Unfinished business: target-based drug discovery

The switch in the mid-1980s/early 1990s from a phenotypic approach to a target-based approach to drug discovery has been followed by low productivity of new drugs entering the market. Reasons for the (necessary) switch and unsolved problems with both approaches to drug discovery are discussed. The S-curve theory of new technology development and introduction can act as guide as to when an upturn in productivity can be expected; this should occur during the next decade leading possibly to a new golden age of drug discovery.     

Navigating tuberculosis drug discovery with target-based screening

Introduction: Mycobacterium tuberculosis kills more people than any other bacterial pathogen. New drugs are required to shorten the treatment time and provide a viable therapy for drug-resistant and latent forms of tuberculosis. The tuberculosis field has advanced considerably since the publication of the M. tuberculosis genome sequence. Today, researchers can build a high definition map of the pathogen's traits and behavior and select individual targets for chemical disruption. Areas covered: This review examines the discovery of current clinical and candidate tuberculosis drugs. It outlines recent developments in the selection of molecular targets for the discovery of new anti-mycobacterial agents. It appraises techniques that incorporate target knowledge into the screening protocol. These techniques include in silico, in vitro enzyme-based, differential antisense sensitivity and gene expression screening systems. The review also looks ahead to further techniques that may be applied in tuberculosis drug discovery. Expert opinion: The adoption of an 'either/or' approach to targeted or random tuberculosis drug screening is not expected. The historical success of random screening in providing the tuberculosis drugs currently in clinical use is likely to ensure that non-targeted protocols retain an important role in drug screening. However, a number of M. tuberculosis inhibitors in lead optimization and preclinical development have been discovered using targeted methods. Realization of the first clinically-approved tuberculosis drugs derived from targeted screening and continued refinements in targeted screening technologies are likely to increase the adoption of targeted approaches in the future.     

Navigating tuberculosis drug discovery with target-based screening

Introduction: Mycobacterium tuberculosis kills more people than any other bacterial pathogen. New drugs are required to shorten the treatment time and provide a viable therapy for drug-resistant and latent forms of tuberculosis. The tuberculosis field has advanced considerably since the publication of the M. tuberculosis genome sequence. Today, researchers can build a high definition map of the pathogen's traits and behavior and select individual targets for chemical disruption.

Areas covered: This review examines the discovery of current clinical and candidate tuberculosis drugs. It outlines recent developments in the selection of molecular targets for the discovery of new anti-mycobacterial agents. It appraises techniques that incorporate target knowledge into the screening protocol. These techniques include in silico, in vitro enzyme-based, differential antisense sensitivity and gene expression screening systems. The review also looks ahead to further techniques that may be applied in tuberculosis drug discovery.

Expert opinion: The adoption of an ‘either/or’ approach to targeted or random tuberculosis drug screening is not expected. The historical success of random screening in providing the tuberculosis drugs currently in clinical use is likely to ensure that non-targeted protocols retain an important role in drug screening. However, a number of M. tuberculosis inhibitors in lead optimization and preclinical development have been discovered using targeted methods. Realization of the first clinically-approved tuberculosis drugs derived from targeted screening and continued refinements in targeted screening technologies are likely to increase the adoption of targeted approaches in the future.     

Peptide ligands in antibacterial drug discovery: use as inhibitors in target validation and target-based screening

There is an urgent need to develop novel classes of antibiotics to counter the inexorable rise of resistant bacterial pathogens. Modern antibacterial drug discovery is focused on the identification and validation of novel protein targets that may have a suitable therapeutic index. In combination with assays for function, the advent of microbial genomics has been invaluable in identifying novel antibacterial drug targets. The major challenge in this field is the implementation of methods that validate protein targets leading to the discovery of new chemical entities. Ligand-directed drug discovery has the distinct advantage of having a concurrent analysis of both the importance of a target in the disease process and its amenability to functional modulation by small molecules. VITA is a process that enables a target-based paradigm by using peptide ligands for direct in vitro and in vivo validation of antibacterial targets and the implementation of high-throughput assays to identify novel inhibitory molecules. This process can establish sufficient levels of confidence indicating that the target is relevant to the disease process and inhibition of the target will lead to effective disease treatment.     

Phage display as a tool for protease ligand discovery

Proteolytic enzymes have been implicated as the pathological agent in a number of disease states. For this reason proteases are attractive therapeutic targets. Phage display of peptide libraries can be used to identify peptides that may be used either directly as inhibitors or serve as leads in the generation of prodrugs and peptidomimetics.     

Highly sensitive target-based whole-cell antibacterial discovery strategy by antisense RNA silencing

Examples of drug-resistant bacteria are increasing while the discovery of new antibiotics with new mechanisms of action has been essentially nonexistent. The antisense-based sensitization of bacterial targets in Staphylococcus aureus is one of the new approaches that provides increased sensitivity for the detection of target-specific antibiotics and whole-cell screening assays based on differential sensitivity of target-depleted strains. The screening of natural product extracts using this type of assay designed for condensing enzyme (FabH/FabF) targets of the fatty acid biosynthesis pathway led to the discovery of a number of target-specific inhibitors including the novel antibiotic platensimycin, which has displayed activity against various drug-resistant bacteria. The antisense-based discovery strategy, rationale and design of screening assays, and the application of such assays for screening of natural product extracts and the discovery of fatty acid condensing enzyme inhibitors are reviewed in this article.     

