Sasaki, Masaki Kobayashi, Ken Iseki, DRUG METABOLISM REVIEWS, 47, 286, 286, 2015年11月
TAYLOR & FRANCIS LTD, 英語, 研究発表ペーパー・要旨（国際会議）

INVOLVEMENT OF MONOCARBOXYLATE TRANSPORTER 4 IN STATIN-INDUCED CYTOTOXICITY
Masaki Kobayashi, Yurika Kikutani, Shotaro Sasaki, Ken Iseki, DRUG METABOLISM REVIEWS, 47, 180, 181, 2015年11月
TAYLOR & FRANCIS LTD, 英語, 研究発表ペーパー・要旨（国際会議）

MCTの役割に着目した疾患と副作用に関する研究
小林正紀, 日本医療薬学会年会講演要旨集, 25th, 11, 11, 2015年10月23日
日本医療薬学会, 日本語

P0988-23-AM オキサリプラチンによる血管痛発現における患者背景の調査(がん薬物療法(他の副作用対策)2,ポスター,一般演題,医療薬学の進歩と未来-次の四半世紀に向けて-)
坂本 和央, 久保田 康生, 速水 章一, 大江 利治, 齋藤 佳敬, 熊井 正貴, 笠師 久美子, 山田 武宏, 小松 嘉人, 柴山 良彦, 小林 正紀, 井関 健, 日本医療薬学会年会講演要旨集, 25, 430, 430, 2015年10月23日
日本医療薬学会, 日本語

大豆イソフラボンによるモノカルボン酸輸送担体の阻害効果を利用したがん細胞増殖の回避戦略
小林正紀, 佐々木将太郎, 二木悠哉, 関口穂乃佳, 鳴海克哉, 高橋夏子, 井関健, 井関健, 日本未病システム学会学術総会抄録集, 22nd, 122, 122, 2015年09月10日
(一社)日本未病学会, 日本語

スポーツと医学(医療),スポーツ薬理学~スポーツにおける適切な薬物治療への支援~3.薬学部における運動(スポーツ)と薬に関する教育
小林正紀, 小林正紀, 医薬ジャーナル, 51, 7, 1723, 1725, 2015年07月01日
日本語

高グルコース環境培養下での大腸癌細胞株のハロゲン置換有機酸感受性の変化
井手野将也, 佐々木将太郎, 小林正紀, 二木悠哉, 井関健, トランスポーター研究会年会抄録集, 10th, 39, 2015年06月20日
日本語

新規抗てんかん薬のLC/MS/MS定量法の確立とヒト胎盤絨毛癌由来BeWo細胞における取り込み機構の解析
古堅 彩子, 西村 あや子, 高村 茂生, 小林 正紀, 山田 武宏, 笠師 久美子, 井関 健, 日本薬学会年会要旨集, 135年会, 4, 117, 117, 2015年03月
(公社)日本薬学会, 日本語

マグネシウムの経静脈前投薬によるシスプラチン起因性腎障害の予防効果の検討
齋藤 佳敬, 小林 正紀, 山田 武宏, 笠師 久美子, 秋田 弘俊, 井関 健, 日本薬学会年会要旨集, 135年会, 4, 59, 59, 2015年03月
(公社)日本薬学会, 日本語

糖尿病病態と乳酸及びアラニンの関連性
樋口 一世, 木村 有希, 鳴海 克哉, 小林 正紀, 三好 秀明, 中村 昭伸, 渥美 達也, 笠師 久美子, 山田 武宏, 井関 健, 日本薬学会年会要旨集, 135年会, 4, 89, 89, 2015年03月
(公社)日本薬学会, 日本語

持続血液ろ過透析療法中のLevofloxacinの薬物動態解析
和田剛志, 小林正紀, 方波見謙一, 小野雄一, 前川邦彦, 柳田雄一郎, 早川峰司, 澤村淳, 井関健, 井関健, 丸藤哲, 日本集中治療医学会学術集会(Web), 42nd, DO18-5 (WEB ONLY), 2015年
日本語

SLC16A1発現抑制による細胞内グルタチオン量の変化
佐々木将太郎, 木村有希, 小林正紀, 二木悠哉, 井関健, 日本薬学会年会要旨集(CD-ROM), 135th, ROMBUNNO.26U-PM04, 2015年
日本語

