Current Medicinal Chemistry - Volume 18, Issue 9, 2011

The bacterium Mycobacterium tuberculosis remains a major challenge to public health systems worldwide, especially affecting developing countries in Asia, Africa, Latin America and Eastern Europe. Tuberculosis (TB) is one the most common bacterial diseases of humans, and World Health Organization estimates that nearly thirty percent of the world's population is infected with M. tuberculosis in a latent form and, as a result, at risk of developing active TB [1]. Furthermore, the appearance of multidrug-resistant (MDR) and extensively drug-resistant (XDR, resistant to first- and second-line anti-TB drugs) strains of M. tuberculosis has worsened the situation. herein. This scenario makes clear the need for development of a new generation of successful drugs against TB. The development of a new drug is result of combination of biological activity and drug-like properties. These qualities can be evaluated by computational approaches in the initial stages of drug discovery and development. Being structure-based virtual screen (SBVS) the major methodology applied for this end. SBVS is a methodology that requires structural information about protein targets, and can be applied to test libraries of small-molecule compounds against important targets for drug design, including targets identified in the Mycobacterium tuberculosis [2]. The use of combination of different drugs is of pivotal importance to stop the appearance of multiple drug resistant (MDR) organisms by spontaneous genetic mutations, which can lead to an ineffective treatment. Therefore new proteins should be targeted for drug development. Several enzymes of the purine and pyrimidine salvage pathways [3,4], shikimate pathway [5], and mycolic acid biosynthetic pathway [6] have been validated as anti-TB targets. The present volume of Current Medicinal Chemistry brings reviews focused on protein targets identified in Mycobacterium tuberculosis. There are reviews about protein-drug interactions and structural basis for inhibition of protein targets identified in the Mycobacterium tuberculosis genome. Among them the enzymes of the shikimate pathway, which represent potential protein targets for developing antibacterial agents, and anti-parasite drugs, because these enzymes are of pivotal importance for bacteria and protozoans, but they are absent from humans [5]. The shikimate pathway is a connection between the metabolism of carbohydrates and the biosynthesis of aromatic compounds through seven metabolic steps. In these pathway phosphoenolpyruvate (PEP) and erythrose 4-phosphate are converted to chorismic acid [7,8]. Due to importance of this pathway, several structural studies focused on these enzymes were carried our [9-19]. This structural information opened the possibility for structure-based virtual screens, which may be able to identity new anti-tubercular drugs [20]. This volume presents two reviews with information about structural studies of shikimate kinase (EC 2.7.1.71) and chorismate synthase (EC 4.2.3.5)...... In addition, a review by Obiol-Pardo and collaborators describes the mechanism of action and inhibitors of the seven enzymes of the methylerythritol phosphate (MEP) pathway for isoprenoid biosynthesis, with special attention to the reported studies in M. tuberculosis. Enzymes of this pathway are also potential targets for antitubercular drugs. Three other metabolic pathways are reviewed here: purine salvage pathway (by Ducati and collaborators), pyrimidine salvage pathway (by Villela and collaborators) and mycolic acid biosynthetic pathway (by Singh and collaborators). It is also discussed here recent development of structure-based virtual screening methodologies, including a review on modern computational approaches to molecular docking simulations. Molecular docking is a computer simulation methodology to predict the conformations of a receptor-ligand complex [21-28]. It is possible to visualize that this simulation is analogous to the key-and-lock problem, where the lock is the receptor and the key the ligand. The goal in this kind of simulation is to adjust the position of the key in the lock. In a computer simulation it is generated many possible positions for the key in the lock, which are called poses. Docking simulations employ one or more of the following methodologies: Monte Carlo (MC) [29], fast shape matching (SM) [30], incremental construction (IC) [31, 32], distance geometry (DG) [33], simulated annealing (SA) [34, 35] and tabu search (TS) [36]. All these computational methodologies have been recently reviewed [37]. Although intense research has been performed on the application of the above mentioned algorithms to the problem of molecular docking simulations, recent results strongly indicate that the most successful approaches are those based on BIAs [27, 38], such as evolutionary programming (EP) [39, 40] and genetic algorithms (GA) [41-43]. These approaches and their application to development of antiTB drugs are discussed here. Structural aspects such as intermolecular hydrogen bonds, contact area and electrostatic interactions can be analyzed from threedimensional structures of complexes involving protein-targets and ligand [44]. Nevertheless, the precise analysis of protein-drug interactions from these structures is inadequate since crystal structures are average structures obtained from the molecules packed in the crystal lattice. It is hard to identify directly from crystallographic structure the flexible parts of the molecule. These limitations can be overcome by application of molecular dynamics simulations. Three-dimensional structures obtained experimentally or by homology modeling can be submitted to this simulation, where dynamical features of the complexes can be analyzed. These methods are also described in the present volume. Several recent applications of SBVS successful identified new drugs, which serve as incentives for development of new methodologies and also for extending the application to a wide range of protein targets and diseases [45-69]. In conclusion, one of the most defying challenges in the post-genomic era is the understanding of dynamics and structural features of the protein-drug interaction. Information obtained from structural studies of protein targets together with molecular docking and dynamics simulations will pave the way for discovery and development of a new generation of drugs. Finally, I would like to express gratitude to the authors for their significant contribution to this special issue, which hopefully will be of significance to researchers working in the development of a new generation of antiTB drugs.

