Current Drug Metabolism - Volume 9, Issue 6, 2008

During the past 80 years, research on Arylamine-Nacetyltransferases (CoASAc; NAT, EC 2.3.1.5) has produced many major discoveries that have helped scientists understand the basis for altered metabolism of drugs and xenobiotics. The first references to the relevance of N-acetylation in humans date back to 1926 [1]. Nearly fifty years ago, Evans et al. demonstrated that acetylation of the isoniazid was bimodally distributed and that in vivo acetylation status was inheritable [2, 3]. The milestone work carried out by Meyer et al. in the Biozentrum of the University of Basel around 1990 caused a dramatic impulse in present knowledge of NAcetyltransferases. This work included the demonstration that common polymorphisms in human NAT2 result in a decreased level of protein in the liver [4], the partial purification of human NAT2 [5], the cloning of NAT1 and NAT2 [5, 6] and the functional analysis of recombinant expressed protein products [4]. Further developments in the role of polymorphisms on NAT2 activity were carried out by Hein et al. [7], and it is not surprising that the number of publications on N-acetyltransferases increased sharply from an average of 30-50 publications per year before 1990 to about 300 publications per year after 1991. Sinclair et al. in 2000 provided the first crystal structure of a NAT protein [8] and recently high resolution crystal structures of human NAT1 and NAT2 have been solved [9], revealing prominent features of NAT enzymes that will be of great help to further studies on structure, function and clinical relevance of NAT enzymes. The complexity of variations in NAT1 and NAT2 genes has put NAT in the forefront of pharmacogenetics research. Four decades after the recognition and effect of NAT2 polymorphism on isoniazide toxicity [10], present genotyping techniques allow the detection of major genetic variations in these genes and permit acetylation phenotypes to be inferred from genotyping data by using bioinformatic tools [11]. The determination of the NAT2 genotype or phenotype was initially proposed to predict adverse reactions in patients with tuberculosis receiving isoniazid [12], before the concomitant administration of procainamide and phenytoin [13], and to analyze the role of NAT2 in drug interactions [14]. These effects together with the high frequency for individuals with impaired NAT2 metabolism [4] make NAT2 a relevant target for pharmacogenomic tests in clinical practice. Since NAT gene polymorphisms have been considered as putative risk factors for some xenobiotic-related cancers because of the prominent role of NAT enzymes in drug and carcinogen activation and detoxification, we can now take advantage of these pharmacogenomic tests to assess cancer risk and the risk of other human diseases. Presently, research on Arylamine- N-acetyltransferases constitutes a major topic in pharmacology and pharmacogenomics and has therapeutic, preventive, anthropologic and even forensic implications. The bibliometric impact of Arylamine-N-acetyltransferases is impressive. In recent years, over 5,000 papers related to drug acetylation and NAT polymorphisms have been published. These papers accumulate over 100,000 citations and an h-index over 120. Nowadays an average of six articles on this topic are published per week. This special issue of Current Drug Metabolism on Nacetyltransferases provides a collection of review articles covering relevant basic and clinical topics in which NATs are of prominent relevance. The articles presented summarize the present knowledge of these topics, and identify further aspects that should be investigated in detail. The topics covered include the most recent advances in the structure of human NATs obtained after crystallization and direct structural analysis of human NAT1 and NAT2 as well as the potential applications of this information to the prediction of therapeutic and toxic effects [15]. A collection of papers related to genetic and non-genetic factors influencing NAT activities is also included in this special issue. The first of such papers is a comprehensive review of structure-function of variant NAT2 enzymes, including an analysis and molecular modeling of the effects of individual single nucleotide polymorphisms (SNPs) on NAT2 function and an update on NAT2 allele nomenclature [16]. The interethnic variability of human NAT2 SNPs, obtained after deconstruction of inferred variant alleles to avoid confounders, is analyzed in another paper that unravels the occurrence of intraethnic variability in NAT2 SNP frequencies and discusses the potential clinical impact of such intraethnic variability [17].

A large body of biochemical, kinetic and molecular information, accumulated over the course of more than 80 years, has produced valuable insights into the relationship between the structures and the catalytic functions of the human arylamine Nacetyltransferases NAT1 and NAT2. Much of the groundwork for the determination of human NAT structures and functions was provided by seminal biochemical and enzyme kinetic studies in both human and non-human model systems, the cloning and primary amino acid sequence determination of eukaryotic and prokaryotic NATs, the characterization of naturally occurring and artificially mutated forms of human NATs, elucidation of the crystal structures of several prokaryotic NAT orthologues, and information that has been derived from cross-species comparisons. In 2007 the progress of these studies was aided substantially by the successful crystallization and direct structural analysis of human NAT1 and NAT2. The purpose of this review is to give a brief historical perspective, to summarize our current understanding of human NAT structures and functions based on both earlier and more recent work, and to provide some future insights into the potential applications of this information to the prediction of therapeutic and toxic outcomes associated with the acetylation of primary aromatic amine- and hydrazine-containing chemicals.

