Nakajima M

References (61)

Title : Role of carboxylesterase and arylacetamide deacetylase in drug metabolism, physiology, and pathology - Nagaoka_2024_Biochem.Pharmacol__116128
Author(s) : Nagaoka M , Sakai Y , Nakajima M , Fukami T
Ref : Biochemical Pharmacology , :116128 , 2024
Abstract : Carboxylesterases (CES1 and CES2) and arylacetamide deacetylase (AADAC), which are expressed primarily in the liver and/or gastrointestinal tract, hydrolyze drugs containing ester and amide bonds in their chemical structure. These enzymes often catalyze the conversion of prodrugs, including the COVID-19 drugs remdesivir and molnupiravir, to their pharmacologically active forms. Information on the substrate specificity and inhibitory properties of these enzymes, which would be useful for drug development and toxicity avoidance, has accumulated. Recently,in vitroandin vivostudies have shown that these enzymes are involved not only in drug hydrolysis but also in lipid metabolism. CES1 and CES2 are capable of hydrolyzing triacylglycerol, and the deletion of their orthologous genes in mice has been associated with impaired lipid metabolism and hepatic steatosis. Adeno-associated virus-mediated human CES overexpression decreases hepatic triacylglycerol levels and increases fatty acid oxidation in mice. It has also been shown that overexpression of CES enzymes or AADAC in cultured cells suppresses the intracellular accumulation of triacylglycerol. Recent reports indicate that AADAC can be up- or downregulated in tumors of various organs, and its varied expression is associated with poor prognosis in patients with cancer. Thus, CES and AADAC not only determine drug efficacy and toxicity but are also involved in pathophysiology. This review summarizes recent findings on the roles of CES and AADAC in drug metabolism, physiology, and pathology.
ESTHER : Nagaoka_2024_Biochem.Pharmacol__116128
PubMedSearch : Nagaoka_2024_Biochem.Pharmacol__116128
PubMedID: 38492781
Gene_locus related to this paper: human-AADAC , human-CES1 , human-CES2

Title : Evaluation of drug-drug interactions via inhibition of hydrolases by orlistat, an anti-obesity drug - Hirosawa_2023_Drug.Metab.Dispos__
Author(s) : Hirosawa K , Fukami T , Nakano M , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , : , 2023
Abstract : Drug-drug interactions (DDI) have a significant impact on drug efficacy and safety. It has been reported that orlistat, an anti-obesity drug, inhibits the hydrolysis of p-nitrophenol acetate, a common substrate of the major drug-metabolizing hydrolases, carboxylesterase (CES) 1, CES2, and arylacetamide deacetylase (AADAC), in vitro The aim of this study was to examine whether orlistat affects the pharmacokinetics of drug(s) metabolized by hydrolases in vivo, after evaluating its inhibitory potencies against CES1, CES2, and AADAC in vitro Orlistat potently inhibited the hydrolysis of acebutolol, a specific substrate of CES2, in a non-competitive manner (K (i) = 2.95 {plus minus} 0.16 nM), whereas it slightly inhibited the hydrolysis of temocapril and eslicarbazepine acetate, specific substrates of CES1 and AADAC, respectively (IC(50) > 100 nM). The in vivo DDI potential was elucidated using mice, in which orlistat showed strong inhibition against acebutolol hydrolase activities in the liver and intestinal microsomes, similar to humans. The AUC of acebutolol was increased by 43%, whereas the AUC of acetolol, a hydrolyzed metabolite of acebutolol, was decreased by 47% by co-administration of orlistat. The ratio of the K (i) value to the maximum unbound plasma concentration of orlistat (< 0.012) is lower than the risk criteria for DDI in the liver defined by the FDA guideline (> 0.02), whereas the ratio of the K (i) value to the estimated intestinal luminal concentration (3.3 x 10(5)) is considerably higher than the risk criteria in the intestine (> 10). Therefore, this suggests that orlistat causes DDI by inhibiting hydrolases in the intestine. Significance Statement This study demonstrated that orlistat, an anti-obesity drug, causes drug-drug interactions in vivo by potently inhibiting carboxylesterase 2 in the intestine. This is the first evidence that inhibition of hydrolases causes drug-drug interactions.
ESTHER : Hirosawa_2023_Drug.Metab.Dispos__
PubMedSearch : Hirosawa_2023_Drug.Metab.Dispos__
PubMedID: 37137721

Title : Characterization of Enzymes Involved in Nintedanib Metabolism in Humans - Nakashima_2023_Drug.Metab.Dispos_51_733
Author(s) : Nakashima S , Sato R , Fukami T , Kudo T , Hashiba S , Morinaga G , Nakano M , Ludwig-Schwellinger E , Matsui A , Ishiguro N , Ebner T , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 51 :733 , 2023
Abstract : Nintedanib, which is used to treat idiopathic pulmonary fibrosis and non-small cell lung cancer, is metabolized to a pharmacologically inactive carboxylate derivative, BIBF1202, via hydrolysis and subsequently by glucuronidation to BIBF1202 acyl-glucuronide (BIBF1202-G). Since BIBF1202-G contains an ester bond, it can be hydrolytically cleaved to BIBF1202. In this study, we sought to characterize these metabolic reactions in the human liver and intestine. Nintedanib hydrolysis was detected in human liver microsomes (HLMs) (Clearance [CL (int)]: 102.8 +/- 18.9 microL/min per mg protein) but not in small intestinal preparations. CES1 was suggested to be responsible for nintedanib hydrolysis according to experiments using recombinant hydrolases and hydrolase inhibitors as well as proteomic correlation analysis using 25 individual HLM. BIBF1202 glucuronidation in HLM (3.6 +/- 0.3 microL/min per mg protein) was higher than that in human intestinal microsomes (1.5 +/- 0.06 microL/min per mg protein). UGT1A1 and gastrointestinal UGT1A7, UGT1A8, and UGT1A10 were able to mediate BIBF1202 glucuronidation. The impact of UGT1A1 on glucuronidation was supported by the finding that liver microsomes from subjects homozygous for the UGT1A1*28 allele showed significantly lower activity than those from subjects carrying the wild-type UGT1A1 allele. Interestingly, BIBF1202-G was converted to BIBF1202 in HLS9 at 70-fold higher rates than the rates of BIBF1202 glucuronidation. An inhibition study and proteomic correlation analysis suggested that beta-glucuronidase is responsible for hepatic BIBF1202-G deglucuronidation. In conclusion, the major metabolic reactions of nintedanib in the human liver and intestine were quantitatively and thoroughly elucidated. This information could be helpful to understand the inter- and intraindividual variability in the efficacy of nintedanib. SIGNIFICANCE STATEMENT: To our knowledge, this is the first study to characterize the enzymes responsible for each step of nintedanib metabolism in the human body. This study found that beta-glucuronidase may contribute to BIBF1202-G deglucuronidation.
ESTHER : Nakashima_2023_Drug.Metab.Dispos_51_733
PubMedSearch : Nakashima_2023_Drug.Metab.Dispos_51_733
PubMedID: 36927840

Title : PPARalpha regulates the expression of human arylacetamide deacetylase involved in drug hydrolysis and lipid metabolism - Morikawa_2022_Biochem.Pharmacol_199_115010
Author(s) : Morikawa T , Fukami T , Gotoh-Saito S , Nakano M , Nakajima M
Ref : Biochemical Pharmacology , 199 :115010 , 2022
Abstract : Human arylacetamide deacetylase (AADAC) hydrolyzes various drugs containing an acetyl group, such as ketoconazole and rifampicin. Knowledge about the role of human AADAC in drug metabolism is accumulating, but the regulatory mechanism of its expression has not been elucidated. In mice, it has been suggested that Aadac expression may be regulated by peroxisome proliferator-activated receptor alpha (Pparalpha). This study examined whether human AADAC is regulated by PPARalpha, which widely regulates the expression of lipid metabolism-related genes. In human hepatoma Huh-7 cells, AADAC mRNA and protein levels were significantly increased by treatment with fenofibric acid and WY-14643, PPARalpha ligands. Knockdown and overexpression of PPARalpha resulted in decreased and increased expression of AADAC, respectively. Luciferase assays revealed that the direct repeat 1 (DR1) at -193/-181 in the AADAC promoter region is responsible for transactivation by PPARalpha. Chromatin immunoprecipitation assays revealed the binding of PPARalpha to DR1. Thus, it was demonstrated that human AADAC is regulated by PPARalpha through binding to DR1. Oil red O staining showed that overexpression of AADAC in Huh-7 cells suppressed lipid accumulation after treatment with free fatty acids. The suppression was restored by treatment with diisopropyl fluorophosphate, an AADAC inhibitor. The WY-14643-mediated suppression of lipid accumulation was restored by AADAC knockdown. These results suggested that AADAC has a role in suppressing cellular lipid accumulation. In conclusion, this study demonstrated the regulation of human AADAC by PPARalpha and its significance in lipid accumulation.
ESTHER : Morikawa_2022_Biochem.Pharmacol_199_115010
PubMedSearch : Morikawa_2022_Biochem.Pharmacol_199_115010
PubMedID: 35314168

Title : Arylacetamide deacetylase knockout mice are sensitive to ketoconazole-induced hepatotoxicity and adrenal insufficiency - Nagaoka_2022_Biochem.Pharmacol_195_114842
Author(s) : Nagaoka M , Fukami T , Kisui F , Yamada T , Sakai Y , Tashiro K , Ogiso T , Konishi K , Honda S , Hirosawa K , Nakano M , Nakajima M
Ref : Biochemical Pharmacology , 195 :114842 , 2022
Abstract : Orally administered ketoconazole may rarely induce liver injury and adrenal insufficiency. A metabolite formed by arylacetamide deacetylase (AADAC)-mediated hydrolysis has been observed in cellulo studies, and it is relevant to ketoconazole-induced cytotoxicity. This study tried to examine the significance of AADAC in ketoconazole-induced toxicity in vivo using Aadac knockout mice. Oral administration of 150 mg/kg ketoconazole resulted in the area under the plasma concentration-time curve values of ketoconazole and N-deacetylketoconazole, a hydrolyzed metabolite of ketoconazole, in Aadac knockout mice being significantly higher and lower than those in wild-type mice, respectively. With the administration of ketoconazole (300 mg/kg/day) for 7 days, Aadac knockout mice showed higher mortality (100%) than wild-type mice (42.9%), and they also showed significantly higher plasma alanine transaminase and lower corticosterone levels, thus representing liver injury and steroidogenesis inhibition, respectively. It was suggested that a higher plasma ketoconazole concentration likely accounts for the inhibition of the synthesis of corticosterone, which has anti-inflammatory effects, in the adrenal gland in Aadac KO mice. In Aadac knockout mice, hepatic mRNA levels of immune- and inflammation-related factors were increased by the administration of 300 mg/kg ketoconazole, and the increase was restored by the replenishment of corticosterone (40 mg/kg, s.c.) along with recoveries of plasma alanine transaminase levels. In conclusion, Aadac defects exacerbate ketoconazole-induced liver injury by inhibiting glucocorticoid synthesis and enhancing the inflammatory response. This in vivo study revealed that the hydrolysis of ketoconazole by AADAC can mitigate ketoconazole-induced toxicities.
ESTHER : Nagaoka_2022_Biochem.Pharmacol_195_114842
PubMedSearch : Nagaoka_2022_Biochem.Pharmacol_195_114842
PubMedID: 34798123
Gene_locus related to this paper: mouse-aryla

Title : Role of human AADAC on hydrolysis of eslicarbazepine acetate and effects of AADAC genetic polymorphisms on hydrolase activity - Hirosawa_2021_Drug.Metab.Dispos__
Author(s) : Hirosawa K , Fukami T , Tashiro K , Sakai Y , Kisui F , Nakano M , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , : , 2021
Abstract : Human arylacetamide deacetylase (AADAC) plays a role in the detoxification or activation of drugs and is sometimes involved in the incidence of toxicity by catalyzing hydrolysis reactions. AADAC prefers compounds with relatively small acyl groups, such as acetyl groups. Eslicarbazepine acetate, an antiepileptic drug, is a prodrug rapidly hydrolyzed to eslicarbazepine. We sought to clarify whether AADAC might be responsible for the hydrolysis of eslicarbazepine acetate. Eslicarbazepine acetate was efficiently hydrolyzed by human intestinal and liver microsomes and recombinant human AADAC. The hydrolase activities in human intestinal and liver microsomes were inhibited by epigallocatechin gallate, a specific inhibitor of AADAC, by 82% and 88% of the control, respectively. The hydrolase activities in liver microsomes from 25 human livers were significantly correlated (r = 0.87, P < 0.001) with AADAC protein levels, suggesting that the enzyme AADAC is responsible for the hydrolysis of eslicarbazepine acetate. The effects of genetic polymorphisms of AADAC on eslicarbazepine acetate hydrolysis were examined by using the constructed recombinant AADAC variants with T74A, V172I, R248S, V281I, N366K, or X400Q. AADAC variants with R248S or X400Q showed lower activity than wild type (5% or 21%, respectively), whereas those with V172I showed higher activity than wild type (174%). Similar tendencies were observed in the other 4 substrates of AADAC; that is, p-nitrophenyl acetate, ketoconazole, phenacetin, and rifampicin. Collectively, we found that eslicarbazepine acetate is specifically and efficiently hydrolyzed by human AADAC, and several AADAC polymorphic alleles would be a factor affecting the enzyme activity and drug response. Significance Statement This is the first study to clarify that AADAC is responsible for the activation of eslicarbazepine acetate, an antiepileptic prodrug, to eslicarbazepine, an active form, in the human liver and intestines. In addition, we found that several AADAC polymorphic alleles would be a factor affecting the enzyme activity and drug response.
ESTHER : Hirosawa_2021_Drug.Metab.Dispos__
PubMedSearch : Hirosawa_2021_Drug.Metab.Dispos__
PubMedID: 33446525
Gene_locus related to this paper: human-AADAC

Title : Hydrolase activities of cynomolgus monkey liver microsomes and recombinant CES1, CES2, and AADAC - Honda_2021_Eur.J.Pharm.Sci__105807
Author(s) : Honda S , Fukami T , Tsujiguchi T , Zhang Y , Nakano M , Nakajima M
Ref : Eur J Pharm Sci , :105807 , 2021
Abstract : The cynomolgus monkey is a nonhuman primate that is often used for pharmacokinetic and toxicokinetic studies of new chemical entities. Species differences in drug metabolism are obstacles for the extrapolation of animal data to humans. This study aimed to characterize hydrolase activities for typical compounds by cynomolgus monkey liver microsomes and recombinant monkey carboxylesterases (CES1 and CES2) and arylacetamide deacetylase (AADAC) compared with the activities in humans. To estimate the contribution of each hydrolase, the ratios of the expression level of each hydrolase in the liver microsomes and recombinant systems were used. For almost all of the tested human CES1 substrates, hydrolase activities in cynomolgus monkey liver microsomes tended to be lower than those in human liver microsomes, and recombinant cynomolgus monkey CES1 showed catalytic activity, but not for all substrates. For human CES2 substrates, hydrolase activities in cynomolgus monkey liver were higher than those in human liver microsomes, and recombinant monkey CES2 was responsible for their hydrolysis. Among human AADAC substrates, phenacetin was mainly hydrolyzed by monkey AADAC, whereas indiplon and ketoconazole were hydrolyzed by AADAC and other unknown enzymes. Flutamide was hydrolyzed by monkey CES2, not by AADAC. Rifamycins were hardly hydrolyzed in monkey liver microsomes. In conclusion, this study characterized the hydrolase activities of cynomolgus monkeys compared with those in humans. The findings would be helpful for pharmacokinetic or toxicokinetic studies of new chemical entities whose main metabolic pathway is hydrolysis.
ESTHER : Honda_2021_Eur.J.Pharm.Sci__105807
PubMedSearch : Honda_2021_Eur.J.Pharm.Sci__105807
PubMedID: 33722734