Applications of biophysical tools to target-based discovery of novel antibacterial leads

New antibacterial drugs are urgently needed to combat the growing problem of multidrug resistant bacterial infections. Major advances in bacterial genomics have uncovered many unexploited targets, leading to the possibility of discovering new antibacterials with novel mechanisms that would circumvent resistance. Many of these targets are soluble enzymes that vary in their degrees of mechanistic complexity. Protein crystallography as well as solution based biophysical methods are playing an increasingly important role in selecting, characterizing and validating promising targets as well as identifying and optimizing lead compounds that inhibit their functions. Advances made in recent years in sensitivity, resolution and throughput of biophysical tools are allowing multiple approaches to screening for hits and rational design of leads based on a deeper understanding of structure-activity relationships. However, the path from a lead compound to a safe and efficacious antibacterial drug still remains challenging. Structural and biophysical approaches have had less of an impact on this later phase of discovery than on the lead generation phase.     

Approaches to antibacterial drug discovery

The discovery of new antibacterial drugs can be based either upon empirical screening methods or structure-based design. Empirical methods utilise both intact bacteria and isolated biochemical targets for high throughput screening of natural product or chemical libraries to detect inhibitor leads. Structure-based methods for drug design are based upon understanding the molecular architecture of the active site in an appropriate target molecule. Empirical methods have been widely applied to screen for antibacterial agents and the introduction of combinatorial methods for the synthesis of chemical libraries considerably expands the potential of empirical screening methods. In contrast, structure-based drug design has not yet been widely applied to the development of antibacterial drugs, although it has proved to be a successful approach in other therapeutic areas. Recent advances in the sequencing of bacterial genomes will assist both empirical and structure-based approaches by identifying new, essential bacterial genes whose products may become the targets of new agents with selective antibacterial activity.     

In silico tools used for compound selection during target-based drug discovery and development

Introduction: The target-based drug discovery process, including target selection, screening, hit-to-lead (H2L) and lead optimization stage gates, is the most common approach used in drug development. The full integration of in vitro and/or in vivo data with in silico tools across the entire process would be beneficial to R&D productivity by developing effective selection criteria and drug-design optimization strategies.

Areas covered: This review focuses on understanding the impact and extent in the past 5 years of in silico tools on the various stage gates of the target-based drug discovery approach.

Expert opinion: There are a large number of in silico tools available for establishing selection criteria and drug-design optimization strategies in the target-based approach. However, the inconsistent use of in vitro and/or in vivo data integrated with predictive in silico multiparameter models throughout the process is contributing to R&D productivity issues. In particular, the lack of reliable in silico tools at the H2L stage gate is contributing to the suboptimal selection of viable lead compounds. It is suggested that further development of in silico multiparameter models and organizing biologists, medicinal and computational chemists into one team with a single accountable objective to expand the utilization of in silico tools in all phases of drug discovery would improve R&D productivity.     

Advances in phage display technology for drug discovery

Introduction: Over the past decade, several library-based methods have been developed to discover ligands with strong binding affinities for their targets. These methods mimic the natural evolution for screening and identifying ligand–target interactions with specific functional properties. Phage display technology is a well-established method that has been applied to many technological challenges including novel drug discovery.

Areas covered: This review describes the recent advances in the use of phage display technology for discovering novel bioactive compounds. Furthermore, it discusses the application of this technology to produce proteins and peptides as well as minimize the use of antibodies, such as antigen-binding fragment, single-chain fragment variable or single-domain antibody fragments like VHHs.

Expert opinion: Advances in screening, manufacturing and humanization technologies demonstrate that phage display derived products can play a significant role in the diagnosis and treatment of disease. The effects of this technology are inevitable in the development pipeline for bringing therapeutics into the market, and this number is expected to rise significantly in the future as new advances continue to take place in display methods. Furthermore, a widespread application of this methodology is predicted in different medical technological areas, including biosensing, monitoring, molecular imaging, gene therapy, vaccine development and nanotechnology.     

Phage display selection and evaluation of cancer drug targets

Techniques for the construction of phage display libraries of combinatorial proteins have dramatically improved. This has allowed researchers to expand the applications to the field of cancer biology. The most direct use of protein phage-displayed libraries is the selection of ligands for individual proteins. This includes identification of peptide ligands for receptor signaling molecules: integrins, cytokine and growth factor receptors. Selected peptides may be used as competitors for natural ligands and for the mapping of binding epitopes. This approach has been exploited for delineation of intracellular signal transduction pathways and for the selection of enzyme substrates and inhibitors. Recently, more complicated biological systems were used as targets for biopanning. This includes combination of soluble proteins, cellular surfaces and even the vasculature of whole organs. cDNA expression libraries in phage-based vectors have been recently introduced. The use of phage as a vector for targeted gene therapy is also considered. These and other applications of phage display for cancer research will be reviewed.     