Millions of deaths worldwide are caused by the aetiological agent of tuberculosis, Mycobacterium tuberculosis. The increasing prevalence of this disease, the emergence of drug-resistant strains, and the devastating effect of human immunodeficiency virus coinfection have led to an urgent need for the development of new and more efficient antimycobacterial drugs. The modern approach to the development of new chemical compounds against complex diseases, especially the neglected endemic ones, such as tuberculosis, is based on the use of defined molecular targets. Among the advantages, this approach allows (i) the search and identification of lead compounds with defined molecular mechanisms against a specific target (e.g. enzymes from defined pathways), (ii) the analysis of a great number of compounds with a favorable cost/benefit ratio, and (iii) the development of compounds with selective toxicity. The present review describes the enzymes of the purine salvage pathway in M. tuberculosis as attractive targets for the development of new antimycobacterial agents. Enzyme kinetics and structural data have been included to provide a thorough knowledge on which to base the search for compounds with biological activity. We have focused on the mycobacterial homologues of this pathway as potential targets for the development of new antitubercular agents.

The interest in analytical ultracentrifugation (AUC) to analyze protein structural parameters and interactions has increased in the past decades as a result of several developments on new generation instrumentation and data analysis tools. In this article, we review AUC principles and applications to study proteins, emphasizing molecular targets of Mycobacterium tuberculosis.

The causative agent of tuberculosis (TB), Mycobacterium tuberculosis, infects one-third of the world population. TB remains the leading cause of mortality due to a single bacterial pathogen. The worldwide increase in incidence of M. tuberculosis has been attributed to the high proliferation rates of multi and extensively drug-resistant strains, and to co-infection with the human immunodeficiency virus. There is thus a continuous requirement for studies on mycobacterial metabolism to identify promising targets for the development of new agents to combat TB. Singular characteristics of this pathogen, such as functional and structural features of enzymes involved in fundamental metabolic pathways, can be evaluated to identify possible targets for drug development. Enzymes involved in the pyrimidine salvage pathway might be attractive targets for rational drug design against TB, since this pathway is vital for all bacterial cells, and is composed of enzymes considerably different from those present in humans. Moreover, the enzymes of the pyrimidine salvage pathway might have an important role in the mycobacterial latent state, since M. tuberculosis has to recycle bases and/or nucleosides to survive in the hostile environment imposed by the host. The present review describes the enzymes of M. tuberculosis pyrimidine salvage pathway as attractive targets for the development of new antimycobacterial agents. Enzyme functional and structural data have been included to provide a broader knowledge on which to base the search for compounds with selective biological activity.

