Martyn Banks

Impact of high-throughput screening in biomedical research

High-throughput screening (HTS) has been postulated in several quarters to be a contributory factor to the decline in productivity in the pharmaceutical industry. Moreover, it has been blamed for stifling the creativity that drug discovery demands. In this article, we aim to dispel these myths and present the case for the use of HTS as part of a proven scientific tool kit, the wider use of which is essential for the discovery of new chemotypes.

HTS approaches to voltage-gated ion channel drug discovery

Voltage-gated ion channels are emerging as a target class of increasing importance to the pharmaceutical industry because of their relevance to a wide variety of diseases in the cardiovascular, CNS and metabolic areas. In the quest to identify novel lead molecules against these targets, drug discovery programmes are increasingly making use of HTS approaches. The authors describe the current technologies available for voltage-gated ion-channel screening, their application to HTS campaigns and the current limitations and emerging technologies within this area.

Synthesis and biological evaluation of a library containing potentially 1600 amides / esters. A strategy for rapid compound generation and screening

A library of potentially 1600amide/ester dimers was prepared by reacting 40 acid chlorides with 40 nucleophiles. The whole library was presented for biological screening in 80 sample mixtures, with each of the possible products appearing in a unique pair of samples. The potential of this strategy for rapid identification of chemical leads was demostrated by the discovery of (1) with micromolar affinity for the NK3 receptor and (2), a weak inhibitor of the matrix metalloprotease MMP-1.

Discovery of positive allosteric modulators and silent allosteric modulators of the μ-opioid receptor

μ-Opioid receptors are among the most studied G protein-coupled receptors because of the therapeutic value of agonists, such as morphine, that are used to treat chronic pain. However, these drugs have significant side effects, such as respiratory suppression, constipation, allodynia, tolerance, and dependence, as well as abuse potential. Efforts to fine tune pain control while alleviating the side effects of drugs, both physiological and psychological, have led to the development of a wide variety of structurally diverse agonist ligands for the μ-opioid receptor, as well as compounds that target κ- and δ-opioid receptors. In recent years, the identification of allosteric ligands for some G protein-coupled receptors has provided breakthroughs in obtaining receptor subtype-selectivity that can reduce the overall side effect profiles of a potential drug. However, positive allosteric modulators (PAMs) can also have the specific advantage of only modulating the activity of the receptor when the orthosteric agonist occupies the receptor, thus maintaining spatial and temporal control of receptor signaling in vivo. This second advantage of allosteric modulators may yield breakthroughs in opioid receptor research and could lead to drugs with improved side-effect profiles or fewer tolerance and dependence issues compared with orthosteric opioid receptor agonists. Here, we describe the discovery and characterization of μ-opioid receptor PAMs and silent allosteric modulators, identified from high-throughput screening using a β-arrestin–recruitment assay.

Case study: impact of technology investment on lead discovery at Bristol–Myers Squibb, 1998–2006

We review strategic approaches taken over an eight-year period at BMS to implement new high-throughput approaches to lead discovery. Investments in compound management infrastructure and chemistry library production capability allowed significant growth in the size, diversity and quality of the BMS compound collection. Screening platforms were upgraded with robust automated technology to support miniaturized assay formats, while workflows and information handling technologies were streamlined for improved performance. These technology changes drove the need for a supporting organization in which critical engineering, informatics and scientific skills were more strongly represented. Taken together, these investments led to significant improvements in speed and productivity as well a greater impact of screening campaigns on the initiation of new drug discovery programs.

Extraction, Identification, and Functional Characterization of a Bioactive Substance From Automated Compound-Handling Plastic Tips

Disposable plastic labware is ubiquitous in contemporary pharmaceutical research laboratories. Plastic labware is routinely used for chemical compound storage and during automated liquid-handling processes that support assay development, high-throughput screening, structure-activity determinations, and liability profiling. However, there is little information available in the literature on the contaminants released from plastic labware upon DMSO exposure and their resultant effects on specific biological assays. The authors report here the extraction, by simple DMSO washing, of a biologically active substance from one particular size of disposable plastic tips used in automated compound handling. The active contaminant was identified as erucamide ((Z)-docos-13-enamide), a long-chain mono-unsaturated fatty acid amide commonly used in plastics manufacturing, by gas chromatography/mass spectroscopy analysis of the DMSO-extracted material. Tip extracts prepared in DMSO, as well as a commercially obtained sample of erucamide, were active in a functional bioassay of a known G-protein-coupled fatty acid receptor. A sample of a different disposable tip product from the same vendor did not release detectable erucamide following solvent extraction, and DMSO extracts prepared from this product were inactive in the receptor functional assay. These results demonstrate that solvent-extractable contaminants from some plastic labware used in the contemporary pharmaceutical research and development (R&D) environment can be introduced into physical and biological assays during routine compound management liquid-handling processes. These contaminants may further possess biological activity and are therefore a potential source of assay-specific confounding artifacts.