P1204-23-AM 妊娠可能年齢の女性における新世代抗てんかん薬の使用状況調査(妊婦・授乳婦,ポスター発表,一般演題,医療薬学の進歩と未来-次の四半世紀に向けて-)
西村 あや子, 高村 茂生, 古堅 彩子, 黒川 瞳, 小林 正紀, 笠師 久美子, 山田 武宏, 井関 健, 日本医療薬学会年会講演要旨集, 25, 0, 466, 466, 2015年
一般社団法人 日本医療薬学会, 日本語

PHARMACOKINETICS OF LEVOFLOXACIN IN CRITICALLY-ILL PATIENTS RECEIVING CONTINUOUS HEMODIAFILTRATION
Takeshi Wada, Masaki Kobayashi, Daisuke Miyamoto, Yuichiro Yanagida, Mineji Hayakawa, Atsushi Sawamura, Ken Iseki, Satoshi Gando, CRITICAL CARE MEDICINE, 42, 12, 2014年12月
LIPPINCOTT WILLIAMS & WILKINS, 英語, 研究発表ペーパー・要旨（国際会議）

ダプトマイシンは骨格筋細胞におけるモノカルボン酸輸送担体の発現を低下させる　　　　　　　　　　　　　　　
石川 修平, 山田 武宏, 鳴海 克哉, 小林 正紀, 山口 浩明, 井関 健, 日本薬学会年会要旨集, 134年会, 3, 63, 63, 2014年03月
(公社)日本薬学会, 日本語

hMCT1を介したコール酸輸送機構の解析
二木悠哉, 佐々木将太郎, 小林正紀, 井関健, 日本薬学会年会要旨集(CD-ROM), 134th, ROMBUNNO.29V-PM12S, 2014年
日本語

SLC16A1を介したプロチレリン代謝物輸送への遺伝子多型(rs1049434)の影響
佐々木将太郎, 二木悠哉, 小林正紀, 井関健, 日本薬剤学会年会講演要旨集(Web), 29th, 20E1-5 (WEB ONLY), 2014年
日本語

担体介在非飽和性グルタミン輸送の評価
佐々木将太郎, 小林正紀, 二木悠哉, 小西徹, 井関健, 生体膜と薬物の相互作用シンポジウム講演要旨集, 35th, 82, 83, 2013年11月21日
日本語

抗MRSA薬ダプトマイシンの骨格筋細胞毒性に関する基礎的検討　　　　　　　　　　　　　　　
石川 修平, 山田 武宏, 鳴海 克哉, 小林 正紀, 山口 浩明, 井関 健, 日本薬学会年会要旨集, 133年会, 3, 169, 169, 2013年03月
(公社)日本薬学会, 日本語

モノカルボン酸輸送担体hMCT4の発現調節機構
小林正紀, 鳴海克哉, 大竹翔, 佐々木将太郎, 井関健, トランスポーター研究会年会抄録集, 8th, 80, 2013年
日本語

モノカルボン酸輸送担体hMCT1,4の機能解析
佐々木将太郎, 小林正紀, 二木悠哉, 井関健, トランスポーター研究会年会抄録集, 8th, 81, 2013年
日本語

小腸虚血再還流により傍細胞経路からの尿酸の排泄は亢進するが, Bcrp を介した経細胞経路からの排泄は低下する
小倉 次郎, 高橋 夏子, 小林 正紀, 山口 浩明, 井関 健, 北海道醫學雜誌 = Acta medica Hokkaidonensia, 88, 1, 42, 42, 2013年01月01日
日本語

医療現場で新しい薬剤師職能を開拓する活動(病院オペ室での活動)と教員としての研究教育活動
小林正紀, 山田勇磨, 佐藤夕紀, 井関健, 日本医療薬学会年会講演要旨集, 22nd, 221, 221, 2012年10月10日
日本医療薬学会, 日本語

手術部サテライトファーマシーにおける麻酔薬の無菌混合調製体制の確立
山崎浩二郎, 須田範行, 新沼朋美, 水口貴史, 小林正紀, 山田勇磨, 山下美妃, 柴山良彦, 山田武宏, 井関健, 日本薬学会年会要旨集, 132nd, 4, 218, 218, 2012年03月05日
(公社)日本薬学会, 日本語