Title : Epicatechin gallate and epigallocatechin gallate are potent inhibitors of human arylacetamide deacetylase - Yasuda_2021_Drug.Metab.Pharmacokinet_39_100397
Author(s) : Yasuda K , Watanabe K , Fukami T , Nakashima S , Ikushiro SI , Nakajima M , Sakaki T
Ref : Drug Metab Pharmacokinet , 39 :100397 , 2021
Abstract : Recently, in addition to carboxylesterases (CESs), we found that arylacetamide deacetylase (AADAC) plays an important role in the metabolism of some clinical drugs. In this study, we screened for food-related natural compounds that could specifically inhibit human AADAC, CES1, or CES2. AADAC, CES1, and CES2 activities in human liver microsomes were measured using phenacetin, fenofibrate, and procaine as specific substrates, respectively. In total, 43 natural compounds were screened for their inhibitory effects on each of these enzymes. Curcumin and quercetin showed strong inhibitory effects against all three enzymes, whereas epicatechin, epicatechin gallate (ECg), and epigallocatechin gallate (EGCg) specifically inhibited AADAC. In particular, ECg and EGCg showed strong inhibitory effects on AADAC (IC(50) values: 3.0 +/- 0.5 and 2.2 +/- 0.2 microM, respectively). ECg and EGCg also strongly inhibited AADAC-mediated rifampicin hydrolase activity in human liver microsomes with IC(50) values of 2.2 +/- 1.4 and 1.7 +/- 0.4 microM, respectively, whereas it weakly inhibited p-nitrophenyl acetate hydrolase activity, which is catalyzed by AADAC, CES1, and CES2. Our results indicate that ECg and EGCg are potent inhibitors of AADAC.
ESTHER : Yasuda_2021_Drug.Metab.Pharmacokinet_39_100397
PubMedSearch : Yasuda_2021_Drug.Metab.Pharmacokinet_39_100397
PubMedID: 34171773

Title : Differences in hydrolase activities in the liver and small intestine between marmosets and humans - Honda_2021_Drug.Metab.Dispos__
Author(s) : Honda S , Fukami T , Hirosawa K , Tsujiguchi T , Zhang Y , Nakano M , Uehara S , Uno Y , Yamazaki H , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , : , 2021
Abstract : For drug development, species differences in drug-metabolism reactions present obstacles for predicting pharmacokinetics in humans. We characterized the species differences in hydrolases among humans and mice, rats, dogs, and cynomolgusmonkeys. In this study, to expand the series of such studies, we attempted to characterize marmoset hydrolases. We measured hydrolase activities for 24 compounds using marmoset liver and intestinal microsomes, as well as recombinant marmoset carboxylesterase (CES) 1, CES2, and arylacetamide deacetylase (AADAC). The contributions of CES1, CES2, and AADAC to hydrolysis in marmoset liver microsomes were estimated by correcting the activities by using the ratios of hydrolase protein levels in the liver microsomes and those in recombinant systems. For 6 out of 8 human CES1 substrates,the activities in marmoset liver microsomes were lower than those in human liver microsomes. For 2 human CES2 substrates and 3 out of 7 human AADAC substrates, the activities in marmoset liver microsomes were higher than those in human liver microsomes. Notably, among the 3 rifamycins, only rifabutin was hydrolyzed by marmoset tissue microsomes and recombinant AADAC. The activities for all substrates in marmoset intestinal microsomes tended to be lower than those in liver microsomes, which suggests that the first-pass effects of the CES and AADAC substrates are dueto hepatic hydrolysis. In most cases, the sums of the values of the contributions of CES1, CES2, and AADAC were below 100%, which indicated the involvement of other hydrolases in marmosets. In conclusion, we clarified the substrate preferences of hydrolases in marmosets. Significance Statement This study confirmed that there are large differences in hydrolase activities between humans and marmosets by characterizing marmoset hydrolase activities for compounds that are substrates of human CES1, CES2, or AADAC. The data obtained in this study may be useful for considering whether marmosets are appropriate for examining the pharmacokinetics and efficacies of new chemical entities in preclinical studies.
ESTHER : Honda_2021_Drug.Metab.Dispos__
PubMedSearch : Honda_2021_Drug.Metab.Dispos__
PubMedID: 34135089

Title : m(6)A modification impacts hepatic drug and lipid metabolism properties by regulating carboxylesterase 2 - Takemoto_2021_Biochem.Pharmacol__114766
Author(s) : Takemoto S , Nakano M , Fukami T , Nakajima M
Ref : Biochemical Pharmacology , :114766 , 2021
Abstract : Methylation of adenosine at the N(6) position to form N(6)-methyladenosine (m(6)A) is the most prevalent epitranscriptomic modification of mammalian mRNA. This modification is catalyzed by a methyltransferase-like 3 (METTL3)-METTL14 complex and is erased by demethylases such as fat mass and obesity-associated protein (FTO) or AlkB homolog 5 (ALKBH5). m(6)A modification regulates mRNA stability, nuclear export, splicing, and/or protein translation via recognition by reader proteins such as members of YT521-B homology (YTH) family. Carboxylesterase 2 (CES2) is a serine esterase responsible for the hydrolysis of drugs and endogenous substrates, such as triglycerides and diacylglycerides. Here, we examined the potential regulation of human CES2 expression by m(6)A modification. CES2 mRNA level was significantly increased by double knockdown of METTL3 and METTL14 but was decreased by knockdown of FTO or ALKBH5 in HepaRG and HepG2 cells, leading to changes in its protein level and hydrolase activity for 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), suggesting that m(6)A modification negatively regulates CES2 expression. Consistent with the changes in CES2 expression, lipid accumulation in the cells was decreased by double knockdown of METTL3 and METTL14 but was increased by knockdown of FTO or ALKBH5. RNA immunoprecipitation assays using an anti-m(6)A antibody showed that adenosines in the 5'-untranslated region (UTR) and the last exon of CES2 are methylated. Luciferase assays revealed that YTHDC2, which degrades m(6)A-containing mRNA, downregulates CES2 expression by recognition of m(6)A in the 5'-UTR of CES2. Collectively, we demonstrated that m(6)A modification has a great impact on the regulation of CES2, affecting pharmacokinetics, drug response and lipid metabolism.
ESTHER : Takemoto_2021_Biochem.Pharmacol__114766
PubMedSearch : Takemoto_2021_Biochem.Pharmacol__114766
PubMedID: 34536357

Title : Arylacetamide deacetylase as a determinant of the hydrolysis and activation of abiraterone acetate in mice and humans - Sakai_2021_Life.Sci_284_119896
Author(s) : Sakai Y , Fukami T , Nagaoka M , Hirosawa K , Ichida H , Sato R , Suzuki K , Nakano M , Nakajima M
Ref : Life Sciences , 284 :119896 , 2021
Abstract : AIM: Abiraterone acetate for metastatic castration-resistant prostate cancer is an acetylated prodrug to be hydrolyzed to abiraterone. Abiraterone acetate is known to be hydrolyzed by pancreatic cholesterol esterase secreted into the intestinal lumen. This study aimed to investigate the possibility that arylacetamide deacetylase (AADAC) expressed in enterocytes contributes to the hydrolysis of abiraterone acetate based on its substrate preference. MATERIALS AND METHODS: Abiraterone acetate hydrolase activity was measured using human intestinal (HIM) and liver microsomes (HLM) as well as recombinant AADAC. Correlation analysis between activity and AADAC expression was performed in 14 individual HIMs. The in vivo pharmacokinetics of abiraterone acetate was examined using wild-type and Aadac knockout mice administered abiraterone acetate with or without orlistat, a pancreatic cholesterol esterase inhibitor. KEY FINDINGS: Recombinant AADAC showed abiraterone acetate hydrolase activity with similar K(m) value to HIM and HLM. The positive correlation between activity and AADAC levels in individual HIMs supported the responsibility of AADAC for abiraterone acetate hydrolysis. The area under the plasma concentration-time curve (AUC) of abiraterone after oral administration of abiraterone acetate in Aadac knockout mice was 38% lower than that in wild-type mice. The involvement of pancreatic cholesterol esterase in abiraterone formation was revealed by the decreased AUC of abiraterone by coadministration of orlistat. Orlistat potently inhibited AADAC, implying its potential as a perpetrator of drug-drug interactions. SIGNIFICANCE: AADAC is responsible for the hydrolysis of abiraterone acetate in the intestine and liver, suggesting that concomitant use of abiraterone acetate and drugs potently inhibiting AADAC should be avoided.
ESTHER : Sakai_2021_Life.Sci_284_119896
PubMedSearch : Sakai_2021_Life.Sci_284_119896
PubMedID: 34450168
Gene_locus related to this paper: mouse-aryla

Title : Strain and sex differences in drug hydrolase activities in rodent livers - Kisui_2019_Eur.J.Pharm.Sci__105143
Author(s) : Kisui F , Fukami T , Nakano M , Nakajima M
Ref : Eur J Pharm Sci , :105143 , 2019
Abstract : Carboxylesterase (CES) 1, CES2, and arylacetamide deacetylase (AADAC) are the major drug hydrolases in humans, and they have different substrate preferences. Because rodents are widely used in preclinical studies, we aimed to clarify the extent of the species, strain, and sex differences in hydrolase activity in rats and mice. Hydrolase activities for 24 compounds were evaluated in Fischer 344, Sprague-Dawley, and Wistar-Imamichi rat liver microsomes (RLM) and Balb/c, C3H/He, C57BL/6J, and ddY mouse liver microsomes (MLM) by comparing the results with the activities in human liver microsomes (HLM). Imidapril hydrolase activities in RLM from all strains were substantially higher than those in MLM and HLM, whereas oseltamivir was hardly hydrolyzed in rodents, although both are specific substrates of CES1 in humans. In rats, males tended to show higher hydrolase activities for most human CES1 substrates than females. Hydrolase activities for irinotecan and procaine, which are CES2 substrates in humans, tended to be higher in RLM and MLM than in HLM. Rifamycins, substrates of human AADAC, were not hydrolyzed in RLM and MLM. The results of this study provide important information about the species, strain, and sex differences in hydrolase activities in rats and mice.
ESTHER : Kisui_2019_Eur.J.Pharm.Sci__105143
PubMedSearch : Kisui_2019_Eur.J.Pharm.Sci__105143
PubMedID: 31726091

Title : Difference in substrate specificity of carboxylesterase and arylacetamide deacetylase between dogs and humans - Yoshida_2018_Eur.J.Pharm.Sci_111_167
Author(s) : Yoshida T , Fukami T , Kurokawa T , Gotoh S , Oda A , Nakajima M
Ref : Eur J Pharm Sci , 111 :167 , 2018
Abstract : Carboxylesterase (CES) and arylacetamide deacetylase (AADAC) are the major enzymes responsible for the hydrolysis of various clinical drugs. Our recent study demonstrated that the identity of the responsible hydrolase can be roughly surmised based on the chemical structures of compounds in humans. Dogs are used for preclinical studies in drug development, but the substrate specificities of dog CES and AADAC remain to be clarified. The purpose of this study is to characterize their substrate specificities. We prepared recombinant dog CES1, CES2, and AADAC. p-Nitrophenyl acetate, a general substrate for esterases, was hydrolyzed by dog CES1 and AADAC, while it was not hydrolyzed by CES2. CES2 protein was not substantially detected in the recombinant system or in the dog liver and intestinal microsomes by Western blot using anti-human CES2 antibodies. In silico analyses demonstrated slight differences in the three-dimensional structures of dog CES2 and human CES2, indicating that dog CES2 might be unstable or inactive. By evaluating the hydrolase activities of 22 compounds, which are known to be substrates of human CES and/or AADAC, we found that the activities of dog recombinant CES1 and AADAC as well as dog tissue preparations for nearly all compounds were lower than those of human enzymes. The dog enzymes that were responsible for the hydrolysis of most compounds corresponded to the human enzymes, but the following differences were observed: oseltamivir, irinotecan, and rifampicin were not hydrolyzed in the dog liver or by any of the recombinant esterases and procaine, a human CES2 substrate, was hydrolyzed by dog CES1. In conclusion, the present study could provide new finding to facilitate our understanding of species differences in drug hydrolysis, which can facilitate drug development and drug safety evaluation.
ESTHER : Yoshida_2018_Eur.J.Pharm.Sci_111_167
PubMedSearch : Yoshida_2018_Eur.J.Pharm.Sci_111_167
PubMedID: 28966098
Gene_locus related to this paper: canfa-CESDD1 , canfa-f1p6w8 , canlf-e2r2h2

Title : Characterization of a helminthosporic acid analog that is a selective agonist of gibberellin receptor - Miyazaki_2018_Bioorg.Med.Chem.Lett_28_2465
Author(s) : Miyazaki S , Tomita K , Yamane H , Kobayashi M , Asami T , Nakajima M
Ref : Bioorganic & Medicinal Chemistry Lett , 28 :2465 , 2018
Abstract : Helminthosporol, a natural growth regulator isolated from a fungus, stimulates hypocotyl growth and seed germination, similar to gibberellin (GA). We recently reported that helminthosporic acid (H-acid), a synthetic analog of helminthosporol, acts as an agonist of GA receptor. In this study, we showed that a H-acid analog, in which the hydroxymethyl group at the C-8 position of H-acid was converted to a keto group, acts as a selective GA receptor agonist. 1) This analog shows higher hypocotyl elongation activity in Arabidopsis than H-acid does, and induces the degradation of DELLA protein and 2) leads to the formation of the GID1-DELLA complex and 3) regulates the expression of GA-related genes. In addition, 4) its hypocotyl elongation activity was not observed in a atgid1a single mutant, and 5) this analog could promote only the interaction between specific GA receptors and DELLA proteins in vitro. Taken together, our results strongly suggest that the selectivity of the reported H-acid analog depends on the specificity of its GA receptor binding activity.
ESTHER : Miyazaki_2018_Bioorg.Med.Chem.Lett_28_2465
PubMedSearch : Miyazaki_2018_Bioorg.Med.Chem.Lett_28_2465
PubMedID: 29907394