Arylamine N-acetyltransferase 2 (NAT2) modifies drug efficacy/toxicity and cancer risk due to its role in bioactivation and detoxification of arylamine and hydrazine drugs and carcinogens. Human NAT2 alleles possess a combination of single nucleotide polymorphisms (SNPs) associated with slow acetylation phenotypes. Clinical and molecular epidemiology studies investigating associations of NAT2 genotype with drug efficacy/toxicity and/or cancer risk are compromised by incomplete and sometimes conflicting information regarding genotype/phenotype relationships. Studies in our laboratory and others have characterized the functional effects of SNPs alone, and in combinations present in alleles or haplotypes. We extrapolate this data generated following recombinant expression in yeast and COS-1 cells to assist in the interpretation of NAT2 structure. Whereas previous structural studies used homology models based on templates of N-acetyltransferase enzyme crystal structures from various prokaryotic species, alignment scores between bacterial and mammalian N-acetyltransferase protein sequences are low (∼ 30%) with important differences between the bacterial and mammalian protein structures. Recently, the crystal structure of human NAT2 was released from the Protein Data Bank under accession number 2PFR. We utilized the NAT2 crystal structure to evaluate the functional effects of SNPs resulting in the protein substitutions R64Q (G191A), R64W (C190T), I114T (T341C), D122N (G364A), L137F (A411T), Q145P (A434C), E167K (G499A), R197Q (C590A), K268R (A803G), K282T (A845C), and G286E (G857A) of NAT2. This analysis advances understanding of NAT2 structure-function relationships, important for interpreting the role of NAT2 genetic polymorphisms in bioactivation and detoxification of arylamine and hydrazine drugs and carcinogens.

Genetic polymorphisms of human arylamine N-acetyltransferase 2 (NAT2) are responsible for interindividual variation in the acetylation of numerous drugs and in the transformation of aromatic and heterocyclic amines into carcinogenic intermediates. Although large interethnic variability in the frequency for NAT2 variant alleles has been reported, comparison of allele frequencies is hampered by differences in the criteria for the assignment of allelic variants. To avoid such sources of bias, in this review we analyze the occurrence of both interethnic and intraethnic variability for the seven commonest single nucleotide polymorphisms (SNP) in the NAT2 gene by using raw SNP data instead of inferred haplotypes. Besides the large interethnic variability observed for all SNPs except C282T, intraethnic variability for NAT2 SNPs was identified for the SNPs G191A among Caucasians (p<0.0001), T341C among Oriental (p<0.001) or African individuals (p<0.012), C481T among Oriental (p<0.001) or African individuals (p<0.001), and G590A among Oriental individuals (p<0.001). In contrast, no major intraethnic differences were identified for the SNPs C282T, A806G or G857A. Intraethnic variability may have relevant clinical implications. For instance, case-control NAT2 studies should not be extrapolated from one Oriental population to another. Nonsynonymous SNPs occur in 32% of alleles in Japanese individuals and in 47% of alleles in Chinese individuals, therefore the frequency of adverse effects and cancer related to slow acetylation is expected to be higher in individuals with Chinese descent than in those with Japanese descent. Intraethnic variability reinforces the need for proper selection of control subjects and points against the use of surrogate control groups for studies involving association of NAT2 alleles with adverse drug effects or spontaneous diseases.

Acetylation catalysed by the arylamine N-acetyltransferases (NATs; 2.3.1.5) is a major biotransformation pathway for arylamine and hydrazine drugs, as well as many carcinogens that we are exposed to on a daily basis. These compounds can either be detoxified by NATs or bioactivated to metabolites that have the potential to cause toxicity such as cancer. As a result, the levels of NATs in the body have clinical importance with regard to drug effect and individual susceptibility to toxicity. Like many other drug metabolising enzymes, the activity of NATs varies considerably between individuals, due in part to genetic polymorphisms. However, it is becoming increasingly evident that non-genetic factors also play an important role in regulating NAT activity in vivo. This review focuses on the nongenetic control of NAT expression, including transcriptional, post-transcriptional/translational, and post-translational regulation. In addition, the dysregulation of NAT1 expression in cancer cells is reviewed, as this is an emerging area that may provide insight into a role for NAT1 in cancer biology.

Arylamine N-acetyltransferases (NAT) are xenobiotic-metabolizing enzymes responsible for the acetylation of many aromatic arylamine and heterocyclic amines, thereby playing an important role in both detoxification and activation of numerous drugs and carcinogens. Two closely related isoforms (NAT1 and NAT2) have been described in humans. NAT2 is mainly expressed in liver and gut, whereas NAT1 is found in a wide range of tissues. Interindividual variations in NAT genes have been shown to be a potential source of pharmacological and/or pathological susceptibility. In addition, there is now evidence that non genetic factors, such as substratedependent inhibition, drug interactions or cellular redox conditions may also contribute to NAT activity. The recent findings reviewed here provide possible mechanisms by which these environmental determinants may affect NAT activity. Interestingly, these data could contribute to the development of selective NAT inhibitors for the treatment of cancer and microbial diseases.