PKCを介した糖代謝制御およびMCT4発現変動への関連性
大竹翔, 鳴海克哉, 小林正紀, 山口浩明, 井関健, 日本医療薬学会年会講演要旨集, 21st, 161, 161, 2011年09月09日
日本医療薬学会, 日本語

抗菌薬アミカシンの用法用量別にみた治療効果の比較　　　　　　　　　　　　　　　
鈴木 貴宏, 川岸 亨, 小倉 次郎, 小林 正紀, 山口 浩明, 山田 武宏, 井関 健, 日本薬学会年会要旨集, 131年会, 4, 177, 177, 2011年03月
(公社)日本薬学会, 日本語

カラムスイッチング-LC/MS/MSによるパクリタキセル血中濃度測定法の確立　　　　　　　　　　　　　　　
藤川 あす歌, 山口 浩明, 伊藤 創, 田中 伸明, 古堅 彩子, 宮森 和明, 高橋 夏子, 小倉 次郎, 小林 正紀, 山田 武宏, 眞野 成康, 井関 健, 日本薬学会年会要旨集, 131年会, 4, 178, 178, 2011年03月
(公社)日本薬学会, 日本語

臨床検査値の変動から推測されるスタチン系薬物由来の副作用発生率の解析研究(一般演題(口頭)43,有害事象・副作用(1),Enjoy Pharmacists' Lifestyles)
落合 愛美, 小林 正紀, 川岸 亨, 笠師 久美子, 山口 浩明, 山田 武宏, 井関 健, 日本医療薬学会年会講演要旨集, 21, 0, 179, 179, 2011年
一般社団法人 日本医療薬学会, 日本語

II型糖尿病モデルラットにおける排出系トランスポーター発現変動に関する研究
渡辺恵布, 小林正紀, 小倉次郎, 山口浩明, 井関健, 日本医療薬学会年会講演要旨集, 20th, 266, 266, 2010年10月25日
日本医療薬学会, 日本語

AMPK活性化が骨格筋MCT1,4発現量に及ぼす影響
古堅彩子, 鳴海克哉, 小林正紀, 板垣史郎, 井関健, 日本薬学会年会要旨集, 130th, 4, 161, 2010年03月05日
日本語

北海道大学薬学部における実務実習事前実習の取り組みとその評価
武隈 洋, 小林 正紀, 山田 勇磨, 板垣 史郎, 吉田 和幸, 井関 健, 菅原 満, 日本薬学会年会要旨集, 130年会, 4, 346, 346, 2010年03月
(公社)日本薬学会, 日本語

骨格筋における糖代謝制御~PKCを標的とした制御メカニズム~
鳴海克哉, 古堅彩子, 渡辺恵布, 小林正紀, 板垣史郎, 井関健, 日本医療薬学会年会講演要旨集, 19th, 262, 262, 2009年09月15日
日本医療薬学会, 日本語

北海道大学病院における内服薬疑義照会率の傾向と分析
齋藤佳敬, 志賀弘康, 小林正紀, 須田範行, 菅原満, 井関健, 日本医療薬学会年会講演要旨集, 19th, 281, 2009年09月15日
日本語

胃内pH変動が薬物動態に及ぼす影響―吸収予測システムによる評価―
渡辺祐子, 井上直樹, 小林正紀, 板垣史郎, 井関健, 日本医療薬学会年会講演要旨集, 19th, 262, 262, 2009年09月15日
日本医療薬学会, 日本語

O9-007 がん化学療法効果減弱に寄与するP-gpの影響(一般演題 口頭発表,感染制御/持参薬管理/院内製剤/高齢者ケア/その他,医療薬学の創る未来 科学と臨床の融合)
黒沼 圭, 小林 正紀, 高橋 夏子, 板垣 史郎, 井岡 健, 日本医療薬学会年会講演要旨集, 19, 0, 272, 272, 2009年09月15日
日本医療薬学会, 日本語

O13-003 北海道大学病院における内服薬疑義照会率の傾向と分析(一般演題 口頭発表,調剤・処方鑑査・リスクマネジメント/医薬品情報・データベース/有害事象・副作用,医療薬学の創る未来 科学と臨床の融合)
齋藤 佳敬, 志賀 弘康, 小林 正紀, 須田 範行, 菅原 満, 井関 健, 日本医療薬学会年会講演要旨集, 19, 0, 281, 281, 2009年09月15日
日本医療薬学会, 日本語