Title : In vitro approach to elucidate the relevance of carboxylesterase 2 and N-acetyltransferase 2 to flupirtine-induced liver injury - Konishi_2018_Biochem.Pharmacol_155_242
Author(s) : Konishi K , Fukami T , Ogiso T , Nakajima M
Ref : Biochemical Pharmacology , 155 :242 , 2018
Abstract : The use of flupirtine, an analgesic, has been restricted in European countries because it causes liver injury in rare cases. Flupirtine is primarily metabolized to D-13223, an acetylamino form. In the process of D-13223 formation, it has been hypothesized that a reactive metabolite is formed which may be involved in flupirtine hepatotoxicity. The purpose of this study was to identify the potential reactive metabolite and the responsible enzymes in the human liver to get a clue to the mechanism of hepatotoxicity. Using recombinant enzymes, we found that D-13223 was formed from flupirtine via hydrolysis by carboxylesterase 2 (CES2) and subsequent acetylation by N-acetyltransferase (NAT) 2. A conjugate of N-acetyl-l-cysteine (NAC), a nucleophile, was detected by incubation of flupirtine with CES2, and the conjugate formation in human liver microsomes was inhibited by CES2 inhibitors, indicating that a reactive metabolite, which may be a quinone diimine, was produced in the process of CES2-mediated hydrolysis of flupirtine. The formation of the NAC conjugate in liver S9 samples from NAT2 slow acetylators was significantly higher than that from NAT2 rapid/intermediate acetylators, indicating that NAT2 could function as a detoxification enzyme for flupirtine. CES2-overexpressing HepG2 cells showed remarkable lactate dehydrogenase leakage under flupirtine treatment, while no cytotoxicity was observed in control cells, suggesting that the reactive metabolite formed by CES2-mediated hydrolysis of flupirtine would be a trigger of hepatotoxicity. NAT2 slow acetylators with high CES2 activity could be highly susceptible to flupirtine-induced liver injury.
ESTHER : Konishi_2018_Biochem.Pharmacol_155_242
PubMedSearch : Konishi_2018_Biochem.Pharmacol_155_242
PubMedID: 30028988

Title : Identification of enzymes responsible for nitrazepam metabolism and toxicity in human - Konishi_2017_Biochem.Pharmacol_140_150
Author(s) : Konishi K , Fukami T , Gotoh S , Nakajima M
Ref : Biochemical Pharmacology , 140 :150 , 2017
Abstract : Nitrazepam (NZP) is a hypnotic agent that rarely causes liver injuries in humans and teratogenicity in rodents. In humans, NZP is primarily metabolized to 7-aminonitrazepam (ANZP) by reduction and subsequently to 7-acetylamino nitrazepam (AANZP) by acetylation. ANZP can be regenerated from AANZP by hydrolysis in rodents, but it is still unclear whether this reaction occurs in humans. In rodents, AANZP may be associated with teratogenicity, while in humans, it is known that drug-induced liver injuries may be caused by NZP reactive metabolite(s). In this study, we attempted to identify the enzymes responsible for NZP metabolism to obtain a basic understanding of this process and the associated metabolite toxicities. We found that the NZP reductase activity in human liver cytosol (HLC) was higher than that in human liver microsomes (HLM). We purified the responsible enzyme(s) from HLC and found that the NZP reductase was aldehyde oxidase 1 (AOX1). The role of AOX1 was confirmed by an observed increase in the NZP reductase activity upon addition of N(1)-methylnicotinamide, an electron donor of AOX1, as well as inhibition of this activity in HLC in the presence of AOX1 inhibitors. ANZP was acetylated to form AANZP by N-acetyltransferase (NAT) 2. An experiment using recombinant esterases in an inhibition study using HLM revealed that AANZP is hydrolyzed by arylacetamide deacetylase (AADAC) in the human liver. N-Hydroxylamino NZP, which is suspected to be a reactive metabolite, was detected as a conjugate with N-acetyl-l-cysteine through NZP reduction and ANZP hydroxylation reactions. In the latter reaction, the conjugate was readily formed by recombinant CYP3A4 among the various P450 isoforms tested. In sum, we found that AOX1, NAT2, AADAC, and CYP3A4 are the determinants for the pharmacokinetics of NZP and that they confer interindividual variability in sensitivity to NZP side effects.
ESTHER : Konishi_2017_Biochem.Pharmacol_140_150
PubMedSearch : Konishi_2017_Biochem.Pharmacol_140_150
PubMedID: 28606603

Title : Human arylacetamide deacetylase hydrolyzes ketoconazole to trigger hepatocellular toxicity - Fukami_2016_Biochem.Pharmacol_116_153
Author(s) : Fukami T , Iida A , Konishi K , Nakajima M
Ref : Biochemical Pharmacology , 116 :153 , 2016
Abstract : Ketoconazole (KC), an antifungal agent, rarely causes severe liver injury when orally administered. It has been reported that KC is mainly hydrolyzed to N-deacetyl ketoconazole (DAK), followed by the N-hydroxylation of DAK by flavin-containing monooxygenase (FMO). Although the metabolism of KC has been considered to be associated with hepatotoxicity, the responsible enzyme(s) remain unknown. The purpose of this study was to identify the responsible enzyme(s) for KC hydrolysis in humans and to clarify their relevance to KC-induced toxicity. Kinetic analysis and inhibition studies using human liver microsomes (HLM) and recombinant enzymes revealed that human arylacetamide deacetylase (AADAC) is responsible for KC hydrolysis to form DAK, and confirmed that FMO3 is the enzyme responsible for DAK N-hydroxylation. In HLM, the clearance of KC hydrolysis occurred to the same extent as DAK N-hydroxylation, which indicates that both processes are not rate-limiting pathways. Cytotoxicity of KC and DAK was evaluated using HepaRG cells and human primary hepatocytes. Treatment of HepaRG cells with DAK for 24h showed cytotoxicity in a dose-dependent manner, whereas treatment with KC did not show due to the low expression of AADAC. Overexpression of AADAC in HepaRG cells with an adenovirus expression system elicited the cytotoxicity of KC. Cytotoxicity of KC in human primary hepatocytes was attenuated by diisopropylfluorophosphate, an AADAC inhibitor. In conclusion, the present study demonstrated that human AADAC hydrolyzes KC to trigger hepatocellular toxicity.
ESTHER : Fukami_2016_Biochem.Pharmacol_116_153
PubMedSearch : Fukami_2016_Biochem.Pharmacol_116_153
PubMedID: 27422753

Title : Arylacetamide Deacetylase is Responsible for Activation of Prasugrel in Human and Dog - Kurokawa_2016_Drug.Metab.Dispos_44_409
Author(s) : Kurokawa T , Fukami T , Yoshida T , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 44 :409 , 2016
Abstract : Prasugrel, a thienopyridine anti-platelet agent, is pharmacologically activated by hydrolysis and hydroxylation. It is efficiently hydrolyzed in the intestine after oral administration, and the enzyme responsible for the hydrolysis in humans was demonstrated to be carboxylesterase (CES)2. Prasugrel hydrolase activity is detected in dog intestines, where CES enzymes are absent; therefore, this prompted us to investigate the involvement of an enzyme(s) other than CES. Human arylacetamide deacetylase (AADAC) is highly expressed in the small intestine, catalyzing the hydrolysis of several clinical drugs containing small acyl moieties. In the present study, we investigated whether AADAC catalyzes prasugrel hydrolysis. Recombinant human AADAC was shown to catalyze prasugrel hydrolysis with a CLint value of 50.0 +/- 1.2 ml/min/mg protein with a similar Km value to human intestinal and liver microsomes, whereas the CLint values of human CES1 and CES2 were 4.6 +/- 0.1 and 6.6 +/- 0.3 ml/min/mg protein, respectively. Inhibition studies using various chemical inhibitors and the relative activity factor approach suggested that the contribution of AADAC to prasugrel hydrolysis in human intestine is comparable to that of CES2. In dog intestine, the expression of AADAC, but not CES1 and CES2, was confirmed by measuring the marker hydrolase activities of each human esterase. The similar Km values and inhibition profiles between recombinant dog AADAC and small intestinal microsomes suggest that AADAC is a major enzyme responsible for prasugrel hydrolysis in dog intestine. Collectively, we found that AADAC largely contributes to prasugrel hydrolysis in both human and dog intestine.
ESTHER : Kurokawa_2016_Drug.Metab.Dispos_44_409
PubMedSearch : Kurokawa_2016_Drug.Metab.Dispos_44_409
PubMedID: 26718653

Title : Comparison of substrate specificity among human arylacetamide deacetylase and carboxylesterases - Fukami_2015_Eur.J.Pharm.Sci_78_47
Author(s) : Fukami T , Kariya M , Kurokawa T , Iida A , Nakajima M
Ref : Eur J Pharm Sci , 78 :47 , 2015
Abstract : Human arylacetamide deacetylase (AADAC) is an esterase responsible for the hydrolysis of some drugs, including flutamide, indiplon, phenacetin, and rifamycins. AADAC is highly expressed in the human liver, where carboxylesterase (CES) enzymes, namely, CES1 and CES2, are also expressed. It is generally recognized that CES1 prefers compounds with a large acyl moiety and a small alcohol or amine moiety as substrates, whereas CES2 prefers compounds with a small acyl moiety and a large alcohol or amine moiety. In a comparison of the chemical structures of known AADAC substrates, AADAC most likely prefers compounds with the same characteristics as does CES2. However, the substrate specificity of human AADAC has not been fully clarified. To expand the knowledge of substrates of human AADAC, we measured its hydrolase activities toward 13 compounds, including known human CES1 and CES2 substrates, using recombinant enzymes expressed in Sf21 cells. Recombinant AADAC catalyzed the hydrolysis of fluorescein diacetate, N-monoacetyldapsone, and propanil, which possess notably small acyl moieties, and these substrates were also hydrolyzed by CES2. However, AADAC could not hydrolyze another CES2 substrate, procaine, which possesses a moderately small acyl moiety. In addition, AADAC did not hydrolyze several known CES1 substrates, including clopidogrel and oseltamivir, which have large acyl moieties and small alcohol moieties. Collectively, these results suggest that AADAC prefers compounds with smaller acyl moieties than does CES2. The role of AADAC in the hydrolysis of drugs has been clarified. For this reason, AADAC should receive attention in ADMET studies during drug development.
ESTHER : Fukami_2015_Eur.J.Pharm.Sci_78_47
PubMedSearch : Fukami_2015_Eur.J.Pharm.Sci_78_47
PubMedID: 26164127
Gene_locus related to this paper: human-AADAC , human-CES1 , human-CES2

Title : Characterization of Species Differences in Tissue Diltiazem Deacetylation Identifies Ces2a as a Rat-Specific Diltiazem Deacetylase - Kurokawa_2015_Drug.Metab.Dispos_43_1218
Author(s) : Kurokawa T , Fukami T , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 43 :1218 , 2015
Abstract : Diltiazem, a calcium channel blocker, is mainly metabolized via demethylation or deacetylation in humans. Diltiazem demethylation is catalyzed by cytochrome P450 2D6 and 3A4. Although it was previously reported that the area under the curve ratio of deacetyldiltiazem to diltiazem after oral dosing with diltiazem in rats was sevenfold higher than in humans, the molecular mechanisms underlying this species difference remain to be clarified. In the present study, we compared the diltiazem deacetylase activity in liver, intestinal, renal, and pulmonary microsome preparations of human and experimental animal tissues to identify the specific deacetylase enzyme(s) involved in deacetylation. Diltiazem deacetylase activity was detected in rat liver and small intestine microsome preparations, but not in those from human, monkey, dog, and mouse tissues. Further purification of rat liver microsome (RLM) proteins identified four carboxylesterase (Ces) enzymes (Ces1d, Ces1e, Ces1f, and Ces2a) as potential candidate deacetylases. On the basis of their tissue distribution, the Ces2a enzyme was considered to be the enzyme that was responsible for diltiazem deacetylation. Furthermore, recombinant rat Ces2a expressed in Sf21 cells displayed efficient diltiazem deacetylase activity with similar Km values as RLM. In addition, the inhibitory characteristics of various chemical inhibitors were similar between recombinant rat Ces2a and RLM. In conclusion, we determined that only rat tissues were able to catalyze diltiazem deacetylation. The characterization of Ces enzymes in animal species, as undertaken in this study, will prove useful to predict the species-specific pharmacokinetics differences between the in vivo models used for drug development.
ESTHER : Kurokawa_2015_Drug.Metab.Dispos_43_1218
PubMedSearch : Kurokawa_2015_Drug.Metab.Dispos_43_1218
PubMedID: 25979260

Title : Comparison of genetic structures and biochemical properties of tandem cutinase-type polyesterases from Thermobifida alba AHK119 - Thumarat_2015_J.Biosci.Bioeng_120_491
Author(s) : Thumarat U , Kawabata T , Nakajima M , Nakajima H , Sugiyama A , Yazaki K , Tada T , Waku T , Tanaka N , Kawai F
Ref : J Biosci Bioeng , 120 :491 , 2015
Abstract : This study described the genetic map of tandem genes (est1 and est119) encoding cutinase-type polyesterases in Thermobifida alba AHK119 and comparison of wild type and mutant enzymes of Est1 and Est119. Two genes were independently and constitutively expressed. The activity of Est1 was higher by approximately 1.6-1.7-fold than that of Est119 towards p-nitrophenyl butyrate, although both enzymes shared 95% sequence identity and 98% similarity and possessed similar 3D structures except that several amino acids in the probable substrate-docking loops were different from each other. Calcium ion enhanced the activity and the thermostability of both enzymes. Based on conserved sequences among Thermobifida cutinases, valine, proline and lysine were introduced into Est1 at Ala68, Thr253 and Met256, respectively. Among wild and mutant enzymes of Est119 and Est1, Est1 (A68V/T253P) possessed three prolines in the substrate-docking loops and displayed the highest thermostability that spotlighted the important effect of proline numbers in the loops. Est1 (A68V/T253P) was stable for 1 h below 60 degrees C and even at 65 degrees C, more than 70% and 50% activities were maintained after 30 and 60 min, respectively. Est1 (A68V/T253P) degraded various aliphatic and aliphatic-co-aromatic polyesters and hydrophilized an amorphous PET film. The enzyme hydrolyzed a PET trimer model compound, indicating its specificity towards an ester bond between terephthalic acid and ethylene glycol.
ESTHER : Thumarat_2015_J.Biosci.Bioeng_120_491
PubMedSearch : Thumarat_2015_J.Biosci.Bioeng_120_491
PubMedID: 25910960
Gene_locus related to this paper: 9acto-d4q9n1 , 9acto-f7ix06

Title : Development of Inhibitors of Salicylic Acid Signaling - Jiang_2015_J.Agric.Food.Chem_63_7124
Author(s) : Jiang K , Kurimoto T , Seo EK , Miyazaki S , Nakajima M , Nakamura H , Asami T
Ref : Journal of Agricultural and Food Chemistry , 63 :7124 , 2015
Abstract : Salicylic acid (SA) plays important roles in the induction of systemic acquired resistance (SAR) in plants. Determining the mechanism of SAR will extend our understanding of plant defenses against pathogens. We recently reported that PAMD is an inhibitor of SA signaling, which suppresses the expression of the pathogenesis-related PR genes and is expected to facilitate the understanding of SA signaling. However, PAMD strongly inhibits plant growth. To minimize the side effects of PAMD, we synthesized a number of PAMD derivatives, and identified compound 4 that strongly suppresses the expression of the PR genes with fewer adverse effects on plant growth than PAMD. We further showed that the adverse effects on plant growth were partially caused the stabilization of DELLA, which is also related to the pathogen responses. These results indicate that compound 4 would facilitate our understanding of SA signaling and its cross talk with other plant hormones.
ESTHER : Jiang_2015_J.Agric.Food.Chem_63_7124
PubMedSearch : Jiang_2015_J.Agric.Food.Chem_63_7124
PubMedID: 26236918