Polymorphic Human arylamine N-acetyltransferase (NAT2) inactivates the anti-tubercular drug isoniazid by acetyltransfer from acetylCoA. There are active NAT proteins encoded by homologous genes in mycobacteria including M. tuberculosis, M. bovis BCG, M. smegmatis and M. marinum. Crystallographic structures of NATs from M. smegmatis and M. marinum, as native enzymes and with isoniazid bound share a similar fold with the first NAT structure, Salmonella typhimurium NAT. There are three approximately equal domains and an active site essential catalytic triad of cysteine, histidine and aspartate in the first two domains. An acetyl group from acetylCoA is transferred to cysteine and then to the acetyl acceptor e.g. isoniazid. M. marinum NAT binds CoA in a more open mode compared with CoA binding to human NAT2. The structure of mycobacterial NAT may promote its role in synthesis of cell wall lipids, identified through gene deletion studies. NAT protein is essential for survival of M. bovis BCG in macrophage as are the proteins encoded by other genes in the same gene cluster (hsaA-D). HsaA-D degrade cholesterol, essential for mycobacterial survival inside macrophage. Nat expression remains to be fully understood but is co-ordinated with hsaA-D and other stress response genes in mycobacteria. Amide synthase genes in the streptomyces are also nat homologues. The amide synthases are predicted to catalyse intramolecular amide bond formation and creation of cyclic molecules, e.g. geldanamycin. Lack of conservation of the CoA binding cleft residues of M. marinum NAT suggests the amide synthase reaction mechanism does not involve a soluble CoA intermediate during amide formation and ring closure.

Human arylamine N-acetyltransferases (CoASAc; NAT, EC 2.3.1.5) NAT1 and NAT2 play a key role in the metabolism of drugs and environmental chemicals and in the metabolic activation and detoxification of procarcinogens. Phenotyping analyses have revealed an association between NAT enzyme activities and the risk of developing several forms of cancer. As genotyping procedures have become available for NAT1 and NAT2 gene variations, hundreds of association studies on NAT polymorphisms and cancer risk have been conducted. Here we review the findings obtained from these studies. Evidence for a putative association of NAT1 polymorphism and myeloma, lung and bladder cancer, as well as association of NAT2 polymorphisms with non-Hodgkin lymphoma, liver, colorectal and bladder cancer have been reported. In contrast, no consistent evidence for a relevant association of NAT polymorphisms with brain, head & neck, breast, gastric, pancreatic or prostate cancer have been described. Although preliminary data are available, further well-powered studies are required to fully elucidate the role of NAT1 in most human cancers, and that of NAT2 in astrocytoma, meningioma, esophageal, renal, cervical and testicular cancers, as well as in leukaemia and myeloma. This review discusses controversial findings on cancer risk and putative causes of heterogeneity in the proposed associations, and it identifies topics that require further investigation, particularly mechanisms underlying association of NAT polymorphisms and risk for subsets of cancer patients with specific exposures, putative epistatic contribution of polymorphism for other xenobiotic-metabolising enzymes such as glutathione S-transferases of Cytochrome P450 enzymes, and genetic plus environmental interaction.

Polymorphic N-acetyl transferases (NAT) 1 and 2 are involved in detoxification of xenobiotic arylamines and hydralazines. These common environmental chemicals may be related to the pathogenesis of many spontaneous disorders, mainly malignancies, but also disimmune or degenerative diseases, for which a polygenic predisposition has been suggested. Hence, polymorphic NAT genes (NAT2 has been the most studied one) may be low-penetrance risk genes for some of these disorders. Although a relation of risk may be definitely discarded for systemic lupus erythematosus (SLE), inflammatory bowel disease and endometriosis, more research is needed for rheumatoid arthritis, Parkinson's, Alzheimer's, Behcet's and periodontal diseases , as current results are inconclusive but suggest a possible relation with NAT2 polymorphism. In diabetes mellitus the possible relation with the rapid phenotype may be due to acquired metabolic changes and more genotyping studies are needed. NAT2 slow metabolizers are more prone to the side effects of polymorphically acetylated drugs, as is the SLE-like syndrome induced by hydralazine and procainamide, the side effects due to sulphasalazine and the skin rash secondary to many sulphonamides. Future research should be based on well-designed studies, with adequate sample sizes and homogeneous recruitment criteria, to obviate the proliferation of small studies that are time- and resource-consuming without offering definite answers.