ルテインの消化管における吸収挙動:トランスポータの関与
佐藤夕紀, 澁谷智美, 小林正紀, 板垣史郎, 野田敏宏, 水野智, 井関健, 薬剤学, 69, Supplement, 98, 2009年04月30日
日本語

小腸虚血再灌流によるP‐gp発現変動に対する食餌の影響
小倉次郎, 藤川あす歌, 丸山創, 小林正紀, 板垣史郎, 井関健, 薬剤学, 69, Supplement, 115, 2009年04月30日
日本語

がん化学療法に伴う口内炎の予防・改善に有効な抗酸化物質の探索
鷹野 瑠美, 平野 剛, 中田 千絵, 笠師 久美子, 菅原 満, 小林 正紀, 板垣 史郎, 井関 健, 医療薬学, 35, 4, 247, 253, 2009年04月
癌化学療法に伴う口内炎の予防・改善に有効な抗酸化物質を探索するため、種々の化合物についてスーパーオキシドアニオン消去作用を評価した。対象物は含嗽として汎用されている3薬剤、抗酸化作用が記載されている8薬剤、カテキンのEGCgを用いた。その結果、含嗽剤ではアロプリノールの消去作用がレバミピドと比較して非常に強く、カモスタットはほとんど消去作用を示さなかった。他剤ではトラニラスト、メサラジン、ピロキシカムは作用が強く、ガベキサート、L-カルボシステインは弱かった。また、EGCgはアロプリノールと同等の強い消去作用を示した。次いで、組織障害に関与するラジカル連鎖反応停止作用を調べた結果、含嗽3薬剤には同作用は認めず、メサラジン、EGCgには強力な作用を認めた。更に、酸化障害性口内炎モデルに対する細胞障害抑制作用を調べたところ、EGCgは強力な効果を、アロプリノール、メサラジンは弱い効果を示し、レバミピド、カモスタットには効果を認めなかった。, (一社)日本医療薬学会, 日本語

多発性骨髄腫における同種造血幹細胞移植後の慢性消化管GVHDに対しベクロメタゾン腸溶性製剤を投与した症例
久保田 康生, 小林 正紀, 笠師 久美子, 高畑 むつみ, 菅原 満, 橋野 聡, 井関 健, 医療薬学, 35, 3, 213, 218, 2009年03月
57歳女。多発性骨髄腫を発症した。MP(メルファラン+プレドニゾロン)療法を開始し、ビスホスホネート製剤を併用したが、病勢のコントロールが不良であった。サリドマイド+デキサメタゾン療法を開始したが奏効しなかった。自己末梢血幹細胞移植は、十分なCD34陽性細胞数を得ることができず施行できなかった。サリドマイド併用によるCP(シクロホスファミド+プレドニゾロン)療法、サリドマイド併用による少量のシクロホスファミドの内服治療を順次施行したが、出血性膀胱炎などの副作用により継続できなかった。非血縁者間同種造血幹細胞移植を施行し、GVHD予防としてFK506の持続点滴を開始した。内視鏡検査と病理検査により回盲部と結腸における急性消化管GVHDの発症を確認した。プロピオン酸ベクロメタゾン(BDP)腸溶性カプセル剤を投与し、改善傾向を認めた。, (一社)日本医療薬学会, 日本語

乳がん化学療法プロトコルの投与スケジュールに着目した評価系の確立
林えり子, 平野剛, 小林正紀, 板垣史郎, 井関健, 日本医療薬学会年会講演要旨集, 18th, 331, 331, 2008年09月01日
日本医療薬学会, 日本語

吸収予測システムを用いた後発医薬品の薬剤学的同等性評価(2)~BCS分類Class IおよびIIIの製剤における比較~
渡辺祐子, 山本千秋, 下山哲哉, 小林正紀, 板垣史郎, 菅原満, 平野剛, 井関健, 日本医療薬学会年会講演要旨集, 18th, 310, 310, 2008年09月01日
一般社団法人 日本医療薬学会, 日本語