Title : A comprehensive review of UDP-glucuronosyltransferase and esterases for drug development - Oda_2015_Drug.Metab.Pharmacokinet_30_30
Author(s) : Oda S , Fukami T , Yokoi T , Nakajima M
Ref : Drug Metab Pharmacokinet , 30 :30 , 2015
Abstract : UDP-glucuronosyltransferase (UGT) and esterases are recognized as the most important non-P450 enzymes because of their high contribution to drug metabolism. UGTs catalyze the transfer of glucuronic acid to hydroxyl, carboxyl, or amine groups of compounds, whereas esterases hydrolyze compounds that contain ester, amide, and thioester bonds. These enzymes, in most cases, convert hydrophobic compounds to water-soluble metabolites to facilitate the elimination of compounds from the body. Information about these enzymes is steadily increasing, although our knowledge is still behind our understanding of P450. This review gives an overview of recent findings in UGT and esterases studies focusing on tissue distribution, gene regulation, substrate and inhibitor specificity, and species differences. In particular, the absolute protein content of UGT isoforms and esterases in human tissues could be available. In the field of esterases, it is becoming clear that enzymes other than carboxylesterase are involved in drug hydrolysis. In addition, there is an interesting interplay between UGTs and esterases in the formation and hydrolytic deglucuronidation of acyl-glucuronide, which is considered to be a reactive metabolite. With the growing awareness of the importance of non-P450 enzymes in drug development, issues that should be resolved are discussed.
ESTHER : Oda_2015_Drug.Metab.Pharmacokinet_30_30
PubMedSearch : Oda_2015_Drug.Metab.Pharmacokinet_30_30
PubMedID: 25760529

Title : P-glycoprotein, CYP3A, and Plasma Carboxylesterase Determine Brain Disposition and Oral Availability of the Novel Taxane Cabazitaxel (Jevtana) in Mice - Tang_2015_Mol.Pharm_12_3714
Author(s) : Tang SC , Kort A , Cheung KL , Rosing H , Fukami T , Durmus S , Wagenaar E , Hendrikx JJ , Nakajima M , van Vlijmen BJ , Beijnen JH , Schinkel AH
Ref : Mol Pharm , 12 :3714 , 2015
Abstract : We aimed to clarify the roles of the multidrug-detoxifying proteins ABCB1, ABCG2, ABCC2, and CYP3A in oral availability and brain accumulation of cabazitaxel, a taxane developed for improved therapy of docetaxel-resistant prostate cancer. Cabazitaxel pharmacokinetics were studied in Abcb1a/1b, Abcg2, Abcc2, Cyp3a, and combination knockout mice. We found that human ABCB1, but not ABCG2, transported cabazitaxel in vitro. Upon oral cabazitaxel administration, total plasma levels were greatly increased due to binding to plasma carboxylesterase Ces1c, which is highly upregulated in several knockout strains. Ces1c inhibition and in vivo hepatic Ces1c knockdown reversed these effects. Correcting for Ces1c effects, Abcb1a/1b, Abcg2, and Abcc2 did not restrict cabazitaxel oral availability, whereas Abcb1a/1b, but not Abcg2, dramatically reduced cabazitaxel brain accumulation (>10-fold). Coadministration of the ABCB1 inhibitor elacridar completely reversed this brain accumulation effect. After correction for Ces1c effects, Cyp3a knockout mice demonstrated a strong (six-fold) increase in cabazitaxel oral availability, which was completely reversed by transgenic human CYP3A4 in intestine and liver. Cabazitaxel markedly inhibited mouse Ces1c, but human CES1 and CES2 only weakly. Ces1c upregulation can thus complicate preclinical cabazitaxel studies. In summary, ABCB1 limits cabazitaxel brain accumulation and therefore potentially therapeutic efficacy against (micro)metastases or primary tumors positioned wholly or partly behind a functional blood-brain barrier. This can be reversed with elacridar coadministration, and similar effects may apply to ABCB1-expressing tumors. CYP3A4 profoundly reduces the oral availability of cabazitaxel. This may potentially be greatly improved by coadministering ritonavir or other CYP3A inhibitors, suggesting the option of patient-friendly oral cabazitaxel therapy.
ESTHER : Tang_2015_Mol.Pharm_12_3714
PubMedSearch : Tang_2015_Mol.Pharm_12_3714
PubMedID: 26317243

Title : A proposed mechanism for the adverse effects of acebutolol: CES2 and CYP2C19-mediated metabolism and antinuclear antibody production - Muta_2015_Biochem.Pharmacol_98_659
Author(s) : Muta K , Fukami T , Nakajima M
Ref : Biochemical Pharmacology , 98 :659 , 2015
Abstract : Acebutolol, a beta-adrenergic receptor-blocker, occasionally causes drug-induced lupus erythematosus (DILE). Acebutolol is mainly metabolized to diacetolol. Because metabolic activation has been considered to be related to acebutolol-induced toxicity, we sought to identify the enzymes that are responsible for acebutolol metabolism and investigate their involvement in acebutolol-induced toxicity. By using human liver microsomes (HLM) or intestinal microsomes and recombinant enzymes, we found that diacetolol was produced via hydrolysis by carboxylesterase 2 (CES2) and subsequent acetylation by N-acetyltransferase 2 (NAT2). When acetolol, a hydrolytic metabolite of acebutolol, was incubated with HLM and an NADPH-generating system, a metabolite conjugated with N-acetylcystein was generated. This metabolite was found to be formed by CYP2C19 based on studies with a panel of recombinant cytochrome P450 enzymes and an inhibition study using HLM with tranylcypromine, a CYP2C19 inhibitor. Because antinuclear antibody (ANA) production is associated with DILE, we investigated whether ANA was detected in plasma from mice treated with acebutolol. Administration of acebutolol (100mg/kg, p.o.) to female C57BL/6 mice for 30 days resulted in ANA production in plasma in seven of thirteen mice. The number of mice that showed ANA production was larger in mice co-treated with pregnenolone 16alpha-carbonitrile, an inducer of P450s, whereas it was lower in mice co-treated with tri-o-tolylphosphate or 1-aminobenzotriazole, which are inhibitors of esterases or P450s, respectively. These results suggested that the hydrolysis and oxidation of acebutolol was associated with ANA production. In summary, this study demonstrated that metabolic activation may be a causal factor of adverse reactions of acebutolol.
ESTHER : Muta_2015_Biochem.Pharmacol_98_659
PubMedSearch : Muta_2015_Biochem.Pharmacol_98_659
PubMedID: 26408002

Title : Indiplon is hydrolyzed by arylacetamide deacetylase in human liver - Shimizu_2014_Drug.Metab.Dispos_42_751
Author(s) : Shimizu M , Fukami T , Ito Y , Kurokawa T , Kariya M , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 42 :751 , 2014
Abstract : Human arylacetamide deacetylase (AADAC) catalyzes the hydrolysis of some clinically used drugs, but the information available on its substrates is limited. To increase our knowledge of the AADAC substrates, we examined whether AADAC catalyzes the hydrolysis of indiplon, which was initially developed as a hypnotic sedative drug. It has been reported that approximately 30-40% of the administered indiplon was hydrolyzed to deacetylindiplon in humans, but the enzyme responsible for this hydrolysis had not been identified. We detected high levels of indiplon hydrolase activity in human liver microsomes (HLMs), but the levels found in human liver cytosol and plasma were scarcely detectable. Recombinant AADAC showed a high level of indiplon hydrolase activity, whereas recombinant carboxylesterase 1 (CES1) and 2 (CES2) showed marginal activity. The indiplon hydrolase activity of HLM was potently inhibited by vinblastine, a potent inhibitor of AADAC and CES2, but it was not inhibited by digitonin and telmisartan, inhibitors of CES1 and CES2, respectively. In a panel of 24 individual HLM samples, the indiplon hydrolase activities were significantly correlated with the hydrolase activities of flutamide, phenacetin, and rifampicin, which are known AADAC substrates. An HLM sample with a homozygous AADAC*3 allele, which was previously found to exhibit decreased enzyme activity, showed the lowest indiplon hydrolase activity among the 24 tested samples. Collectively, we found that human AADAC is responsible for the hydrolysis of indiplon. Thus, we can add indiplon to the list of human AADAC substrates.
ESTHER : Shimizu_2014_Drug.Metab.Dispos_42_751
PubMedSearch : Shimizu_2014_Drug.Metab.Dispos_42_751
PubMedID: 24464802

Title : An Orphan Esterase ABHD10 Modulates Probenecid Acyl Glucuronidation in Human Liver - Ito_2014_Drug.Metab.Dispos_42_2109
Author(s) : Ito Y , Fukami T , Yokoi T , Nakajima M
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 42 :2109 , 2014
Abstract : Probenecid, a widely used uricosuric agent, is mainly metabolized to probenecid acyl glucuronide (PRAG), which is considered a causal substance of severe allergic or anaphylactoid reactions. PRAG can be hydrolyzed (deglucuronidated) to probenecid. The purpose of this study was to identify enzymes responsible for probenecid acyl glucuronidation and PRAG deglucuronidation in human livers and to examine the effect of deglucuronidation in PRAG formation. In human liver homogenates (HLHs), the intrinsic clearance (CLint) of PRAG deglucuronidation was much greater (497-fold) than that of probenecid acyl glucuronidation. Evaluation of PRAG formation by recombinant UDP-glucuronosyltransferase (UGT) isoforms and an inhibition study using HLHs as an enzyme source demonstrated that multiple UGT isoforms, including UGT1A1, UGT1A9, and UGT2B7, catalyzed probenecid acyl glucuronidation. We found that recombinant alpha/beta hydrolase domain containing 10 (ABHD10) substantially catalyzed PRAG deglucuronidation activity, whereas carboxylesterases did not. Similar inhibitory patterns by chemicals between HLHs and recombinant ABHD10 supported the major contribution of ABHD10 to PRAG deglucuronidation in human liver. Interestingly, it was demonstrated that the CLint value of probenecid acyl glucuronidation in HLHs was increased by 1.7-fold in the presence of phenylmethylsulfonyl fluoride, which potently inhibited ABHD10 activity. In conclusion, we found that PRAG deglucuronidation catalyzed by ABHD10 suppressively regulates PRAG formation via multiple UGT enzymes in human liver. The balance of activities by these enzymes is important for the formation of PRAG, which may be associated with the adverse reactions observed after probenecid administration.
ESTHER : Ito_2014_Drug.Metab.Dispos_42_2109
PubMedSearch : Ito_2014_Drug.Metab.Dispos_42_2109
PubMedID: 25217485
Gene_locus related to this paper: human-ABHD10 , mouse-abhda

Title : Screening of specific inhibitors for human carboxylesterases or arylacetamide deacetylase - Shimizu_2014_Drug.Metab.Dispos_42_1103
Author(s) : Shimizu M , Fukami T , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 42 :1103 , 2014
Abstract : Esterases catalyze the hydrolysis of therapeutic drugs containing esters or amides in their structures. Human carboxylesterase (CES) and arylacetamide deacetylase (AADAC) are the major enzymes that catalyze the hydrolysis of drugs in the liver. Characterization of the enzyme(s) responsible for drug metabolism is required in drug development and to realize optimal drug therapy. Because multiple enzymes may show a metabolic potency for a given compound, inhibition studies using chemical inhibitors are useful tools to determine the contribution of each enzyme in human tissue preparations. The purpose of this study was to find specific inhibitors for human CES1, CES2, and AADAC. We screened 542 chemicals for the inhibition potency toward hydrolase activities of p-nitrophenyl acetate by recombinant CES1, CES2, and AADAC. We found that digitonin and telmisartan specifically inhibited CES1 and CES2 enzyme activity, respectively. Vinblastine potently inhibited both AADAC and CES2, but no specific inhibitor of AADAC was found. The inhibitory potency and specificity of these compounds were also evaluated by monitoring the effects on hydrolase activity of probe compounds of each enzyme (CES1: lidocaine, CES2: CPT-11, AADAC: phenacetin) in human liver microsomes. Telmisartan and vinblastine strongly inhibited the hydrolysis of CPT-11 and/or phenacetin, but digitonin did not strongly inhibit the hydrolysis of lidocaine, indicating that the inhibitory potency of digitonin was different between recombinant CES1 and liver microsomes. Although we could not find a specific inhibitor of AADAC, the combined use of telmisartan and vinblastine could predict the responsibility of human AADAC to drug hydrolysis.
ESTHER : Shimizu_2014_Drug.Metab.Dispos_42_1103
PubMedSearch : Shimizu_2014_Drug.Metab.Dispos_42_1103
PubMedID: 24751575
Gene_locus related to this paper: human-AADAC , human-CES1 , human-CES2

Title : N-Glycosylation during translation is essential for human arylacetamide deacetylase enzyme activity - Muta_2014_Biochem.Pharmacol_87_352
Author(s) : Muta K , Fukami T , Nakajima M , Yokoi T
Ref : Biochemical Pharmacology , 87 :352 , 2014
Abstract : Human arylacetamide deacetylase (AADAC) can hydrolyze clinical drugs such as flutamide, phenacetin, and rifamycins. AADAC is a glycoprotein, but the role of glycosylation remains unclear. In the present study, we investigated the effect of glycosylation on AADAC enzyme activity. Immunoblot analysis of mutant AADACs that contained an asparagine (N, Asn) to glutamine (Q, Gln) substitution at either residue 78 or 282 (N78Q or N282Q) showed a different migration compared with the wild-type protein. A mutant AADAC that contained N to Q substitutions at both residue 78 and 282 (N78Q/N282Q) showed a similar migration to AADAC in human liver microsomes (HLM) treated with endoglycosidase H (Endo H), which produces deglycosylated proteins. This result indicated that AADAC was glycosylated at both N78 and N282. Mutant types of AADAC with the N282Q and the N78Q/N282Q substitutions showed dramatically lower phenacetin hydrolase activity than did the wild-type protein. The treatment of wild-type AADAC-expressing HuH-7 cells with tunicamycin, which produces unglycosylated protein, decreased AADAC enzyme activity. However, the treatment of the HLM with Endo H caused no decrease of AADAC activity. Thus, the oligosaccharide chain, per se, was not important for AADAC activity in the mature form. The mutant types of AADAC containing the N282Q and the N78Q/N282Q substitutions were not detected by immunoblotting analysis after non-reducing SDS-PAGE, suggesting that the glycosylation of AADAC at N282 was important for proper protein folding. Overall, this study found that the translational, but not post-translational, N-glycosylation of AADAC plays a crucial role in regulating AADAC enzyme activity.
ESTHER : Muta_2014_Biochem.Pharmacol_87_352
PubMedSearch : Muta_2014_Biochem.Pharmacol_87_352
PubMedID: 24125761
Gene_locus related to this paper: human-AADAC