There are a lot of pharmaceutical substances nowadays on the market. More than 1000 drugs have been implicated in causing liver diseases in more than one occasion. The liver is the most massive and important internal organ of human body. The morphological and functional integrity of the liver is vital to the health of the human organism. Xenobiotic biotransformation is the principal mechanism for maintaining homeostasis during exposure of organisms to small foreign molecules, such as drugs. Most drugs are lipophilic and they become more hydrophilic by xenobiotic metabolizing enzymes. Arylamine N acetyltransferases (NAT) convert aromatic amines or hydrazines to aromatic amides and hydrazides. A lot of generally used drugs contain aromatic amine or hydrazine groups. Drug-induced liver injury (DILI) is the grave problem in the present world. The frequency of DILI is 15-40 cases per 100000 persons per year with 6 % mortality rate on average. This review is devoted to the analyses of arylamine N-acetyltransferases role in DILI. The NAT gene polymorphism and slow phenotype are associated with predisposition to hepatotoxicity during drug-specific treatment. NAT activity is changed by smoking, viral infections and variety of drugs. It is shown that the involving of NAT in pathogenic processes of DILI such as inflammatory or immune response, formation reactive metabolites, oxidative stress, cholestasis.

An increasing prevalence of asthma noted worldwide has stimulated research on the phenotypic complexity resulting from interaction between the genetic and environmental components. Particularly, an increase in the prevalence of allergic rhinitis and asthma in industrialized countries indicate the importance of pulmonary metabolism of environmental xenobiotics. The arylamine Nacetyltransferases (NATs) are a unique family of enzymes that are involved in the biotransformation and detoxification of hydrazine and arylamine drugs/xenobiotics and could have a major role to play in atopy/asthma pathogenesis. Association studies on NAT1 and NAT2 polymorphisms focused in this review indicate the genetic significance of slow acetylation phenotype in bronchial and occupational asthma as well as in other allergic diseases in different populations worldwide. In contrast, fast acetylators have been found to have higher susceptibility to contact allergic dermatitis. Further in-depth research on the functional role of N- acetylation phenotype in disease pathogenesis is the requisite of the day, so that N-acetylation polymorphisms could serve as a genetic marker. Also, such genetic variations may have important implications in the efficacy of drugs for asthma treatment. The present review also makes a comment on the role of Arylalkylamine N-Acetyltransferase, an important enzyme involved in the conversion of serotonin to melatonin, in asthma pathogenesis.

Coronary artery disease continues to be an important cause of mortality and morbidity. Sirolimus and paclitaxel eluting stents have become an important treatment for patients undergoing revascularization from coronary blockages. These drug eluting stents have enjoyed great success initially in preventing recurrences of adverse cardiac events and decreasing the incidences of repeat revascularizations. However, adverse effects, such as thrombosis, emanating from the use of these drug eluting stents has recently come to focus. Hence a better understanding of the mechanism of action of these drugs in preventing restenosis is important for the long term success and potential betterment of drug eluting stent technology. Herein we review and discuss the pathophysiology of restenosis, the basic mechanism of action of sirolimus and paclitaxel eluting stents and their limitations so as to create a scope for more efficient and novel drug eluting stents in the future.

As more and more evidence has become available, the link between gene and emergent disease has been made including cancer, heart disease and parkinsonism. Analyzing the diseases and designing drugs with respect to the gene and protein level obviously help to find the underlying causes of the diseases, and to improve their rate of cure. The development of modern molecular biology, biochemistry, data collection and analysis techniques provides the scientists with a large amount of gene data. To draw a link between genes and their relation to disease outcomes and drug discovery is a big challenge: How to analyze large datasets and extract useful knowledge? Combining bioinformatics with drug discovery is a promising method to tackle this issue. Most techniques of bioinformatics are used in the first two phases of drug discovery to extract interesting information and find important genes and/or proteins for speeding the process of drug discovery, enhancing the accuracy of analysis and reducing the cost. Gene identification is a very fundamental and important technique among them. In this paper, we have reviewed gene identification algorithms and discussed their usage, relationships and challenges in drug discovery and development.

Recent studies have shown that many promising new drug candidates were abandoned due to poor pharmacokinetic properties (PKs). Therefore, it is important to predict the PKs of compounds during the early stages of drug development. The volume of distribution (VD) is one of the most important PK parameters. When considered along with systemic clearance, the VD determines the biological half-life, which is used for designing suitable dosage regimens and rational formulations. At present, the methods used to predict VD include (i) the extrapolation of animal data, (ii) physiologically based pharmacokinetic (PBPK) modeling and (iii) in silico approaches that employ quantitative structure - pharmacokinetic relationships (QSPR). In this article, the latest progress in the field of VD prediction is summarized in terms of the above three areas, respectively, and these approaches are expected to be valuable for screening new drugs during the early stages of drug discovery and development.