Best Practices in Compound Management for Preserving Compound Integrity and Accurately Providing Samples for Assays

Preserving the integrity of the compound collection and providing high-quality materials for drug discovery in an efficient and cost-effective manner are 2 major challenges faced by compound management (CM) at Bristol-Myers Squibb (BMS). The demands on CM include delivering hundreds of thousands of compounds a year to a variety of operations. These operations range from single-compound requests to hit identification support and just-in-time assay plate provision for lead optimization. Support needs for these processes consist of the ability to rapidly provide compounds as solids or solutions in a variety of formats, establishing proper long- and short-term storage conditions and creating appropriate methods for handling concentrated, potent compounds for delivery to sensitive biological assays. A series of experiments evaluating the effects of processing compounds with volatile solvents, storage conditions that can induce freeze/thaw cycles, and the delivery of compounds were performed. This article presents the results of these experiments and how they affect compound integrity and the accuracy of compound management processes. (Journal of Biomolecular Screening 2009:476-484)

Use of Cryopreserved Transiently Transfected Cells in High-Throughput Pregnane X Receptor Transactivation Assay

Cryopreserved, transiently transfected HepG2 cells were compared to freshly transfected HepG2 cells for use in a pregnane X receptor (PXR) transactivation assay. Assay performance was similar for both cell preparations; however, cryopreserved cells demonstrated less interassay variation. Validation with drugs of different PXR activation potencies and efficacies demonstrated an excellent correlation (r 2 > 0.95) between cryopreserved and fresh cells. Cryopreservation did not change the effect of known CYP3A4 inducers that have poor cell permeability, indicating that cryopreservation had little effect on membrane permeability. In addition, cryopreserved HepG2 cells did not exhibit enhanced susceptibility to cytotoxic compounds compared to transiently transfected control cells. The use of cryopreserved cells enables this assay to run with enhanced efficiency.

An ultra-HTS process for the identification of small molecule modulators of orphan G-protein-coupled receptors

G-protein-coupled receptors (GPCRs) are the most successful target proteins for drug discovery research to date. More than 150 orphan GPCRs of potential therapeutic interest have been identified for which no activating ligands or biological functions are known. One of the greatest challenges in the pharmaceutical industry is to link these orphan GPCRs with human diseases. Highly automated parallel approaches that integrate ultra-high throughput and focused screening can be used to identify small molecule modulators of orphan GPCRs. These small molecules can then be employed as pharmacological tools to explore the function of orphan receptors in models of human disease. In this review, we describe methods that utilize powerful ultra-high-throughput screening technologies to identify surrogate ligands of orphan GPCRs.

Discovery, synthesis, and molecular pharmacology of selective positive allosteric modulators of the δ-opioid receptor

Allosteric modulators of G protein-coupled receptors (GPCRs) have a number of potential advantages compared to agonists or antagonists that bind to the orthosteric site of the receptor. These include the potential for receptor selectivity, maintenance of the temporal and spatial fidelity of signaling in vivo, the ceiling effect of the allosteric cooperativity which may prevent overdose issues, and engendering bias by differentially modulating distinct signaling pathways. Here we describe the discovery, synthesis, and molecular pharmacology of δ-opioid receptor-selective positive allosteric modulators (δ PAMs). These δ PAMs increase the affinity and/or efficacy of the orthosteric agonists leu-enkephalin, SNC80 and TAN67, as measured by receptor binding, G protein activation, β-arrestin recruitment, adenylyl cyclase inhibition, and extracellular signal-regulated kinases (ERK) activation. As such, these compounds are useful pharmacological tools to probe the molecular pharmacology of the δ receptor and to explore the therapeutic potential of δ PAMs in diseases such as chronic pain and depression.