消化管吸収予測システムを用いた後発医薬品の薬剤学的同等性評価(1)
下山哲哉, 渡辺祐子, 山本千秋, 小林正紀, 板垣史郎, 菅原満, 平野剛, 井関健, 日本医療薬学会年会講演要旨集, 18th, 267, 267, 2008年09月01日
一般社団法人 日本医療薬学会, 日本語

放射線療法に伴う口腔粘膜障害の評価系の確立
中田千絵, 平野剛, 鷹野瑠美, 小林正紀, 板垣史郎, 井関健, 日本医療薬学会年会講演要旨集, 18th, 252, 252, 2008年09月01日
日本医療薬学会, 日本語

小腸虚血再灌流障害時における小腸吸収動態の変化
丸山創, 小倉次郎, 小林正紀, 板垣史郎, 平野剛, 井関健, 日本医療薬学会年会講演要旨集, 18th, 265, 265, 2008年09月01日
日本医療薬学会, 日本語

肝切除による有機アニオントランスポーター発現変動と遠隔臓器への影響
藤井達郎, 板垣史郎, 小林正紀, 平野剛, 井関健, 日本医療薬学会年会講演要旨集, 18th, 265, 265, 2008年09月01日
日本医療薬学会, 日本語

ルテイン長期投与による肺蓄積機構の解明
澁谷智美, 佐藤夕紀, 小林正紀, 板垣史郎, 平野剛, 井関健, 日本医療薬学会年会講演要旨集, 18th, 280, 280, 2008年09月01日
日本医療薬学会, 日本語

20D-28 放射線療法に伴う口腔粘膜障害の評価系の確立(がん薬物療法(副作用対策),来るべき時代への道を拓く)
中田 千絵, 平野 剛, 鷹野 瑠美, 小林 正紀, 板垣 史郎, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 252, 252, 2008年09月01日
日本医療薬学会, 日本語

20I-24 小腸虚血再灌流障害時における小膓吸収動態の変化(薬物動態(基礎),来るべき時代への道を拓く)
丸山 創, 小倉 次郎, 小林 正紀, 板垣 史郎, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 265, 265, 2008年09月01日
日本医療薬学会, 日本語

20I-25 肝切除による有機アニオントランスポーター発現変動と遠隔臓器への影響(薬物動態(基礎),来るべき時代への道を拓く)
藤井 達郎, 板垣 史郎, 小林 正紀, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 265, 265, 2008年09月01日
日本医療薬学会, 日本語

21C-12 消化管吸収予測システムを用いた後発医薬品の薬剤学的同等性評価(1)(後発医薬品,来るべき時代への道を拓く)
下山 哲哉, 渡辺 祐子, 山本 千秋, 小林 正紀, 板垣 史郎, 菅原 満, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 267, 267, 2008年09月01日
日本医療薬学会, 日本語

21I-13 ルテイン長期投与による肺蓄積機構の解明(薬物動態(基礎・臨床・遺伝子多型),来るべき時代への道を拓く)
澁谷 智美, 佐藤 夕紀, 小林 正紀, 板垣 史郎, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 280, 280, 2008年09月01日
日本医療薬学会, 日本語

20-P1-168 吸収予測システムを用いた後発医薬品の薬剤学的同等性評価(2) : BCS分類class IおよびIIIの製剤における比較(後発医薬品,来るべき時代への道を拓く)
渡辺 祐子, 山本 千秋, 下山 哲哉, 小林 正紀, 板垣 史郎, 菅原 満, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 310, 310, 2008年09月01日
日本医療薬学会, 日本語

20-P2-290 乳がん化学療法プロトコルの投与スケジュールに着目した評価系の確立(がん薬物療法(その他),来るべき時代への道を拓く)
林 えり子, 平野 剛, 小林 正紀, 板垣 史郎, 井関 健, 日本医療薬学会年会講演要旨集, 18, 0, 331, 331, 2008年09月01日
日本医療薬学会, 日本語

スタチン系薬物間における筋障害およびコレステロール低下作用の比較
知嵜郁美, 小林正紀, 板垣史郎, 平野剛, 井関健, J Toxicol Sci, 33, Supplement, S169, 2008年06月
日本語

N‐モノデスエチルアミオダロンの肺細胞への取り込み機構と肺障害性との関連
関悟, 板垣史郎, 小林正紀, 平野剛, 井関健, 薬剤学, 68, Supplement, 268, 2008年04月30日
日本語

