Twenty Years of Structural Biology in Drug Discovery

Approximately once a term, scientists working in structural biology and related areas in and near London, meet under the auspices of the London Structural Biology Club to discuss their recent research. These Club meetings generally involve four or five lectures followed by an informal discussion over beer and pizza, generously provided by a sponsoring company.

Most speakers at LSBC meetings are academic researchers and a typical presentation will highlight a novel structure or two, emphasising, for a specialist audience, some of the trickier aspects of how they were solved. The most recent meeting, however – held on 3 July 2012 in Birkbeck’s Clore Management Centre, and sponsored by specialist light scattering company Avid Nano – included one rather unusual talk. Dave Brown recently left the pharmaceutical giant Pfizer after sixteen years in its structural biology group. He now combines an academic post at the University of Kent at Canterbury with work in a new biotech company, Cangenix Molecular Solutions. During his time at Pfizer, Brown contributed his expertise in structural biology to drug discovery programmes for a wide range of cardiovascular, inflammatory and infectious diseases. The topic of his LSBC presentation was the evolution of structural biology in the pharmaceutical industry as he had experienced it during the last two decades.

The history of structural biology in drug discovery goes back a little longer than that, to the use of NMR by companies such as Abbott and Agouron to determine structure-activity relationships for compound series. The first drugs to be largely designed based on structural principles were the HIV protease inhibitors, which are covered in some depth in both section 5 and section 10 of PPS. The first protease inhibitor to be licensed for treating AIDS was Saquinavir (Invirase™), which entered the clinic in 1995, only ten years after the protease gene was first detected in the newly sequenced HIV genome.

By the mid-1990s, most major pharma companies had specialist structural biology groups. The most important developments since then have been driven by technical improvements in molecular biology, gene cloning and protein purification, and in diffraction technology. Synchrotrons such as the UK’s Diamond Light Source make their facilities available for commercial use at a price that large companies, at least, have no trouble affording. X-ray data collection is now extremely fast and can largely be automated so it has become accessible to non-expert users. Although dedicated structural biology groups in industry are no larger than they were in the 1990s, there are many more researchers there – medicinal chemists and molecular biologists – who spend some of their working life doing structural biology. Arguably, a reasonable knowledge of protein structure is essential for all scientists working in drug design.

But what, exactly, does structural biology contribute to the complex process of drug discovery? Once a likely protein target for a new discovery programme has been identified – an enzyme to be targeted by an inhibitor, for example – knowing its three-dimensional structure can help both in the selection of likely starting molecules or “hits” and in their development into “lead” compounds that are potent enough inhibitors to go forward to the later stages of drug discovery. In the related technique of fragment-based drug discovery, a huge number of very small compounds or fragments are screened against a target and those that bind to different parts of the protein’s active site are selected and linked together to form a larger and theoretically more potent compound. Typically, individual fragments bind to their target so weakly that they can only be identified experimentally rather than through computer modelling. Structural experiments can also be used to probe unexpected ligand-binding interactions, perhaps even identifying previously unknown functional binding sites, and to build selectivity for one member of a large protein family into a drug molecule. Selectivity is essential for the development of safe, non-toxic drugs that bind proteins such as kinases and G-protein coupled receptors (see section 11) that have hundreds of homologs in the human proteome.

Brown then went on to describe some of the work that he had been involved with during his career at Pfizer. Phosphodiesterases are enzymes that catalyse the breaking of phosphodiester bonds; this function is responsible for regulating the concentration of the essential cyclic nucleotide monophosphates in cells, and enzymes in this class are drug targets for a range of diseases. Cyclic GMP-specific phosphodiesterase type 5 or PDE5 is the target of Pfizer’s most famous drug, sildefanil, which is known worldwide as Viagra™ (link is to the Wikipedia entry). Researchers at Pfizer have used structure-based techniques to modify the basic structure of this compound into drugs with the potential to treat very different diseases, including Reynaud’s syndrome (in which the blood supply to the extremities is reduced in cold temperatures) and stroke recovery. The recent structure-led discovery of the mechanism by which another protein in this family, PDE4, regulates cyclic AMP may even lead to the design of drugs for a range of devastating neurological conditions including Alzheimer’s disease and schizophrenia.

Brown summed up his lecture with a series of recommendations for structural biologists working in industry (and by extension, for industrial researchers doing structural biology there). He advised them to get involved in drug discovery programmes at the earliest stage, to use structure to understand mechanisms of action rather than simply to design inhibitors and to collaborate between disciplines and sometimes even across companies. And, overall (and more prosaically), to work as quickly and cheaply as reasonable. These are not bad recommendations for academic researchers, either.

Commemorating World TB Day: Drugs from Plants

This post was written first for the Birkbeck College events blog and is re-posted here with permission.