Title : Prilocaine- and lidocaine-induced methemoglobinemia is caused by human carboxylesterase-, CYP2E1-, and CYP3A4-mediated metabolic activation - Higuchi_2013_Drug.Metab.Dispos_41_1220
Author(s) : Higuchi R , Fukami T , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 41 :1220 , 2013
Abstract : Prilocaine and lidocaine are classified as amide-type local anesthetics for which serious adverse effects include methemoglobinemia. Although the hydrolyzed metabolites of prilocaine (o-toluidine) and lidocaine (2,6-xylidine) have been suspected to induce methemoglobinemia, the metabolic enzymes that are involved remain uncharacterized. In the present study, we aimed to identify the human enzymes that are responsible for prilocaine- and lidocaine-induced methemoglobinemia. Our experiments revealed that prilocaine was hydrolyzed by recombinant human carboxylesterase (CES) 1A and CES2, whereas lidocaine was hydrolyzed by only human CES1A. When the parent compounds (prilocaine and lidocaine) were incubated with human liver microsomes (HLM), methemoglobin (Met-Hb) formation was lower than when the hydrolyzed metabolites were incubated with HLM. In addition, Met-Hb formation when prilocaine and o-toluidine were incubated with HLM was higher than that when lidocaine and 2,6-xylidine were incubated with HLM. Incubation with diisopropyl fluorophosphate and bis-(4-nitrophenyl) phosphate, which are general inhibitors of CES, significantly decreased Met-Hb formation when prilocaine and lidocaine were incubated with HLM. An anti-CYP3A4 antibody further decreased the residual formation of Met-Hb. Met-Hb formation after the incubation of o-toluidine and 2,6-xylidine with HLM was only markedly decreased by incubation with an anti-CYP2E1 antibody. o-Toluidine and 2,6-xylidine were further metabolized by CYP2E1 to 4- and 6-hydroxy-o-toluidine and 4-hydroxy-2,6-xylidine, respectively, and these metabolites were shown to more efficiently induce Met-Hb formation than the parent compounds. Collectively, we found that the metabolites produced by human CES-, CYP2E1-, and CYP3A4-mediated metabolism were involved in prilocaine- and lidocaine-induced methemoglobinemia.
ESTHER : Higuchi_2013_Drug.Metab.Dispos_41_1220
PubMedSearch : Higuchi_2013_Drug.Metab.Dispos_41_1220
PubMedID: 23530020

Title : Chemical screening of an inhibitor for gibberellin receptors based on a yeast two-hybrid system - Yoon_2013_Bioorg.Med.Chem.Lett_23_1096
Author(s) : Yoon JM , Nakajima M , Mashiguchi K , Park SH , Otani M , Asami T
Ref : Bioorganic & Medicinal Chemistry Lett , 23 :1096 , 2013
Abstract : We applied a yeast two-hybrid (Y2H) system to the high-throughput monitoring of two proteins' interaction, a receptor for phytohormone gibberellin (GA) and its direct signal transducer DELLA. With this system, we screened inhibitors to the interaction. As a result, we discovered a chemical, 3-(2-thienylsulfonyl)pyrazine-2-carbonitrile (TSPC), and we confirmed that TSPC is an inhibitor for GA perception by in vitro and in planta evaluations.
ESTHER : Yoon_2013_Bioorg.Med.Chem.Lett_23_1096
PubMedSearch : Yoon_2013_Bioorg.Med.Chem.Lett_23_1096
PubMedID: 23298808

Title : Contributions of arylacetamide deacetylase and carboxylesterase 2 to flutamide hydrolysis in human liver - Kobayashi_2012_Drug.Metab.Dispos_40_1080
Author(s) : Kobayashi Y , Fukami T , Shimizu M , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 40 :1080 , 2012
Abstract : Flutamide, an antiandrogen drug, is widely used for the treatment of prostate cancer. The major metabolic pathways of flutamide are hydroxylation and hydrolysis. The hydrolyzed metabolite, 5-amino-2-nitrobenzotrifluoride (FLU-1), is further metabolized to N-hydroxy FLU-1, an assumed hepatotoxicant. Our previous study demonstrated that arylacetamide deacetylase (AADAC), one of the major serine esterases expressed in the human liver and gastrointestinal tract, catalyzes the flutamide hydrolysis. However, the enzyme kinetics in human tissue microsomes were not consistent with the kinetics by recombinant human AADAC. Thus, it seemed that AADAC is not the sole enzyme responsible for flutamide hydrolysis in human. In the present study, we found that recombinant carboxylesterase (CES) 2 could hydrolyze flutamide at low concentrations of flutamide. In the inhibition assay, the flutamide hydrolase activities at a flutamide concentration of 5 muM in human liver and jejunum microsomes were strongly inhibited by a selective CES2 inhibitor, 10 muM loperamide, with the residual activities of 22.9 +/- 3.5 and 18.6 +/- 0.7%, respectively. These results suggest that CES2 is also involved in the flutamide hydrolysis in human tissues. Using six individual human livers, the contributions of AADAC and CES2 to flutamide hydrolysis were estimated by using the relative activity factor. The relative contribution of CES2 was approximately 75 to 99% at the concentration of 5 muM flutamide. In contrast, the relative contribution of AADAC increased in parallel with the concentration of flutamide. Thus, CES2, rather than AADAC, largely contributed to the flutamide hydrolysis in clinical therapeutics.
ESTHER : Kobayashi_2012_Drug.Metab.Dispos_40_1080
PubMedSearch : Kobayashi_2012_Drug.Metab.Dispos_40_1080
PubMedID: 22446520

Title : An unusual spliced variant of DELLA protein, a negative regulator of gibberellin signaling, in lettuce - Sawada_2012_Biosci.Biotechnol.Biochem_76_544
Author(s) : Sawada Y , Umetsu A , Komatsu Y , Kitamura J , Suzuki H , Asami T , Fukuda M , Honda I , Mitsuhashi W , Nakajima M , Toyomasu T
Ref : Biosci Biotechnol Biochem , 76 :544 , 2012
Abstract : DELLA proteins are negative regulators of the signaling of gibberellin (GA), a phytohormone regulating plant growth. DELLA degradation is triggered by its interaction with GID1, a soluble GA receptor, in the presence of bioactive GA. We isolated cDNA from a spliced variant of LsDELLA1 mRNA in lettuce, and named it LsDELLA1sv. It was deduced that LsDELLA1sv encodes truncated LsDELLA1, which has DELLA and VHYNP motifs at the N terminus but lacks part of the C-terminal GRAS domain. The recombinant LsDELLA1sv protein interacted with both Arabidopsis GID1 and lettuce GID1s in the presence of GA. A yeast two-hybrid assay suggested that LsDELLA1sv interacted with LsDELLA1. The ratio of LsDELLA1sv to LsDELLA1 transcripts was higher in flower samples at the late reproductive stage and seed samples (dry seeds and imbibed seeds) than in the other organ samples examined. This study suggests that LsDELLA1sv is a possible modulator of GA signaling in lettuce.
ESTHER : Sawada_2012_Biosci.Biotechnol.Biochem_76_544
PubMedSearch : Sawada_2012_Biosci.Biotechnol.Biochem_76_544
PubMedID: 22451398

Title : Human alpha\/beta hydrolase domain containing 10 (ABHD10) is responsible enzyme for deglucuronidation of mycophenolic acid acyl-glucuronide in liver - Iwamura_2012_J.Biol.Chem_287_9240
Author(s) : Iwamura A , Fukami T , Higuchi R , Nakajima M , Yokoi T
Ref : Journal of Biological Chemistry , 287 :9240 , 2012
Abstract : Mycophenolic acid (MPA), the active metabolite of the immunosuppressant mycophenolate mofetil (MMF), is primarily metabolized by glucuronidation to a phenolic glucuronide (MPAG) and an acyl glucuronide (AcMPAG). It is known that AcMPAG, which may be an immunotoxic metabolite, is deglucuronidated in human liver. However, it has been reported that recombinant beta-glucuronidase does not catalyze this reaction. AcMPAG deglucuronidation activity was detected in both human liver cytosol (HLC) and microsomes (HLM). In this study, the enzyme responsible for AcMPAG deglucuronidation was identified by purification from HLC with column chromatographic purification steps. The purified enzyme was identified as alpha/beta hydrolase domain containing 10 (ABHD10) by amino acid sequence analysis. Recombinant ABHD10 expressed in Sf9 cells efficiently deglucuronidated AcMPAG with a K(m) value of 100.7 +/- 10.2 muM, which was similar to those in HLM, HLC, and human liver homogenates (HLH). Immunoblot analysis revealed ABHD10 protein expression in both HLC and HLM. The AcMPAG deglucuronidation by recombinant ABHD10, HLC, and HLH were potently inhibited by AgNO(3), CdCl(2), CuCl(2), PMSF, bis-p-nitrophenylphosphate, and DTNB. The CL(int) value of AcMPAG formation from MPA, which was catalyzed by human UGT2B7, in HLH was increased by 1.8-fold in the presence of PMSF. Thus, human ABHD10 would affect the formation of AcMPAG, the immunotoxic metabolite.
ESTHER : Iwamura_2012_J.Biol.Chem_287_9240
PubMedSearch : Iwamura_2012_J.Biol.Chem_287_9240
PubMedID: 22294686
Gene_locus related to this paper: human-ABHD10 , mouse-abhda

Title : Species differences in tissue distribution and enzyme activities of arylacetamide deacetylase in human, rat, and mouse - Kobayashi_2012_Drug.Metab.Dispos_40_671
Author(s) : Kobayashi Y , Fukami T , Nakajima A , Watanabe A , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 40 :671 , 2012
Abstract : Human arylacetamide deacetylase (AADAC) is a major esterase responsible for the hydrolysis of clinical drugs such as flutamide, phenacetin, and rifampicin. Thus, AADAC is considered to be a relevant enzyme in preclinical drug development, but there is little information about species differences with AADAC. This study investigated the species differences in the tissue distribution and enzyme activities of AADAC. In human, AADAC mRNA was highly expressed in liver and the gastrointestinal tract, followed by bladder. In rat and mouse, AADAC mRNA was expressed in liver at the highest level, followed by the gastrointestinal tract and kidney. The expression levels in rat tissues were approximately 7- and 10-fold lower than those in human and mouse tissues, respectively. To compare the catalytic efficiency of AADAC among three species, each recombinant AADAC was constructed, and enzyme activities were evaluated by normalizing with the expression levels of AADAC. Flutamide and phenacetin hydrolase activities were detected by the recombinant AADAC of all species. In flutamide hydrolysis, liver microsomes of all species showed similar catalytic efficiencies, despite the lower AADAC mRNA expression in rat liver. In phenacetin hydrolysis, rat liver microsomes showed approximately 4- to 6.5-fold lower activity than human and mouse liver microsomes. High rifampicin hydrolase activity was detected only by recombinant human AADAC and human liver and jejunum microsomes. Taken together, the results of this study clarified the species differences in the tissue distribution and enzyme activities of AADAC and facilitate our understanding of species differences in drug hydrolysis.
ESTHER : Kobayashi_2012_Drug.Metab.Dispos_40_671
PubMedSearch : Kobayashi_2012_Drug.Metab.Dispos_40_671
PubMedID: 22207054
Gene_locus related to this paper: human-AADAC , mouse-aryla , ratno-aryla

Title : A novel polymorphic allele of human arylacetamide deacetylase leads to decreased enzyme activity - Shimizu_2012_Drug.Metab.Dispos_40_1183
Author(s) : Shimizu M , Fukami T , Kobayashi Y , Takamiya M , Aoki Y , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 40 :1183 , 2012
Abstract : Human arylacetamide deacetylase (AADAC) is responsible for the hydrolysis of clinically used drugs such as flutamide, phenacetin, and rifamycins. Our recent studies suggested that human AADAC is a relevant enzyme pharmacologically and toxicologically. To date, the genetic polymorphisms that affect enzyme activity in AADAC have been unknown. In this study, we found single-nucleotide polymorphisms in the human AADAC gene in a liver sample that showed remarkably low flutamide hydrolase activity. Among them, g.13651G > A (V281I) and g.14008T > C (X400Q) were nonsynonymous. The latter would be predicted to cause a C-terminal one-amino acid (glutamine) extension. The AADAC*2 allele (g.13651G > A) was found in all populations investigated in this study (European American, African American, Korean, and Japanese), at allelic frequencies of 52.6 to 63.5%, whereas the AADAC*3 allele (g.13651G > A/g.14008T > C) was found in European American (1.3%) and African American (2.0%) samples. COS7 cells expressing AADAC.1 (wild-type) exhibited flutamide, phenacetin, and rifampicin hydrolase activities with intrinsic clearance (CLint) values of 1.31 +/- 0.06, 1.00 +/- 0.02, and 0.39 +/- 0.02 mul x min(-1) x unit(-1), respectively. AADAC.2, which is a protein produced from the AADAC*2 allele, showed moderately lower or similar CLint values, compared with AADAC.1, but AADAC.3 showed substantially lower CLint values (flutamide hydrolase, 0.21 +/- 0.02 mul x min(-1) x unit(-1); phenacetin hydrolase, 0.12 +/- 0.00 mul x min(-1) x unit(-1); rifampicin hydrolase, 0.03 +/- 0.01 mul x min(-1) x unit(-1), respectively). Microsomes from a liver sample genotyped as AADAC*3/AADAC*3 showed decreased enzyme activities, compared with those genotyped as AADAC*1/AADAC*1, AADAC*1/AADAC*2, and AADAC*2/AADAC*2. In conclusion, we found an AADAC allele that yielded decreased enzyme activity. This study should provide useful information on interindividual variations in AADAC enzyme activity.
ESTHER : Shimizu_2012_Drug.Metab.Dispos_40_1183
PubMedSearch : Shimizu_2012_Drug.Metab.Dispos_40_1183
PubMedID: 22415931
Gene_locus related to this paper: human-AADAC

Title : Human arylacetamide deacetylase is responsible for deacetylation of rifamycins: rifampicin, rifabutin, and rifapentine - Nakajima_2011_Biochem.Pharmacol_82_1747
Author(s) : Nakajima A , Fukami T , Kobayashi Y , Watanabe A , Nakajima M , Yokoi T
Ref : Biochemical Pharmacology , 82 :1747 , 2011
Abstract : Rifamycins such as rifampicin, rifabutin, and rifapentine are used for the treatment of tuberculosis and induce various drug-metabolizing enzymes. Rifamycins have been reported to be mainly deacetylated by esterase(s) expressed in human liver microsomes (HLM) to 25-deacetylrifamycins, but the responsible enzyme remained to be determined. In this study, we found that recombinant human arylacetamide deacetylase (AADAC) could efficiently deacetylate rifamycins, whereas human carboxylesterases, which are enzymes responsible for the hydrolysis of many prodrugs, showed no activity. The involvement of AADAC in the deacetylation of rifamycins in HLM was verified by the similarities of the K(m) and K(i) values and the inhibitory characteristics between recombinant AADAC and HLM. Rifamycins exhibited potent cytotoxicity to HepG2 cells, but their 25-deacetylated metabolites did not. Luciferase assay using a reporter plasmid containing CYP3A4 direct repeat 3 and everted repeat 6 motifs revealed that 25-deacetylrifamycins have lesser potency to transactivate CYP3A4 compared with the parent drugs. Supporting these results, HepG2 cells infected with a recombinant adenovirus expressing human AADAC showed low cytotoxicity and induction potency of CYP3A4 by rifamycins. In addition, CYP3A4 induction in human hepatocytes by rifamycins was increased by transfecting siRNA for human AADAC. Thus, we found that human AADAC was the enzyme responsible for the deacetylation of rifamycins and would affect the induction rate of drug-metabolizing enzymes by rifamycins and their induced hepatotoxicity.
ESTHER : Nakajima_2011_Biochem.Pharmacol_82_1747
PubMedSearch : Nakajima_2011_Biochem.Pharmacol_82_1747
PubMedID: 21856291
Gene_locus related to this paper: human-AADAC