北大病院における内服薬疑義照会率の傾向と分析
齋藤 佳敬, 榊原 則寛, 志賀 弘康, 沖 洋充, 小林 正紀, 川合 真次, 深井 敏隆, 菅原 満, 井関 健, 日本薬学会年会要旨集, 128年会, 4, 163, 163, 2008年03月
(公社)日本薬学会, 日本語

20D-30 がん化学療法における口内炎の予防に対する抗酸化物質の応用(がん薬物療法(副作用対策),来るべき時代への道を拓く)
鷹野 瑠美, 平野 剛, 中田 千絵, 笠師 久美子, 菅原 満, 小林 正紀, 板垣 史郎, 井関 健, 大江 利治, 日本医療薬学会年会講演要旨集, 18, 0, 253, 253, 2008年
一般社団法人 日本医療薬学会, 日本語

20D-30 がん化学療法における口内炎の予防に対する抗酸化物質の応用(がん薬物療法(副作用対策),来るべき時代への道を拓く)
鷹野 瑠美, 平野 剛, 中田 千絵, 笠師 久美子, 菅原 満, 小林 正紀, 板垣 史郎, 井関 健, 大江 利治, 日本医療薬学会年会講演要旨集, 18, 0, 253, 253, 2008年
一般社団法人 日本医療薬学会, 日本語

29-C1-10-2 Coenzyme Q10の消化管吸収挙動:排出系トランスポーターの関与(薬物相互作用・薬物動態,社会の期待に応える医療薬学を)
落合 彰子, 小林 正紀, 板垣 史郎, 平野 剛, 菅原 満, 井関 健, 日本医療薬学会年会講演要旨集, 17, 196, 196, 2007年09月01日
日本医療薬学会, 日本語

29-C1-10-4 ルテインの吸収動態の解明と酸化障害抑制効果に対する検討(薬物相互作用・薬物動態,社会の期待に応える医療薬学を)
佐藤 夕紀, 澁谷 智美, 小林 正紀, 板垣 史郎, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 17, 196, 196, 2007年09月01日
日本医療薬学会, 日本語

30-A2-13-5 小腸虚血再灌流によるABCトランスポーター発現変動(がん薬物療法・薬物動態,社会の期待に応える医療薬学を)
小倉 次郎, 小林 正紀, 板垣 史郎, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 17, 199, 199, 2007年09月01日
日本医療薬学会, 日本語

細胞内シグナル伝達系を介したMCT1,4の調節機構
鳴海克哉, 小林正紀, 香川俊樹, 板垣史郎, 平野剛, 井関健, 薬剤学, 67, Supplement, 260, 260, 2007年05月10日
日本語

ABCG2プロモータ活性に及ぼす性ホルモンの影響
安田哲, 小林正紀, 板垣史郎, 平野剛, 井関健, 薬剤学, 67, Supplement, 258, 258, 2007年05月10日
日本語

トランスポーター活性制御によるアミオダロンの肺障害性の軽減
関 悟, 板垣 史郎, 小林 正紀, 平野 剛, 井関 健, 薬剤学 = Journal of Pharmaceutical Science and Technology, Japan, 67, 244, 244, 2007年05月10日
日本語

30-D-05 多発性骨髄腫患者に対するサリドマイド経口投与後の両エナンチオマー血中濃度測定方法の確立(癌薬物療法(外来化学療法、緩和ケア等),医療薬学の扉は開かれた)
千本木 要, 田中 雅則, 櫻田 惠右, 小林 正紀, 板垣 史郎, 平野 剛, 井関 健, 日本医療薬学会年会講演要旨集, 16, 0, 315, 315, 2006年09月01日
日本医療薬学会, 日本語

Pravastatinおよびrosuvastatinの骨格筋細胞障害性にpH変動が及ぼす影響
小林正紀, 香川俊樹, 鷹野瑠美, 板垣史郎, 平野剛, 井関健, J Toxicol Sci, 31, Supplement, S113, 2006年06月10日
日本語

ヒトPEPT1の安定大量発現系の構築とその有用性
藤沢佑紀, 館岡涼子, 宮内正二, 久保さやか, 大塚幸夫, 小林正紀, 井関健, 奈良敏文, 加茂直樹, 生体膜と薬物の相互作用シンポジウム講演要旨集, 23rd, 205, 208, 2001年11月08日
日本語