World TB Day is held on 24 March every year, to mark the day in 1882 when Robert Koch, one of the fathers of microbiology, first announced that he had discovered the cause of tuberculosis (TB) – the bacterium now known as Mycobacterium tuberculosis (link to the TB proteome page in PPS section 5). Over 125 years since its discovery, and despite billions of dollars of investment in drug discovery, this bacterium is still a killer. The World Health Organisation estimates that about two billion people are infected with latent tuberculosis; in 2010, the last year for which full figures are available, over eight million people became ill with active tuberculosis, and 1.4 million people died from the disease. Two factors help make TB particularly deadly: it often occurs in people infected with the HIV virus, where it is one of the major causes of death, and drug resistant forms are becoming more common. In January 2012, Nature reported the identification in India of so-called “totally drug resistant” (TDR) tuberculosis, resistant to all anti-TB drugs in general use.

 Image of Mycobacterium tuberculosis bacteria

Photo credit: Janice Carr, Centers for Disease Control and Prevention, USA

In 2012 at Birkbeck, World TB Day coincided with the start of the College’s annual Science Week. Dr Sanjib Bhakta, head of the Mycobacteria Research Laboratory in the Department of Biological Sciences, organised a well-attended symposium on tuberculosis and its treatment. Besides two scientific presentations, the symposium featured a short video, Tuberculosis: The Real Story, highlighting the views of people affected by TB in the UK, and a panel discussion led by the grassroots volunteer organisation Results UK on some of the political challenges raised by tuberculosis.

Both science lectures focused on plants as a source of potential new drugs for tuberculosis. Professor Franz Bucar from the University of Graz in Austria highlighted the extreme chemical diversity of compounds that could be extracted from plants, particularly as compared to those found in the average synthetic compound library. Plants have always existed alongside their own microbial pathogens and have evolved natural antibiotics to protect themselves. Our ancestors, before the dawn of scientific medicine, used plant extracts to treat infectious disease, often quite successfully. The sub-discipline of ethnomedicine involves “mining” these traditional or historical remedies for pure chemicals that can be developed as, or modified into, drugs.

Bucar described a European herb, elecampane or Inula helenium, which is known to have been used to treat lung disease in the sixteenth century. He explained how a complex mixture of natural products derived from this plant had been tested against mycobacteria. Compounds found to have anti-mycobacterial activity were extracted and purified. Other plants have also yielded useful lead compounds; extracts of bark from a small tree with the Latin name of Berchemia discolor have even been shown to inhibit multi-drug resistant strains of Mycobacterium tuberculosis at useful concentrations.

Discovering antibacterial products in plant extracts, however, is only a first step towards drug discovery. Even when natural products like these compounds are found to be selective for bacterial over human cells, it is necessary to discover their mechanism of action; to modify them to optimize their activity; and, since plant sources are often scarce and extraction processes costly, to determine methods of synthesizing them in the laboratory.

The second scientific presentation was given by Dr. Bhakta himself and described current work in Birkbeck’s Mycobacteria Research Laboratory in searching for potential drugs for TB. These are needed not only to combat resistant forms of the bacteria but to improve current treatment regimens for “standard”, drug-sensitive TB. This requires a combination of four drugs to be taken for two months followed by two drugs for another four months, and many patients, particularly poorer and less well educated ones, fail to complete such a long and complex regimen. This in turn can lead to the development of further resistant strains.

Ideally, new drugs are required that target proteins not targeted by existing drugs, as resistance will be harder to develop. Mycobacteria have extremely complex cell walls, unlike those of other types of bacteria; they are essential for the bacteria to survive, and the enzymes used to synthesise them have no equivalents in mammalian genomes. These enzymes, therefore, have many of the characteristics of excellent drug targets.  Bhakta and his group have been exploring ways to inhibit the synthesis of the peptidoglycan that is one of the most important constituents of that cell wall. This molecule has been described as the bacterium’s “Achilles heel”, but no drugs targeting its synthesis have yet entered the clinic.

Mycobacteria synthesise peptidoglycan via a series of enzymes known as ligases, each of which adds a new link to the growing peptidoglycan chain. Bhaka’s group has focused on one of these ligases, termed MurE. This enzyme is essential for the bacterium to survive and it is conserved in all Mycobacterium tuberculosis strains. Working in collaboration with Professor Nick Keep, also in the Department of Biology, Bhakta solved the structure of MurE (PDB 2XTA) and showed it to have an active site that could in theory, at least, be occupied, and blocked, by a relatively small, “drug-like” molecule. He and his co-workers are now searching libraries of natural products for compounds that might inhibit this enzyme. They have identified promising MurE inhibitors from plants endemic to both Colombia and China, and are synthesizing analogues of these compounds for further testing.

It is unlikely that the next generation of anti-tuberculosis drugs will include any unchanged natural products. It is extremely likely, however, that natural products will yield the “scaffolds” on which these desperately needed drugs may be built, and perhaps one of these will be generated from within Bucar’s or Bhakta’s groups.