Title : Arylacetamide deacetylase is a determinant enzyme for the difference in hydrolase activities of phenacetin and acetaminophen - Watanabe_2010_Drug.Metab.Dispos_38_1532
Author(s) : Watanabe A , Fukami T , Takahashi S , Kobayashi Y , Nakagawa N , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 38 :1532 , 2010
Abstract : Phenacetin was withdrawn from the market because it caused renal failure in some patients. Many reports indicated that the nephrotoxicity of phenacetin is associated with the hydrolyzed metabolite, p-phenetidine. Acetaminophen (APAP), the major metabolite of phenacetin, is also hydrolyzed to p-aminophenol, which is a nephrotoxicant. However, APAP is safely prescribed if used in normal therapeutic doses. This background prompted us to investigate the difference between phenacetin and APAP hydrolase activities in human liver. In this study, we found that phenacetin is efficiently hydrolyzed in human liver microsomes (HLM) [CL(int) 1.08 +/- 0.02 microl/(min . mg)], whereas APAP is hardly hydrolyzed [0.02 +/- 0.00 microl/(min . mg)]. To identify the esterase involved in their hydrolysis, the activities were measured using recombinant human carboxylesterase (CES) 1A1, CES2, and arylacetamide deacetylase (AADAC). Among these, AADAC showed a K(m) value (1.82 +/- 0.02 mM) similar to that of HLM (3.30 +/- 0.16 mM) and the highest activity [V(max) 6.03 +/- 0.14 nmol/(min . mg)]. In contrast, APAP was poorly hydrolyzed by the three esterases. The large contribution of AADAC to phenacetin hydrolysis was demonstrated by the prediction with a relative activity factor. In addition, the phenacetin hydrolase activity by AADAC was activated by flutamide (5-fold) as well as that in HLM (4-fold), and the activity in HLM was potently inhibited by eserine, a strong inhibitor of AADAC. In conclusion, we found that AADAC is the principal enzyme responsible for the phenacetin hydrolysis, and the difference of hydrolase activity between phenacetin and APAP is largely due to the substrate specificity of AADAC.
ESTHER : Watanabe_2010_Drug.Metab.Dispos_38_1532
PubMedSearch : Watanabe_2010_Drug.Metab.Dispos_38_1532
PubMedID: 20542992

Title : In vitro evaluation of inhibitory effects of antidiabetic and antihyperlipidemic drugs on human carboxylesterase activities - Fukami_2010_Drug.Metab.Dispos_38_2173
Author(s) : Fukami T , Takahashi S , Nakagawa N , Maruichi T , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 38 :2173 , 2010
Abstract : Human carboxylesterase (CES) 1A is responsible for the biotransformation of angiotensin-converting enzyme (ACE) inhibitors such as imidapril and temocapril. Because antidiabetic or antihyperlipidemic drugs are often coadministered with ACE inhibitors in clinical pharmacotherapy, the inhibitory effect of these drugs on CES1A1 enzyme activity was investigated. In addition, the inhibitory effect on CES2 enzyme activity was evaluated to compare it with that on CES1A1. The inhibitory effects were evaluated with 11 antidiabetic and 12 antihyperlipidemic drugs. The imidapril hydrolase activity by recombinant CES1A1 was substantially inhibited by lactone ring-containing statins such as simvastatin and lovastatin and thiazolidinediones such as troglitazone and rosiglitazone. The activity in human liver microsomes was also strongly inhibited by simvastatin and troglitazone (K(i) = 0.8 +/- 0.1 and 5.6 +/- 0.2 muM, respectively). However, statins containing no lactone ring such as pravastatin and fluvastatin did not show strong inhibition. 7-Ethyl-10-[4-(1-piperidono)-1-piperidono]carbonyloxycamptothecin hydrolase activity by recombinant human CES2 was substantially inhibited by fenofibrate (K(i) = 0.04 +/- 0.01 muM) as well as by simvastatin (0.67 +/- 0.09 muM). Other fibrates such as clinofibrate and bezafibrate did not show strong inhibition. Thus, the inhibitory effects of the thiazolidinediones and fenofibrate on CES1A1 and CES2 were different. Some statins such as simvastatin and lovastatin, thiazolidinediones, and fenofibrate might attenuate the drug efficacy of prodrugs biotransformed by CES1A and CES2.
ESTHER : Fukami_2010_Drug.Metab.Dispos_38_2173
PubMedSearch : Fukami_2010_Drug.Metab.Dispos_38_2173
PubMedID: 20810539

Title : Transcriptional regulation of human carboxylesterase 1A1 by nuclear factor-erythroid 2 related factor 2 (Nrf2) - Maruichi_2010_Biochem.Pharmacol_79_288
Author(s) : Maruichi T , Fukami T , Nakajima M , Yokoi T
Ref : Biochemical Pharmacology , 79 :288 , 2010
Abstract : Human carboxylesterase (CES) 1A, which is predominantly expressed in liver and lung, plays an important role in the hydrolysis of endogenous compounds and xenobiotics. CES1A is reported to be induced in human hepatocytes by butylated hydroxyanisole, ticlopidine and diclofenac, and the induction is assumed to be caused by oxidative stress. However, the molecular mechanism remains to be determined. In this study, we sought to investigate whether CES1A is regulated by nuclear factor-erythroid 2 related factor 2 (Nrf2), which is a transcriptional factor activated by oxidative stress, and clarify the molecular mechanism. Real-time reverse transcription-PCR assays revealed that CES1A1 mRNA was significantly induced by tert-butylhydroquinone (tBHQ) and sulforaphane (SFN), which are representative activators of Nrf2 in HepG2, Caco-2 and HeLa cells. The induction was completely suppressed with small interfering RNA for Nrf2. In HepG2 cells, the CES1A protein level and imidapril hydrolase activity, which is specifically catalyzed by CES1A, were also significantly induced by tBHQ and SFN. Luciferase assays revealed that the antioxidant response element (ARE) at -2025 in the CES1A1 gene was responsible for the transactivation by Nrf2. In addition, electrophoretic mobility shift assays and chromatin immunoprecipitation assays revealed that Nrf2 binds to the ARE in the CES1A1 gene. These findings clearly demonstrated that human CES1A1 is induced by Nrf2. This is the first study to demonstrate the molecular mechanism of the inducible regulation of human CES1A1.
ESTHER : Maruichi_2010_Biochem.Pharmacol_79_288
PubMedSearch : Maruichi_2010_Biochem.Pharmacol_79_288
PubMedID: 19715681
Gene_locus related to this paper: human-CES1

Title : Human arylacetamide deacetylase is a principal enzyme in flutamide hydrolysis - Watanabe_2009_Drug.Metab.Dispos_37_1513
Author(s) : Watanabe A , Fukami T , Nakajima M , Takamiya M , Aoki Y , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 37 :1513 , 2009
Abstract : Flutamide, an antiandrogen drug, is widely used for the treatment of prostate cancer. The initial metabolic pathways of flutamide are hydroxylation and hydrolysis. It was recently reported that the hydrolyzed product, 4-nitro-3-(trifluoromethyl)phenylamine (FLU-1), is further metabolized to N-hydroxy FLU-1, an assumed hepatotoxicant. However, the esterase responsible for the flutamide hydrolysis has not been characterized. In the present study, we found that human arylacetamide deacetylase (AADAC) efficiently hydrolyzed flutamide using recombinant AADAC expressed in COS7 cells. In contrast, carboxylesterase1 (CES1) and CES2, which are responsible for the hydrolysis of many drugs, could not hydrolyze flutamide. AADAC is specifically expressed in the endoplasmic reticulum. Flutamide hydrolase activity was highly detected in human liver microsomes (K(m), 794 +/- 83 microM; V(max), 1.1 +/- 0.0 nmol/min/mg protein), whereas the activity was extremely low in human liver cytosol. The flutamide hydrolase activity in human liver microsomes was strongly inhibited by bis-(p-nitrophenyl)phosphate [corrected], diisopropylphosphorofluoride, and physostigmine sulfate (eserine) but moderately inhibited by sodium fluoride, phenylmethylsulfonyl fluoride, and disulfiram. The same inhibition pattern was obtained with the recombinant AADAC. Moreover, human liver and jejunum microsomes showing AADAC expression could hydrolyze flutamide, but human pulmonary and renal microsomes, which do not express AADAC, showed slight activity. In human liver microsomal samples (n = 50), the flutamide hydrolase activities were significantly correlated with the expression levels of AADAC protein (r = 0.66, p < 0.001). In conclusion, these results clearly showed that flutamide is exclusively hydrolyzed by AADAC. AADAC would be an important enzyme responsible for flutamide-induced hepatotoxicity.
ESTHER : Watanabe_2009_Drug.Metab.Dispos_37_1513
PubMedSearch : Watanabe_2009_Drug.Metab.Dispos_37_1513
PubMedID: 19339378
Gene_locus related to this paper: human-AADAC

Title : Different inhibitory effects in rat and human carboxylesterases - Takahashi_2009_Drug.Metab.Dispos_37_956
Author(s) : Takahashi S , Katoh M , Saitoh T , Nakajima M , Yokoi T
Ref : Drug Metabolism & Disposition: The Biological Fate of Chemicals , 37 :956 , 2009
Abstract : In vitro inhibition studies on drug-metabolizing enzyme activity are useful for understanding drug-drug interactions and for drug development. However, the profile of the inhibitory effects of carboxylesterase (CES) activity has not been fully investigated concerning species and tissue differences. In the present study, we measured the inhibitory effects of 15 drugs and 1 compound on CES activity using liver and jejunum microsomes and cytosol in human and rat. In addition, the inhibition constant (K(i) values) and patterns were determined for the compounds exhibiting strong inhibition. Hydrolysis of imidapril and irinotecan hydrochloride (CPT-11) is catalyzed mainly by CES1 and CES2, respectively. In the inhibition study, imidaprilat formation from imidapril in human liver was strongly inhibited by nordihydroguaiaretic acid (NDGA) and procainamide. The inhibition profile and pattern were similar in human liver and rat liver. The compounds showing potent inhibition were similar between liver and jejunum. The K(i) value of NDGA (K(i) = 13.3 +/- 1.5 microM) in human liver microsomes was 30-fold higher than that in rat liver microsomes (K(i) = 0.4 +/- 0.0 microM). On the other hand, 7-ethyl-10-hydroxycamptothecin (SN-38) formation from CPT-11 was not inhibited except by carvedilol, manidipine, and physostigmine. The K(i) value of physostigmine (K(i) = 0.3 +/- 0.0 microM) in human jejunum cytosol was 10-fold lower than that in rat jejunum cytosol (K(i) = 3.1 +/- 0.4 microM) and was similar to that for manidipine. The present study clarified the species differences in CES inhibition. These results are useful for the development of prodrugs.
ESTHER : Takahashi_2009_Drug.Metab.Dispos_37_956
PubMedSearch : Takahashi_2009_Drug.Metab.Dispos_37_956
PubMedID: 19225040

Title : Differential expression and affinities of Arabidopsis gibberellin receptors can explain variation in phenotypes of multiple knock-out mutants - Suzuki_2009_Plant.J_60_48
Author(s) : Suzuki H , Park SH , Okubo K , Kitamura J , Ueguchi-Tanaka M , Iuchi S , Katoh E , Kobayashi M , Yamaguchi I , Matsuoka M , Asami T , Nakajima M
Ref : Plant J , 60 :48 , 2009
Abstract : In Arabidopsis, three receptors exist for the phytohormone gibberellin. Of the three, only a double loss-of-function mutant (atgid1a atgid1c) shows a dwarf phenotype, while other double and all single mutants show no abnormality in height. In this study we show that the expression of AtGID1b-GUS mRNA, driven by the AtGID1b promoter, is low in inflorescence stems, but may be 10% of AtGID1a-GUS mRNA, driven by the AtGID1a promoter. However, AtGID1b-GUS enzymatic activity does not exist in them. This factor strongly suggests that atgid1a atgid1c lacks sufficient AtGID1b protein for normal stem growth. In the stamens of pAtGID1c::AtGID1c-GUS transformants, we detected clear AtGID1c-GUS activity, while another atgid1a atgid1b, which has short stamens in its flowers, causes the adhesion of little pollen to stigmas thus leading to its low fertility. We then evaluated the affinity of the AtGID1-DELLA interaction by a competitive yeast three-hybrid system and also by QCM apparatus. AtGID1c showed a quite lower affinity to RGL2, the major DELLA protein in floral buds, than AtGID1a or AtGID1b. The low affinity of the AtGID1c-RGL2 interaction is likely to be responsible for the failure of AtGID1c to hold RGL2, which is required for normal stamen development. Taken together with expressional information of DELLA genes, we propose that in a double loss-of-function mutant of gibberellin receptors, the emergence of any phenotype(s) depends on the abundance of the remaining receptor and its preference to DELLA proteins existing at a target site.
ESTHER : Suzuki_2009_Plant.J_60_48
PubMedSearch : Suzuki_2009_Plant.J_60_48
PubMedID: 19500306
Gene_locus related to this paper: arath-AT5G27320 , arath-GID1B

Title : Allosteric kinetics of human carboxylesterase 1: species differences and interindividual variability - Takahashi_2008_J.Pharm.Sci_97_5434
Author(s) : Takahashi S , Katoh M , Saitoh T , Nakajima M , Yokoi T
Ref : J Pharm Sci , 97 :5434 , 2008
Abstract : Esterified drugs such as imidapril, derapril, and oxybutynin hydrolyzed by carboxylesterase 1 (CES1) are extensively used in clinical practice. The kinetics using the CES1 substrates have not fully clarified, especially concerning species and tissue differences. In the present study, we performed the kinetic analyses in humans and rats in order to clarify these differences. The imidaprilat formation from imidapril exhibited sigmoidal kinetics in human liver microsomes (HLM) and cytosol (HLC) but Michaelis-Menten kinetics in rat liver microsomes and cytosol. The 2-cyclohexyl-2-phenylglycolic acid (CPGA) formation from oxybutynin were not detected in enzyme sources from rats, although HLM showed high activity. The kinetics were clarified to be different among species, tissues, and preparations. In individual HLM and HLC, there was large interindividual variability in imidaprilat (31- and 24-fold) and CPGA formations (15- and 9-fold). Imidaprilat formations exhibited Michaelis-Menten kinetics in HLM and HLC with high activity but sigmoidal kinetics in those with low activity. CPGA formations showed sigmoidal kinetics in high activity HLM but Michaelis-Menten kinetics in HLM with low activity. We revealed that the kinetics were different between individuals. These results could be useful for understanding interindividual variability and for the development of oral prodrugs.
ESTHER : Takahashi_2008_J.Pharm.Sci_97_5434
PubMedSearch : Takahashi_2008_J.Pharm.Sci_97_5434
PubMedID: 18383336