The enzymes of the shikimate pathway represent potential molecular targets for the development of non-toxic antimicrobial agents and anti-parasite drugs. One of the most promising of these enzymes is shikimate kinase (EC 2.7.1.71), which is responsible for the fifth step in the shikimate pathway. This enzyme phosphorylates shikimic acid to yield shikimate-3-phosphate, using ATP as a substrate. In this work, the conformational dynamics of the shikimate kinase from Mycobacterium tuberculosis was investigated in its apostate in solution. For this study, the enzyme was subjected to a gradient of temperatures from 15° C to 45 ° C in the presence or absence of deuterium oxide, and the amide H/D exchange was monitored using ESI-mass spectrometry. We observed: i) the phosphate binding domain in the apo-enzyme is fairly rigid and largely protected from solvent access, even at relatively high temperatures; ii) the shikimate binding domain is highly flexible, as indicated by the tendency of the apo-enzyme to exhibit large conformational changes to permit LID closure after the shikimate binding; iii) the nucleotide binding domain is initially conformationally rigid, which seems to favour the initial orientation of ADP/ATP, but becomes highly flexible at temperatures above 30°C, which may permit domain rotation; iv) part of the LID domain, including the phosphate binding site, is partially rigid, while another part is highly flexible and accessible to the solvent.

Tuberculosis is considered a worldwide health problem mainly due to co-infection with HIV and proliferation of multi-drugresistant strains. The enzymes of the shikimate pathway are potential targets for the development of new therapies because they are essential for bacteria, but absent from mammals. The last step in this pathway is performed by chorismate synthase (CS), which catalyzes the conversion of 5-enolpyruvylshikimate-3-phosphate (EPSP) to chorismate. The aim of this article is to review the available information on chorismate synthase from Mycobacterium tuberculosis.

The continuous preventive measures and control of tuberculosis are often hampered by re-emergence of multi-drug-resistant (MDR) strains of Mycobacterium tuberculosis. A novel drug approach is desperately needed to combat the global threat posed by MDR strains. In spite of current advancement in biological techniques viz. microarray and proteomics data for tuberculosis, no such potent drug has been developed in the past decades yet. Therefore, mycolic acid is an essential constituent which is involved in the formation of cell wall of Mycobacterium species. The biosynthesis of mycolic acid is involved in two fatty acid synthase systems, the multifunctional polypeptide fatty acid synthase I (FASI) which performs de novo fatty acid synthesis and dissociate FASII system. FASII system consists of monofunctional enzymes and acyl carrier protein (ACP), elongating FASI products to long chain mycolic acid precursor. In this review, the β-ketoacyl-ACP synthases (fadH, kasA and kasB) are distinct and play a vital role in mycolic acid synthesis, cell wall synthesis, biofilm formation and also pathogenesis. On the basis of substantial observation we suggest that these enzymes may be used as promising and attractive targets for novel anti-TB drugs designing and discovery.

Tuberculosis remains a major infectious disease to humans. It accounts for approximately 8-9 million new cases worldwide and an estimated 1.6 million deaths annually. Effective treatments for tuberculosis consist of a combination of several drugs administered over long periods of time. Since Mycobacterium tuberculosis often acquires multiple drug resistant mechanisms, development of new drugs with innovative actions is urgently required. The 2C-methyl-D-erythritol 4-phosphate (MEP) pathway, in charge of the essential biosynthesis of isoprenoids, represents a promising and selective target for developing new drugs against tuberculosis. To date, only fosmidomycin, a molecule that targets the second enzyme of the MEP pathway, has reached clinical trials but recent advances elucidating the structure and kinetics of the MEP enzymes are likely to change this scenario. This review describes the structure, mechanism of action and inhibitors of the seven enzymes of the MEP pathway, with special attention to the reported studies in M. tuberculosis.

Nature as a source of inspiration has been shown to have a great beneficial impact on the development of new computational methodologies. In this scenario, analyses of the interactions between a protein target and a ligand can be simulated by biologically inspired algorithms (BIAs). These algorithms mimic biological systems to create new paradigms for computation, such as neural networks, evolutionary computing, and swarm intelligence. This review provides a description of the main concepts behind BIAs applied to molecular docking simulations. Special attention is devoted to evolutionary algorithms, guided-directed evolutionary algorithms, and Lamarckian genetic algorithms. Recent applications of these methodologies to protein targets identified in the Mycobacterium tuberculosis genome are described.

