Current Drug Targets - Infectious Disorders - Volume 2, Issue 4, 2002

Genomic research is playing a critical role in the discovery of new antimicrobial drugs. The rapid increase in bacterial and eukaryotic genome sequences allows for new and innovative ways for obtaining antimicrobial protein targets. Here, we describe a two level strategy for target identification and validation using computers (in silico). First, large scale comparative analyses of genome sequences were used to identify highly conserved genes which might be essential for in vitro and / or in vivo survival of bacterial pathogens. Lab-based experiments provided confirmation or validation of the hypothesis of in silico essentiality for over 350 individual genes. Over 200 validated, broad spectrum, yet highly specific gene targets, were identified in community infection pathogens. The second part of the target discovery strategy is an in-depth evolutionary, structural and cellular analysis of key drug targets. As an example, phylogenetic and structural analyses suggest that sequence and binding-pocket conservation in FabH (β-ketoacyl-ACP synthase III) would allow for the development of small molecule inhibitors not only effective against a broad species spectrum of community bacterial pathogens but also as potential new therapies for tuberculosis and malaria.

The appearance of antibiotic resistant pathogens, including vancomycin resistant Staphylococcus aureus, in the clinic has necessitated the development of new antibiotics. The golden age of antibiotic discovery, in which potent selective compounds were readily extracted from natural product extracts is over and novel approaches need to be implemented to cover the therapeutic shortfall. The generation of huge quantities of bacterial sequence data has allowed the identification of all the possible targets for therapeutic intervention and allowed the development of screens to identify inhibitors. Here, we described a number of target classes in which genomics has contributed to its identification. As a result of analyzing sequence data, all of the tRNA synthetases and all of the two-component signal transduction systems were readily isolated, which would not have been easily identified if whole genome sequences were not available. Fatty acid biosynthesis is a known antibacterial target, but genomics showed which genes in that pathway had the appropriate spectrum to be considered as therapeutic targets. Genes of unknown function may seem untractable targets, but if those that are broad spectrum and essential are identified, it becomes valuable to invest time and effort to determine their cellular role. In addition, we discuss the role of genomics in developing technologies that assist in the discovery of new antibiotics including microarray gridding technology. Genomics can also increase the chemical diversity against which the novel targets can be screened.

The limitations of the therapeutic antifungals are becoming increasingly apparent in the clinic due to their modest efficacy against life-threatening systemic fungal infections. These antifungals belong to only a few structural classes that affect a small range of targets, some are quite toxic in humans while the use of others, particularly the azole drugs, has encouraged the emergence of resistant clinical isolates and the selection of innately resistant fungal pathogens. Only a few new drugs based on novel targets are in clinical development, and these may be insufficient to overcome the changing tide of fungal disease. In parallel with the successful completion of the Saccharomyces cerevisiae and human genome sequencing projects, an increasing number of genome sequencing projects are being initiated and completed for significant fungal pathogens. The growing repository of genomic information, which is complemented by decades of genetic and biochemical study, is now available for genome-wide analysis of gene function and for incisive inter-genomic comparison, with the S. cerevisiae and human genomes providing key points of reference. Functional genomic and comparative genomic techniques, many of which were developed with S. cerevisiae, are being applied to fungal pathogens with the aim of obtaining an integrated view of fungal biology and to extract targets suitable for drug discovery. This review describes some of these techniques, their limitations and their increasing contribution to the antifungal discovery process through effective gene annotation, target identification and prioritization, and in the optimization of antifungal leads.

Helicobacter pylori infects the gastric mucosa of almost half of the worlds population and infection is associated with several gastrointestinal diseases, ranging in severity from superficial and chronic gastritis to duodenal ulceration and gastric adenocarcinoma. Developing new therapeutics against a bacterium with such a unique niche has proven challenging, and the current therapy is complex and increase of bacterial resistance to current antimicrobials and treatment failure has identified a need for newer, more potent compounds. Access to the genomic sequence of several H. pylori isolates has allowed a more focused, target-specific approach to the development of new therapeutics.

The three-dimensional structures of Haemophilus influenzae proteins whose biological functions are unknown are being determined as part of a structural genomics project to ask whether structural information can assist in assigning the functions of proteins. The structures of the hypothetical proteins are being used to guide further studies and narrow the field of such studies for ultimately determining protein function. An outline of the structural genomics methodological approach is provided along with summaries of a number of completed and in progress crystallographic and NMR structure determinations. With more than twenty-five structures determined at this point and with many more in various stages of completion, the results are encouraging in that some level of functional understanding can be deduced from experimentally solved structures. In addition to aiding in functional assignment, this effort is identifying a number of possible new targets for drug development.

The genus Mycobacterium contains two of the most important human pathogens, Mycobacterium tuberculosis and Mycobacterium leprae, the etiologic agents of tuberculosis and leprosy, respectively. Other mycobacteria are mostly saprophytic organisms, living in soil and water, but some of them can cause opportunistic infections. The increasing incidence of tuberculosis as well as infections with non-tuberculous mycobacteria (NTM) in AIDS patients has renewed interest in molecular mechanisms of drug resistance in these pathogens. Mycobacteria show a high degree of intrinsic resistance to most common antibiotics. For instance, species from the M. tuberculosis complex (MTC) are intrinsically resistant to macrolides. Nevertheless, some semi-synthetic macrolides as the erythromycin derivatives clarithromycin, azithromycin and most recently the ketolides, are active against NTM, particularly Mycobacterium avium, and some of them are widely used for infection treatment. However, shortly after the introduction of these new drugs, resistant strains appeared due to mutations in the macrolide target, the ribosome. The mycobacterial cell wall with its specific composition and structure is considered to be a major factor in promoting the natural resistance of mycobacteria to various antibiotics. However, to explain the difference in macrolide sensitivity between the MTC and NTM, the synergistic contribution of a specific resistance mechanism might be required, in addition to possible differences in cell wall permeability.This mini-review summarizes the current knowledge on the natural and acquired macrolide resistance in mycobacteria, gives an overview of potential mechanisms implicated in the intrinsic resistance and brings recent data concerning a macrolide resistance determinant in the MTC.