Title : Structural basis for gibberellin recognition by its receptor GID1 - Shimada_2008_Nature_456_520
Author(s) : Shimada A , Ueguchi-Tanaka M , Nakatsu T , Nakajima M , Naoe Y , Ohmiya H , Kato H , Matsuoka M
Ref : Nature , 456 :520 , 2008
Abstract : Gibberellins (GAs) are phytohormones essential for many developmental processes in plants. A nuclear GA receptor, GIBBERELLIN INSENSITIVE DWARF1 (GID1), has a primary structure similar to that of the hormone-sensitive lipases (HSLs). Here we analyse the crystal structure of Oryza sativa GID1 (OsGID1) bound with GA(4) and GA(3) at 1.9 A resolution. The overall structure of both complexes shows an alpha/beta-hydrolase fold similar to that of HSLs except for an amino-terminal lid. The GA-binding pocket corresponds to the substrate-binding site of HSLs. On the basis of the OsGID1 structure, we mutagenized important residues for GA binding and examined their binding activities. Almost all of them showed very little or no activity, confirming that the residues revealed by structural analysis are important for GA binding. The replacement of Ile 133 with Leu or Val-residues corresponding to those of the lycophyte Selaginella moellendorffii GID1s-caused an increase in the binding affinity for GA(34), a 2beta-hydroxylated GA(4). These observations indicate that GID1 originated from HSL and was further modified to have higher affinity and more strict selectivity for bioactive GAs by adapting the amino acids involved in GA binding in the course of plant evolution.
ESTHER : Shimada_2008_Nature_456_520
PubMedSearch : Shimada_2008_Nature_456_520
PubMedID: 19037316
Gene_locus related to this paper: orysa-gid1

Title : Regulation of insulin-like growth factor binding protein-1 and lipoprotein lipase by the aryl hydrocarbon receptor - Minami_2008_J.Toxicol.Sci_33_405
Author(s) : Minami K , Nakajima M , Fujiki Y , Katoh M , Gonzalez FJ , Yokoi T
Ref : Journal of Toxicological Sciences , 33 :405 , 2008
Abstract : The aryl hydrocarbon receptor (Ahr), a ligand-activated transcriptional factor, mediates the transcriptional activation of a battery of genes encoding drug metabolism enzymes. In the present study, we investigated the hepatic mRNA expression profile in Ahr-null (Ahr KO) mice compared to wild-type mice by microarray analysis to find new Ahr target genes. Pooled total RNA samples of liver extracted from 7- and 60-week-old Ahr KO or wild-type mice were studied by DNA microarray representing 19,867 genes. It was demonstrated that 23 genes were up-regulated and 20 genes were down-regulated over 2 fold in Ahr KO mice compared with wild-type mice commonly within the different age groups. We focused on insulin-like growth factor binding protein-1 (Igfbp-1) and lipoprotein lipase (Lpl) that were up-regulated in Ahr KO mice. The higher expression in Ahr KO mice compared to wild-type mice were confirmed by real-time RT-PCR analysis. In the wild-type mice but not in the Ahr KO mice, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) treatment increased the Igfbp-1 and Lpl mRNA levels. The expression profile of Igfbp-1 protein was consistent with that of Igfbp-1 mRNA. Since Lpl is the primary enzyme responsible for hydrolysis of lipids in lipoproteins, the serum triglyceride levels were determined. Indeed, the serum triglyceride levels in Ahr KO mice was lower than that in wild-type mice in accordance with the Lpl mRNA levels. Contrary to our expectation, TCDD treatment significantly increased the serum triglyceride levels in wild-type, but did not in Ahr KO mice. These results suggest that serum triglyceride levels are not correlated with hepatic Lpl expression levels. In the present study, we found that Ahr paradoxically regulates Igfbp-1 and Lpl expressions in the liver.
ESTHER : Minami_2008_J.Toxicol.Sci_33_405
PubMedSearch : Minami_2008_J.Toxicol.Sci_33_405
PubMedID: 18827440

Title : Structure and characterization of human carboxylesterase 1A1, 1A2, and 1A3 genes - Fukami_2008_Pharmacogenet.Genomics_18_911
Author(s) : Fukami T , Nakajima M , Maruichi T , Takahashi S , Takamiya M , Aoki Y , McLeod HL , Yokoi T
Ref : Pharmacogenet Genomics , 18 :911 , 2008
Abstract : OBJECTIVE: Human carboxylesterase (CES) 1A1 gene (14 exons) and CES1A3 pseudogene (six exons) are inverted and duplicated genes in a reference sequence (NT_010498). In contrast, earlier studies reported the CES1A2 gene (14 exons) instead of the CES1A3 pseudogene. The sequences of the CES1A2 gene downstream and upstream of intron 1 are identical with those of the CES1A1 and CES1A3 genes, respectively. A CES1A1 variant of which exon 1 is converted with that of the CES1A3 gene (the transcript is CES1A2) has recently been identified. We sought to clarify the confusing gene structure of human CES1A.
METHODS: A panel of 55 human liver as well as 318 blood samples (104 Caucasians, 107 African-Americans, and 107 Japanese) was used to clarify the gene structures of CES1A1, CES1A2, and CES1A3. Real-time reverse transcription-PCR and western blot analysis were carried out. Imidapril hydrolase activity in human liver microsomes and cytosol was determined by liquid chromatography-mass spectrometry (LC-MS)/MS.
RESULTS: By PCR analyses, we found that the CES1A2 gene is a variant of the CES1A3 gene. Four haplotypes, A (CES1A1 wild type and CES1A3), B (CES1A1 wild type and CES1A2), C (CES1A1 variant and CES1A3), and D (CES1A1 variant and CES1A2), were demonstrated. Ethnic differences were observed in allele frequencies of CES1A1 variant (17.3% in Caucasians and African-Americans and 25.2% in Japanese) and CES1A2 gene (14.4% in Caucasians, 5.1% in African-Americans, and 31.3% in Japanese). In human livers whose diplotype was A/A and C/C or C/D, no CES1A2 and CES1A1 mRNA was detected, respectively. In the other participants, the CES1A1 mRNA levels were higher than the CES1A2 mRNA levels. The CES1A proteins translated from CES1A1 mRNA and CES1A2 mRNA were detected in both human liver microsomes and cytosol fractions suggesting that the differences in exon 1 encoding a signal peptide did not affect the subcellular localization. Imidapril hydrolase activities reflected the CES1A protein levels. CONCLUSION: We found that the CES1A2 gene is a variant of the CES1A3 pseudogene. The findings presented here significantly increase our understanding about the gene structure and expression properties of human CES1A.
ESTHER : Fukami_2008_Pharmacogenet.Genomics_18_911
PubMedSearch : Fukami_2008_Pharmacogenet.Genomics_18_911
PubMedID: 18794728
Gene_locus related to this paper: human-CES1

Title : Molecular interactions of a soluble gibberellin receptor, GID1, with a rice DELLA protein, SLR1, and gibberellin - Ueguchi-Tanaka_2007_Plant.Cell_19_2140
Author(s) : Ueguchi-Tanaka M , Nakajima M , Katoh E , Ohmiya H , Asano K , Saji S , Hongyu X , Ashikari M , Kitano H , Yamaguchi I , Matsuoka M
Ref : Plant Cell , 19 :2140 , 2007
Abstract : GIBBERELLIN INSENSITIVE DWARF1 (GID1) encodes a soluble gibberellin (GA) receptor that shares sequence similarity with a hormone-sensitive lipase (HSL). Previously, a yeast two-hybrid (Y2H) assay revealed that the GID1-GA complex directly interacts with SLENDER RICE1 (SLR1), a DELLA repressor protein in GA signaling. Here, we demonstrated, by pull-down and bimolecular fluorescence complementation (BiFC) experiments, that the GA-dependent GID1-SLR1 interaction also occurs in planta. GA(4) was found to have the highest affinity to GID1 in Y2H assays and is the most effective form of GA in planta. Domain analyses of SLR1 using Y2H, gel filtration, and BiFC methods revealed that the DELLA and TVHYNP domains of SLR1 are required for the GID1-SLR1 interaction. To identify the important regions of GID1 for GA and SLR1 interactions, we used many different mutant versions of GID1, such as the spontaneous mutant GID1s, N- and C-terminal truncated GID1s, and mutagenized GID1 proteins with conserved amino acids replaced with Ala. The amino acid residues important for SLR1 interaction completely overlapped the residues required for GA binding that were scattered throughout the GID1 molecule. When we plotted these residues on the GID1 structure predicted by analogy with HSL tertiary structure, many residues were located at regions corresponding to the substrate binding pocket and lid. Furthermore, the GA-GID1 interaction was stabilized by SLR1. Based on these observations, we proposed a molecular model for interaction between GA, GID1, and SLR1.
ESTHER : Ueguchi-Tanaka_2007_Plant.Cell_19_2140
PubMedSearch : Ueguchi-Tanaka_2007_Plant.Cell_19_2140
PubMedID: 17644730

Title : Multiple loss-of-function of Arabidopsis gibberellin receptor AtGID1s completely shuts down a gibberellin signal - Iuchi_2007_Plant.J_50_958
Author(s) : Iuchi S , Suzuki H , Kim YC , Iuchi A , Kuromori T , Ueguchi-Tanaka M , Asami T , Yamaguchi I , Matsuoka M , Kobayashi M , Nakajima M
Ref : Plant J , 50 :958 , 2007
Abstract : Arabidopsis carries three receptor genes for the phytohormone gibberellin (GA), AtGID1a, AtGID1b and AtGID1c. Expression of each gene in the rice gid1-1 mutant for GA receptors causes reversion of its severely dwarfed phenotype and GA insensitivity to a normal level, even though each loss-of-function mutant shows no clear phenotype in Arabidopsis (Nakajima et al., 2006). In this paper, we report the functional redundancy and specificity of each AtGID1 by analyzing the multiple mutants for loss of function. Seeds of the double knockout mutants atgid1a atgid1b, atgid1a atgid1c and atgid1b atgid1c germinated normally. The double knockout mutant atgid1a atgid1c showed a dwarf phenotype, while other double mutants were of normal height compared to the wild-type. The stamens of the double knockout mutant atgid1a atgid1b were significantly shorter than those of the wild-type, and this leads to low fertility. A severe disarrangement of the pattern on its seed surface was also observed. The triple knockout mutant atgid1a atgid1b atgid1c did not germinate voluntarily, and only started to grow when the seed coat was peeled off after soaking. Seedlings of the triple knockout mutants were severe dwarfs, only a few millimeters high after growing for 1 month. Moreover, the triple knockout seedlings completely lost their ability to respond to exogenously applied GA. These results show that all AtGID1s function as GA receptors in Arabidopsis, but have specific role(s) for growth and development.
ESTHER : Iuchi_2007_Plant.J_50_958
PubMedSearch : Iuchi_2007_Plant.J_50_958
PubMedID: 17521411
Gene_locus related to this paper: arath-AT5G27320 , arath-GID1B

Title : Identification and characterization of Arabidopsis gibberellin receptors - Nakajima_2006_Plant.J_46_880
Author(s) : Nakajima M , Shimada A , Takashi Y , Kim YC , Park SH , Ueguchi-Tanaka M , Suzuki H , Katoh E , Iuchi S , Kobayashi M , Maeda T , Matsuoka M , Yamaguchi I
Ref : Plant J , 46 :880 , 2006
Abstract : Three gibberellin (GA) receptor genes (AtGID1a, AtGID1b and AtGID1c), each an ortholog of the rice GA receptor gene (OsGID1), were cloned from Arabidopsis, and the characteristics of their recombinant proteins were examined. The GA-binding activities of the three recombinant proteins were confirmed by an in vitro assay. Biochemical analyses revealed similar ligand selectivity among the recombinants, and all recombinants showed higher affinity to GA(4) than to other GAs. AtGID1b was unique in its binding affinity to GA(4) and in its pH dependence when compared with the other two, by only showing binding in a narrow pH range (pH 6.4-7.5) with 10-fold higher affinity (apparent K(d) for GA(4) = 3 x 10(-8) m) than AtGID1a and AtGID1c. A two-hybrid yeast system only showed in vivo interaction in the presence of GA(4) between each AtGID1 and the Arabidopsis DELLA proteins (AtDELLAs), negative regulators of GA signaling. For this interaction with AtDELLAs, AtGID1b required only one-tenth of the amount of GA(4) that was necessary for interaction between the other AtGID1s and AtDELLAs, reflecting its lower K(d) value. AtDELLA boosted the GA-binding activity of AtGID1 in vitro, which suggests the formation of a complex between AtDELLA and AtGID1-GA that binds AtGID1 to GA more tightly. The expression of each AtGID1 clone in the rice gid1-1 mutant rescued the GA-insensitive dwarf phenotype. These results demonstrate that all three AtGID1s functioned as GA receptors in Arabidopsis.
ESTHER : Nakajima_2006_Plant.J_46_880
PubMedSearch : Nakajima_2006_Plant.J_46_880
PubMedID: 16709201
Gene_locus related to this paper: arath-AT5G27320 , arath-AT5G62180 , arath-GID1B

Title : GIBBERELLIN INSENSITIVE DWARF1 encodes a soluble receptor for gibberellin - Ueguchi-Tanaka_2005_Nature_437_693
Author(s) : Ueguchi-Tanaka M , Ashikari M , Nakajima M , Itoh H , Katoh E , Kobayashi M , Chow TY , Hsing YI , Kitano H , Yamaguchi I , Matsuoka M
Ref : Nature , 437 :693 , 2005
Abstract : Gibberellins (GAs) are phytohormones that are essential for many developmental processes in plants. It has been postulated that plants have both membrane-bound and soluble GA receptors; however, no GA receptors have yet been identified. Here we report the isolation and characterization of a new GA-insensitive dwarf mutant of rice, gid1. The GID1 gene encodes an unknown protein with similarity to the hormone-sensitive lipases, and we observed preferential localization of a GID1-green fluorescent protein (GFP) signal in nuclei. Recombinant glutathione S-transferase (GST)-GID1 had a high affinity only for biologically active GAs, whereas mutated GST-GID1 corresponding to three gid1 alleles had no GA-binding affinity. The dissociation constant for GA4 was estimated to be around 10(-7) M, enough to account for the GA dependency of shoot elongation. Moreover, GID1 bound to SLR1, a rice DELLA protein, in a GA-dependent manner in yeast cells. GID1 overexpression resulted in a GA-hypersensitive phenotype. Together, our results indicate that GID1 is a soluble receptor mediating GA signalling in rice.
ESTHER : Ueguchi-Tanaka_2005_Nature_437_693
PubMedSearch : Ueguchi-Tanaka_2005_Nature_437_693
PubMedID: 16193045
Gene_locus related to this paper: orysa-gid1