Application of molecular dynamics simulation technique has become a conventional computational methodology to calculate significant processes at the molecular level. This computational methodology is particularly useful for analyzing the dynamics of proteinligand systems. Several uses of molecular dynamics simulation makes possible evaluation of important structural features found at interface between a ligand and a protein, such as intermolecular hydrogen bonds, contact area and binding energy. Considering structurebased virtual screening, molecular dynamics simulations play a pivotal role in understanding the features that are important for ligandbinding affinity. This information could be employed to select higher-affinity ligands obtained in screening processes. Many protein targets such as enoyl-[acyl-carrier-protein] reductase (InhA), purine nucleoside phosphorylase (PNP), and shikimate kinase have been submitted to these simulations and will be analyzed here. All command files used in this review are available for download at http://azevedolab.net/md_75.html.

Charge is an important characteristic of drug molecules, since ionization sites determine the pKa at a particular pH. The pKa in turn can affect many parameters, including solubility, dissolution rate, reaction kinetics, formulation, cell permeability, tissue distribution, renal elimination, metabolism, protein binding and receptor interactions. The impact of charge dynamics is amplified in human solid tumors that exhibit the glycolytic phenotype and associated acidic extracellular microenvironment. This phenotype is driven by hypoxia and creates a pH gradient in tumors that favors uptake of weak acids and exclusion of weak bases. Established anticancer drugs exhibit a range of pKa's and thus variable ability to exploit the tumor pH gradient. The camptothecins are a prime example as they represent a diverse class of approved anticancer drugs and drug candidates whose charge distribution varies with pH. An in silico method was used to predict charge distribution of camptothecins at physiological versus acidic pH in both the lactone and carboxylate forms. A significant amount of uncharged carboxylate was predicted at acidic pH that could enter tumor cells and accumulate in mitochondria to inhibit mitochondrial topoisomerase I. A model is presented to describe the charge dynamics of a new camptothecin analog and the impact on nuclear and mitochondrial mechanism(s) of action. This example illustrates the importance of integrating tumor physiology and charge dynamics into anticancer drug development.

Biologically active peptides and proteins have a great potential to act as targeted drug therapies in the treatment of a variety of diseases, including cancer. However, their use in vivo is limited by their low stability and cell permeability. Thus, it is necessary to develop efficient and safe peptide/protein delivery systems that can overcome these problems and increase a therapy's bioavailability. The search for promising vectors has led to the use of compounds called cell-penetrating peptides or protein transduction domains. The cellpenetrating peptides, as effective transporter, are utilized to enhance uptake of various biologically active peptide/protein cargos upon fusion or attachment to its sequences. Cell-penetrating peptides have been the subject of investigation of many researchers, however this review only focuses on the arginine-rich and amphipathic carriers and their potential therapeutic use.

The treatment of schizophrenia, one of the most debilitating mental illnesses, began by the serendipitous discovery of chlorpromazine. Since then, researchers have endeavored to find the cause of the illness but it remains unresolved. As a result, literature on the etiology of schizophrenia is littered with hypotheses and theories that are constantly reviewed, modified and rejected. Two hypotheses, however, have withstood the test of time and serve as the basis for the drug treatment, namely the dopamine and serotonin hypotheses. This review introduces the disease state, summarizes in detail the two leading hypotheses on schizophrenia, presents drugs that are currently available for treatment, and discusses some of the promising drug candidates based on their pre and early clinical trial results.

Vitamin A serves as substrate for the biosynthesis of several derivates (retinoids) which are important for cell growth and cell differentiation as well as for vision. Retinoic acid is the major physiologically active form of vitamin A regulating the expression of different genes. At present, hundreds of genes are known to be regulated by retinoic acid. This regulation is very complex and is, in turn, regulated on many levels. To date, two families of retinoid nuclear receptors have been identified: retinoic acid receptors and retinoid X receptors, which are members of the steroid hormone receptor superfamily of ligand-activated transcription factors. In order to regulate gene expression, all-trans retinal needs to be oxidized to retinoic acid. All-trans retinal, in turn, can be produced during oxidation of alltrans retinol or in a retinol-independent metabolic pathway through cleavage of β-carotene with all-trans retinal as an intermediate metabolite. Recently it has been shown that not only retinoic acid is an active form of vitamin A, but also that all-trans retinal can play an important role in gene regulation. In this review we comprehensively summarize recent literature on regulation of gene expression by retinoids, biochemistry of retinoid receptors, and molecular mechanisms of retinoid-mediated effects on gene regulation.