In vitro protein display in drug discovery

Nucleic acid-encoded libraries have been used at different stages of the drug discovery process for the identification of polypeptide ligands and for target identification. Traditionally, phage display screening systems have been used to explore large libraries of peptides and proteins. Lately, novel protein selection technologies have been developed that work entirely in vitro and use the polymerase chain reaction (PCR) rather than cells to amplify genetic material. The simplicity of the linkage between the protein and its encoding nucleic acid leads to several advantages, including the use of larger libraries without the biases of cell-based amplification, greater control over binding conditions and the ease with which PCR-based mutagenesis and recombination can be incorporated. This review focuses on the latest improvements in this new generation of in vitro protein display techniques and discusses their applications to the drug discovery process.     

Structural biology approaches to antibacterial drug discovery

Antibacterial drug discovery has undertaken a major experiment in the 12 years since the first bacterial genomes were sequenced. Genome mining has identified hundreds of potential targets that have been distilled to a relatively small number of broad-spectrum targets (‘low-hanging fruit’) using the genetics tools of modern microbiology. Prosecuting these targets with high-throughput screens has led to a disappointingly small number of lead series that have mostly evaporated under closer scrutiny. In the meantime, multi-drug resistant pathogens are becoming a serious challenge in the clinic and the community and the number of pharmaceutical firms pursuing antibacterial discovery has declined. Filling the antibacterial development pipeline with novel chemical series is a significant challenge that will require the collaboration of scientists from many disciplines. Fortunately, advancements in the tools of structural biology and of in silico modeling are opening up new avenues of research that may help deal with the problems associated with discovering novel antibiotics.     

In vitro display technologies - new tools for drug discovery

Over the past decade, several ligand discovery techniques have been developed that mimic the process of natural evolution. Phage display technology is the most established of these methods and has been applied to numerous technological problems including the discovery of novel drugs. More recently, some new display technologies have emerged which, unlike phage display, operate entirely in vitro and have concomitant advantages. This review describes this new generation of display technologies and indicates how they might fit into the modern drug discovery process.     

Bacterial DNA replication enzymes as targets for antibacterial drug discovery

INTRODUCTION: The bacterial replisome is composed of a large number of enzymes, which work in exquisite coordination to accomplish chromosomal replication. Effective inhibition inside the bacterial cell of any of the 'essential' enzymes of the DNA replication pathway should be detrimental to cell survival. AREAS COVERED: This review covers DNA replication enzymes that have been shown to have a potential for delivering antibacterial compounds or drug candidates including: type II topoisomerases, a clinically validated target family, and DNA ligase, which has yielded inhibitors with in vivo efficacy. A few of the 'replisome' enzymes that are structurally and functionally well characterized and have been subjects of antibacterial discovery efforts are also discussed. EXPERT OPINION: Identification of several essential genes in the bacterial replication pathway raised hopes that targeting these gene products would lead to novel antibacterials. However, none of these novel, single gene targets have delivered antibacterial drug candidates into clinical trials. This lack of productivity may be due to the target properties and inhibitor identification approaches employed. For DNA primase, DNA helicase and other replisome targets, with the exception of DNA ligase, the exploitation of structure for lead generation has not been tested to the same extent that it has for DNA gyrase. Utilization of structural information should be considered to augment HTS efforts and initiate fragment-based lead generation. The complex protein-protein interactions involved in regulation of replication may explain why biochemical approaches have been less productive for some replisome targets than more independently functioning targets such as DNA ligase or DNA gyrase.     

Bacterial DNA replication enzymes as targets for antibacterial drug discovery

Introduction: The bacterial replisome is composed of a large number of enzymes, which work in exquisite coordination to accomplish chromosomal replication. Effective inhibition inside the bacterial cell of any of the ‘essential’ enzymes of the DNA replication pathway should be detrimental to cell survival.

Areas covered: This review covers DNA replication enzymes that have been shown to have a potential for delivering antibacterial compounds or drug candidates including: type II topoisomerases, a clinically validated target family, and DNA ligase, which has yielded inhibitors with in vivo efficacy. A few of the ‘replisome’ enzymes that are structurally and functionally well characterized and have been subjects of antibacterial discovery efforts are also discussed.

Expert opinion: Identification of several essential genes in the bacterial replication pathway raised hopes that targeting these gene products would lead to novel antibacterials. However, none of these novel, single gene targets have delivered antibacterial drug candidates into clinical trials. This lack of productivity may be due to the target properties and inhibitor identification approaches employed. For DNA primase, DNA helicase and other replisome targets, with the exception of DNA ligase, the exploitation of structure for lead generation has not been tested to the same extent that it has for DNA gyrase. Utilization of structural information should be considered to augment HTS efforts and initiate fragment-based lead generation. The complex protein–protein interactions involved in regulation of replication may explain why biochemical approaches have been less productive for some replisome targets than more independently functioning targets such as DNA ligase or DNA gyrase.