Title : Empirical analysis of transcriptional activity in the Arabidopsis genome - Yamada_2003_Science_302_842
Author(s) : Yamada K , Lim J , Dale JM , Chen H , Shinn P , Palm CJ , Southwick AM , Wu HC , Kim C , Nguyen M , Pham P , Cheuk R , Karlin-Newmann G , Liu SX , Lam B , Sakano H , Wu T , Yu G , Miranda M , Quach HL , Tripp M , Chang CH , Lee JM , Toriumi M , Chan MM , Tang CC , Onodera CS , Deng JM , Akiyama K , Ansari Y , Arakawa T , Banh J , Banno F , Bowser L , Brooks S , Carninci P , Chao Q , Choy N , Enju A , Goldsmith AD , Gurjal M , Hansen NF , Hayashizaki Y , Johnson-Hopson C , Hsuan VW , Iida K , Karnes M , Khan S , Koesema E , Ishida J , Jiang PX , Jones T , Kawai J , Kamiya A , Meyers C , Nakajima M , Narusaka M , Seki M , Sakurai T , Satou M , Tamse R , Vaysberg M , Wallender EK , Wong C , Yamamura Y , Yuan S , Shinozaki K , Davis RW , Theologis A , Ecker JR
Ref : Science , 302 :842 , 2003
Abstract : Functional analysis of a genome requires accurate gene structure information and a complete gene inventory. A dual experimental strategy was used to verify and correct the initial genome sequence annotation of the reference plant Arabidopsis. Sequencing full-length cDNAs and hybridizations using RNA populations from various tissues to a set of high-density oligonucleotide arrays spanning the entire genome allowed the accurate annotation of thousands of gene structures. We identified 5817 novel transcription units, including a substantial amount of antisense gene transcription, and 40 genes within the genetically defined centromeres. This approach resulted in completion of approximately 30% of the Arabidopsis ORFeome as a resource for global functional experimentation of the plant proteome.
ESTHER : Yamada_2003_Science_302_842
PubMedSearch : Yamada_2003_Science_302_842
PubMedID: 14593172
Gene_locus related to this paper: arath-AT2G42690 , arath-AT4g30610 , arath-At5g13640 , arath-AT5G20520 , arath-AT5G27320 , arath-CGEP , arath-clh1 , arath-clh2 , arath-CXE12 , arath-CXE15 , arath-SCP25 , arath-F14F8.240 , arath-MES6 , arath-LCAT1 , arath-PLA11 , arath-PLA15 , arath-PLA16 , arath-PLA17 , arath-SCP8 , arath-SCP11 , arath-SCP40 , arath-MES14 , arath-AXR4 , arath-SFGH , arath-B9DFR3 , arath-pae2

Title : [Simultaneous determination of propanil, carbaryl and 3,4-dichloroaniline in human serum by HPLC with UV detector following solid phase extraction] - Hori_2002_Yakugaku.Zasshi_122_247
Author(s) : Hori Y , Nakajima M , Fujisawa M , Shimada K , Hirota T , Yoshioka T
Ref : Yakugaku Zasshi , 122 :247 , 2002
Abstract : In case of poisoning by herbicide compounded with Propanil (DCPA) and Carbaryl (NAC), we attempted simultaneous solid-phase extractions of DCPA, NAC, and 3,4-dichloroaniline (DCA), a metabolite of DCPA, from the patient's serum, and quantitative analytical method using HPLC-UV detection. With this HPLC method, the quantitative detection limits in the serum are 0.005 microgram/ml for DCPA and DCA and 0.001 microgram/ml for NAC, and the UV spectra of all three compounds could easily be obtained using a diode-array detection limit of 0.05 microgram/ml. When the three compounds were added to serum at concentrations ranging from 0.1-10.0 micrograms/ml, the recovery rates were satisfactory at between 91.1% and 101.9%. On analysis of the serum of patient who had ingested Kusanon A Emulsion, the ingested substance apparently caused an increase in the DCA concentration, which led to the appearance of methemoglobinemia. The possibility that the DCA concentration might be used for prognostic purposes was suggested.
ESTHER : Hori_2002_Yakugaku.Zasshi_122_247
PubMedSearch : Hori_2002_Yakugaku.Zasshi_122_247
PubMedID: 11905049

Title : Functional annotation of a full-length Arabidopsis cDNA collection - Seki_2002_Science_296_141
Author(s) : Seki M , Narusaka M , Kamiya A , Ishida J , Satou M , Sakurai T , Nakajima M , Enju A , Akiyama K , Oono Y , Muramatsu M , Hayashizaki Y , Kawai J , Carninci P , Itoh M , Ishii Y , Arakawa T , Shibata K , Shinagawa A , Shinozaki K
Ref : Science , 296 :141 , 2002
Abstract : Full-length complementary DNAs (cDNAs) are essential for the correct annotation of genomic sequences and for the functional analysis of genes and their products. We isolated 155,144 RIKEN Arabidopsis full-length (RAFL) cDNA clones. The 3'-end expressed sequence tags (ESTs) of 155,144 RAFL cDNAs were clustered into 14,668 nonredundant cDNA groups, about 60% of predicted genes. We also obtained 5' ESTs from 14,034 nonredundant cDNA groups and constructed a promoter database. The sequence database of the RAFL cDNAs is useful for promoter analysis and correct annotation of predicted transcription units and gene products. Furthermore, the full-length cDNAs are useful resources for analyses of the expression profiles, functions, and structures of plant proteins.
ESTHER : Seki_2002_Science_296_141
PubMedSearch : Seki_2002_Science_296_141
PubMedID: 11910074
Gene_locus related to this paper: arath-CXE15

Title : Small-angle X-ray scattering analysis of stearic acid modified lipase - Maruyama_2001_Biosci.Biotechnol.Biochem_65_1003
Author(s) : Maruyama T , Nakajima M , Ichikawa S , Sano Y , Nabetani H , Furusaki S , Seki M
Ref : Biosci Biotechnol Biochem , 65 :1003 , 2001
Abstract : Stearic acid modified lipase (from Rhizopus japonicus) exhibited remarkable interesterification activity in n-hexane, but crude native lipase did not. The structure of the fatty acid modified lipase had not been analyzed until now. We analyzed the modified lipase by small-angle X-ray scattering (SAXS) measurements in order to clarify the structure. SAXS measurements showed that the modified lipase consisted of a lipid lamellar structure and implied that the lipase was incorporated into the lamellar structure of stearic acid. The long spacings in the lamellar structures of the modified lipase and stearic acid were measured.
ESTHER : Maruyama_2001_Biosci.Biotechnol.Biochem_65_1003
PubMedSearch : Maruyama_2001_Biosci.Biotechnol.Biochem_65_1003
PubMedID: 11388447

Title : [A case of carbamate poisoning in which GCMS was useful to identify causal substance and to decide the appropriate treatment] - Kinoshita_2001_Chudoku.Kenkyu_14_343
Author(s) : Kinoshita H , Hirose Y , Tanaka T , Hori Y , Nakajima M , Fujisawa M , Oseki M
Ref : Chudoku Kenkyu , 14 :343 , 2001
Abstract : We often have cases of insecticide poisoning where the patient is unconscious and the causal substances are unknown We report an 83-year-old unconscious man who had apparently ingested several agricultural chemicals possibly organophosphate or carbamate According to his family there were three kinds of containers of agricultural chemicals with their caps opened around him When he was transferred to our hospital he presented hypertension hypersalivation and muscle fasciculation His pupils were markedly miotic In order to identify the substances ingested we used a gas chromatographymass spectrometer GCMS using his gastric content Within 30 minutes we were able to identify the causal substance as methomyl one of the popular carbamates thereby eliminating the need to use pralidoxime PAM GCMS makes it possible to identify unknown substances quickly and accurately and is therefore extremely useful in deciding the appropriate treatment
ESTHER : Kinoshita_2001_Chudoku.Kenkyu_14_343
PubMedSearch : Kinoshita_2001_Chudoku.Kenkyu_14_343
PubMedID: 11806102

Title : Effect of hydrocarbon-water interfaces on synthetic and hydrolytic activities of lipases - Maruyama_2001_J.Biosci.Bioeng_92_242
Author(s) : Maruyama T , Nakajima M , Seki M
Ref : J Biosci Bioeng , 92 :242 , 2001
Abstract : We have developed a new method of lipase activation for interesterification, in which lipase, in contact with a hydrocarbon-water interface, has been found to have high interesterification activity in an anhydrous solvent. We have applied this activation method to various lipase and obtained high synthetic activity in n-hexane, and have investigated the effect of various hydrocarbon-water interfaces on the synthetic and hydrolytic activities of lipases. The esterification and/or interesterification activity of lipases tested was improved by this activation method, using an n-tetradecane-water interface. From the initial group of lipases, three representative lipases (from Rhizopus japonicus, Chromobacterium viscosum and porcine pancreas) were selected for further study. The effect of various hydrocarbon-water interfaces on synthetic (interesterification or esterification) activity was studied. We demonstrated that the resulting synthetic activity was affected by the choice of hydrocarbon-water interface and that there were differences in the effects of interfaces on the synthetic activity of these lipases.
ESTHER : Maruyama_2001_J.Biosci.Bioeng_92_242
PubMedSearch : Maruyama_2001_J.Biosci.Bioeng_92_242
PubMedID: 16233091

Title : Sarin-like and soman-like organophosphorous agents activate PLCgamma in rat brains - Niijima_1999_Toxicol.Appl.Pharmacol_156_64
Author(s) : Niijima H , Nagao M , Nakajima M , Takatori T , Matsuda Y , Iwase H , Kobayashi M
Ref : Toxicol Appl Pharmacol , 156 :64 , 1999
Abstract : We report that there is a time-related change in the phospholipase C (PLC) activities of rat brain cytosol and membrane fractions after iv injection of a soman-like or a sarin-like organophosphorous agent (bis(isopropyl methyl)phosphonate [BIMP] and bis(pinacolyl methyl)phosphonate [BPMP]). PLCgamma was activated in the brain cytosol fraction from BPMP-injected rats. The phosphorylating activity of rat brain membrane fractions were enhanced by BPMP treatment. The brain membrane fractions from BPMP-treated rats phosphorylated several proteins, including supposedly PLCgamma in the brain cytosol fraction from control rats in vitro. These results suggest that soman and sarin may stimulate a membrane tyrosine kinase, including growth factor receptors, directly or indirectly.
ESTHER : Niijima_1999_Toxicol.Appl.Pharmacol_156_64
PubMedSearch : Niijima_1999_Toxicol.Appl.Pharmacol_156_64
PubMedID: 10101100

Title : Detection of the sarin hydrolysis product in formalin-fixed brain tissues of victims of the Tokyo subway terrorist attack - Matsuda_1998_Toxicol.Appl.Pharmacol_150_310
Author(s) : Matsuda Y , Nagao M , Takatori T , Niijima H , Nakajima M , Iwase H , Kobayashi M , Iwadate K
Ref : Toxicol Appl Pharmacol , 150 :310 , 1998
Abstract : One of the hydrolysis products of sarin (isopropyl methylphosphonofluoridate) was detected in formalin-fixed brain tissues of victims poisoned in the Tokyo subway terrorist attack. Part of this procedure, used for the detection of sarin hydrolysis products in erythrocytes of sarin victims, has been described previously. The test materials were four individual cerebellums, which had been stored in formalin fixative for about 2 years. Sarin-bound acetylcholinesterase (AChE) was solubilized from these cerebellums, purified by immunoaffinity chromatography, and digested with trypsin. Then the sarin hydrolysis products bound to AChE were released by alkaline phosphatase digestion, subjected to trimethylsilyl derivatization (TMS), and detected by gas chromatography-mass spectrometry. Peaks at m/z 225 and m/z 240, which are indicative of TMS-methylphosphonic acid, were observed within the retention time range of authentic methylphosphonic acid. However, no isopropyl methylphosphonic acid was detected in the formalin-fixed cerebellums of these 4 sarin victims, probably because the isopropoxy group of isopropyl methylphosphonic acid underwent chemical hydrolysis during storage. This procedure will be useful for the forensic diagnosis of poisoning by protein-bound, highly toxic agents, such as sarin, which are easily hydrolysed. This appears to be the first time that intoxication by a nerve agent has been demonstrated by analyzing formalin-fixed brains obtained at autopsy.
ESTHER : Matsuda_1998_Toxicol.Appl.Pharmacol_150_310
PubMedSearch : Matsuda_1998_Toxicol.Appl.Pharmacol_150_310
PubMedID: 9653062

Title : Definitive evidence for the acute sarin poisoning diagnosis in the Tokyo subway - Nagao_1997_Toxicol.Appl.Pharmacol_144_198
Author(s) : Nagao M , Takatori T , Matsuda Y , Nakajima M , Iwase H , Iwadate K
Ref : Toxicol Appl Pharmacol , 144 :198 , 1997
Abstract : A new method was developed to detect sarin hydrolysis products from erythrocytes of four victims of sarin (isopropylmethylphosphonofluoridate) poisoning resulting from the terrorist attack on the Tokyo subway. Sarin-bound acetylcholinesterase (AChE) was solubilized from erythrocyte membranes of sarin victims, digested with trypsin, the sarin hydrolysis products bound to AChE were released by alkaline phosphatase digestion, and the digested sarin hydrolysis products were subjected to trimethylsilyl derivatization and detected by gas chromatography-mass spectrometry. Isopropylmethylphosphonic acid, which is a sarin hydrolysis product, was detected in all sarin poisoning, victims we examined and methylphosphonic acid, which is a sarin and soman hydrolysis product, was determined in all victims. Postmortem examinations revealed no macroscopic and microscopic findings specific to sarin poisoning and sarin and its hydrolysis products were almost undetectable in their blood. We think that the procedure described below will be useful for the forensic diagnosis of acute sarin poisoning.
ESTHER : Nagao_1997_Toxicol.Appl.Pharmacol_144_198
PubMedSearch : Nagao_1997_Toxicol.Appl.Pharmacol_144_198
PubMedID: 9169085

Title : Detection of sarin hydrolysis products from sarin-like organophosphorus agent-exposed human erythrocytes - Nagao_1997_J.Chromatogr.B.Biomed.Sci.Appl_701_9
Author(s) : Nagao M , Takatori T , Matsuda Y , Nakajima M , Niijima H , Iwase H , Iwadate K , Amano T
Ref : Journal of Chromatography B Biomed Sci Appl , 701 :9 , 1997
Abstract : A sarin-like organophosphorus agent, [bis(isopropyl methyl)phosphonate; BIMP], was synthesized. This agent has the same phosphonate group as sarin and also has the same anti-acetylcholinesterase activity potency as sarin. The ID50 and LD50 values of BIMP in mice after intravenous injection were 3.9 nM and 0.8 mg/kg, respectively. The AChE activities of their red blood cells and brains were dose-dependently reduced by intravenous BIMP. After preparation of experimental BIMP-exposed human red blood cells, BIMP-bound acetylcholinesterase (AChE) was solubilized from erythrocyte membranes, purified by immunoaffinity chromatography, digested with trypsin, and the sarin hydrolysis products bound to AChE were released by alkaline phosphatase digestion. The digested sarin hydrolysis products were subjected to trimethylsilyl (TMS) derivatization and detected by gas chromatography-mass spectrometry. Isopropyl methylphosphonic- and methylphosphonic acids, which are the sarin hydrolysis products, were detected in experimental BIMP-exposed human red blood cells. This new method, which enables sarin's hydrolysis products to be detected in BIMP-exposed erythrocytes, is a useful tool for studying sarin-poisoning victims.
ESTHER : Nagao_1997_J.Chromatogr.B.Biomed.Sci.Appl_701_9
PubMedSearch : Nagao_1997_J.Chromatogr.B.Biomed.Sci.Appl_701_9
PubMedID: 9389333