医療概論　　　　　　　　　　　　　　　
2020年04月

薬物治療学Ⅲ　　　　　　　　　　　　　　　
2020年04月

薬局実習　　　　　　　　　　　　　　　
北海道大学薬学部

病院実習　　　　　　　　　　　　　　　
北海道大学薬学部

救命救急実習　　　　　　　　　　　　　　　
北海道大学薬学部

実務実習事前実習　　　　　　　　　　　　　　　
北海道大学薬学部

臨床薬学事前実習　　　　　　　　　　　　　　　
北海道大学薬学部

OSCE対応演習　　　　　　　　　　　　　　　
北海道大学薬学部

臨床薬物動態解析演習　　　　　　　　　　　　　　　
北海道大学薬学部

医療情報解析演習　　　　　　　　　　　　　　　
北海道大学薬学部

薬物治療学Ⅱ　　　　　　　　　　　　　　　
北海道大学薬学部

エネルギー源輸送を担うMCTsの機能解析と治療薬の探索
科学研究費助成事業 基盤研究(C)
2020年04月01日 - 2023年03月31日
小林 正紀
本邦において高齢者に頻発する糖尿病や認知症等の疾患に対する有効な予防・治療法の開発が求められている。この対策として栄養介入による効果を期待するものが数多く報告され、これは生体内のエネルギー源の重要性を示唆するものである。最近エネルギー源として認知されてきた乳酸やピルビン酸とこれらを輸送するヒトモノカルボン酸輸送担体（hMCT）は、上記疾患と関連することが示唆されている。しかしながらhMCTの機能および生体内の役割については未だ不明な点が多い。そこで本研究ではhMCTの中でもhMCT2, 11に着目し、細胞における機能・発現を明らかにすることで、これらを標的とした化合物の探索を行う。昨年度は、hMCT2の評価系の樹立に成功したことから、今年度はhMCT11の評価系の構築を目的とするとともに、臨床研究における知見との比較・考察を試みた。
日本学術振興会, 基盤研究(C), 北海道大学, 20K07171

食品成分による分子標的薬のリスク回避と免疫予測マーカーの探索
科学研究費助成事業
2017年04月01日 - 2021年03月31日
鈴木 夏子, 小林 正紀
本研究では、分子標的薬曝露後のalpha defensin (Defa)5および toll-like receptor (TLR) 4発現量の変動、Defa5の産生を促進する食品成分の探索、さらに分子標的薬による腸管免疫低下に対する食品成分の有用性を評価した。分子標的薬の曝露により、Defa5およびTLR4タンパク質発現量は有意に減少した。また、食品成分曝露によるDefa等への影響については、一部の食品成分でDefa5およびTLR4タンパク質発現量を増加させることを見出した。最後に、食品成分を併用することで、分子標的薬による腸管免疫低下を回復する可能性が示唆された。
日本学術振興会, 基盤研究(C), 17K00871

食品成分の抗酸化作用に着目した記憶・腸管免疫賦活作用の機序に関する研究　　　　　　　　　　　　　　　
2019年研究助成
2019年 - 2020年
小林正紀
一般財団法人 旗影会, 研究代表者, 競争的資金

食品成分のα-ディフェンシン分泌促進作用による小腸移植後感染症の予防戦略
科学研究費助成事業 基盤研究(C)
2016年04月01日 - 2019年03月31日
井関 健, 小林 正紀
本研究では、小腸虚血再還流(I/R)後のalpha-defensin (Defa)5産生量の変動、Defa5産生促進物質の探索、食品成分のDefa5産生促進作用による小腸I/R予防効果の3点について行ってきた。最初の検討により小腸I/R処置において小腸絨毛の顕著な脱落と炎症性マーカーの上昇を確認し、本条件下においてDefa5 mRNA量が有意に低下することを見出した。一方で一部の食品成分がDefa5 mRNA量を増大させることを明らかとした。最後に本食品成分をI/R前に前投与することで、I/Rによる腸管免疫低下ならびに絨毛の脱落を軽減させることを見出した。
日本学術振興会, 基盤研究(C), 北海道大学, 16K08388

コーヒー成分が記憶形成と腸管免疫に及ぼす影響に関する研究　　　　　　　　　　　　　　　
2018年助成対象研究
2018年 - 2019年
小林正紀
一般社団法人 全日本コーヒー協会, 研究代表者, 競争的資金

食品成分による記憶と腸管免疫の維持・改善に関する研究　　　　　　　　　　　　　　　
平成28年度学術研究奨励金
2016年 - 2017年
小林正紀
公益財団法人 三島海雲記念財団, 研究代表者, 競争的資金

MCTの機能・発現阻害に基づいたがん細胞の転移・浸潤の抑制効果の検証　　　　　　　　　　　　　　　
基盤研究(C)
2015年 - 2017年
小林正紀
独立行政法人 日本学術振興会, 研究代表者, 競争的資金

胆汁酸の輸送特性を反映した胎児毒性の回避戦略　　　　　　　　　　　　　　　
2015年度 研究助成
2015年 - 2016年
小林正紀
公益財団法人 秋山記念生命科学振興財団, 研究代表者, 競争的資金

5 -オキソプロリンの輸送担体を利用したグリア細胞の酸化ストレス抵抗性に関する研究　　　　　　　　　　　　　　　
2015年 研究助成
2015年 - 2016年
小林正紀
一般財団法人 旗影会, 研究代表者, 競争的資金

薬剤性筋障害の早期発見を目的としたバイオマーカーの探索と臨床応用　　　　　　　　　　　　　　　
平成26年度研究助成
2014年 - 2015年
小林正紀
一般財団法人 横山臨床薬理研究助成基金, 研究代表者, 競争的資金

記憶の維持・改善を指向した食品成分によるエネルギー源輸送の阻害・促進メカニズムの解明　　　　　　　　　　　　　　　
平成26年度公募研究
2014年 - 2015年
小林正紀
一般財団法人 糧食研究会, 研究代表者, 競争的資金

大豆ポリフェノールによる乳酸輸送担体の阻害を利用したがんの浸潤・転移の回避戦略　　　　　　　　　　　　　　　
平成26年度 研究助成
2014年 - 2014年
小林正紀
タカノ財団, 研究代表者, 競争的資金

乳酸輸送体の機能解析と個別化治療へ向けた臨床応用に関する研究　　　　　　　　　　　　　　　
若手研究人材・ネットワーク育成補助金
2013年 - 2014年
小林正紀
ノーステック財団, 研究代表者, 競争的資金

乳酸輸送担体(MCT)の基質認識性の解明および機能解析に基づく薬剤性筋障害制圧へ向けた新規戦略　　　　　　　　　　　　　　　
若手研究者自立支援
2013年 - 2013年
小林正紀
北海道大学重点配分経費, 研究代表者, 競争的資金

筋MCTの役割に着目した薬剤性筋障害の回避戦略　　　　　　　　　　　　　　　
科学研究費補助金(若手研究(B))
2012年 - 2013年
小林正紀
文部科学省, 研究代表者, 競争的資金

トランスポータの発現変動を利用した薬物濃度調節　　　　　　　　　　　　　　　
第23回中富健康科学振興財団・研究助成金
2010年 - 2011年
小林正紀
公益財団法人 中富健康科学振興財団, 研究代表者, 競争的資金

骨格筋細胞内pH調節因子の評価培養系モデルの構築　　　　　　　　　　　　　　　
笹川科学研究助成
2009年 - 2010年
小林正紀
公益財団法人 日本科学協会, 研究代表者, 競争的資金

糖尿病治療を標的としたGLUTおよびMCT発現調節機構解明　　　　　　　　　　　　　　　
若手研究人材・ネットワーク育成補助金
2008年 - 2009年
小林正紀
ノーステック財団, 研究代表者, 競争的資金

高脂血症薬の副作用回避ストラテジーの構築～新規薬物相互作用の解明～
科学研究費補助金(若手研究(B))
2008年 - 2009年
小林正紀
本研究はトランスポータの発現制御を担う核内受容体LXR(liver X receptor)に着目し、LXRの活性に影響を及ぼす脂質異常症治療薬(スタチン)投与時の薬物動態変動の可能性を明らかにし、薬物-薬物相互作用の要因の一端を明らかにすることを目的としている。本研究により脂質異常症に対する安全かつ適正な薬物治療の一助となる。
文部科学省, 若手研究(B), 北海道大学, 研究代表者, 競争的資金, 20790126