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THE DEGREE TO WHICH combinatorial chemistry has become fully integrated into the drug discovery process, as opposed to being an optional add-on that has yet to prove itself, is exemplified by the sophisticated "compound factory" that has been created at GlaxoSmithKline, in Harlow, England. Researchers there have designed a flexible combinatorial chemistry environment with a wide range of automated instruments for library synthesis, analysis, and purification, and they have developed a powerful informatics system to handle the huge volume of combinatorial data generated at the center. At the Cambridge conference, David Hunter of the Discovery Research Chemistry unit at Harlow gave attendees an unusually detailed view of the combinatorial state of the art at a major pharmaceutical company. Maintaining profitability in a highly competitive and tightly regulated environment necessitates that drug companies generate drug leads efficiently, minimize the failure rate of compounds in clinical trials by identifying strong candidates, and move drugs into the marketing pipeline quickly. Combinatorial chemistry is helping GlaxoSmithKline meet these goals, Hunter says. In the past couple of years, the company has become significantly more dependent on lead compounds generated by the combinatorial unit and less dependent on leads from the corporate compound library--the venerable mainstay of drug discovery in big pharma. An early emphasis in GlaxoSmithKline's combinatorial group on the synthesis of very large libraries as mixtures of compounds has matured into a more focused approach on the use of solid- and solution-phase techniques to synthesize smaller libraries of well-characterized individual compounds. Automated synthesizers from Zymark, Hopkinton, Mass., and Bohdan, Vernon Hills, Ill., and the FlexChem system from Robbins Scientific, Sunnyvale, Calif., are used for most of the group's solution-phase chemistry. Synthesizers from Advanced ChemTech, Louisville, Ky., are used for training purposes at the company. COMBI LAB Combinatorial researchers at GlaxoSmithKline's Discovery Research Chemistry unit, in Harlow, England, work at a Tecan Genesis workup station (front) and a Myriad Core System robotic synthesizer. MULTIPLEXED Micromass MUX inlet permits effluents from as many as eight liquid chromatographs to be directed sequentially into an electrospray mass spectrometer for combinatorial library analysis. THE RESEARCHERS also make extensive use of both the Myriad Core System from Mettler-Toledo Myriad, Melbourn, England--a parallel solution- and solid-phase system for automated synthesis in 192 reaction vessels--and Myriad personal synthesizers, which are similar but have only 24 reaction vessels. The unique cap design of the Myriad reactors makes these instruments particularly useful for syntheses involving corrosive, air-sensitive, and moisture-sensitive reagents and reactive intermediates, Hunter says. Hunter says that encoding technology from the Irori unit of Discovery Partners International, San Diego, Calif., is ideal for creating large libraries by split-and-mix synthesis--an approach in which pools of compounds are iteratively mixed, reacted, and divided up to generate large numbers of diverse products very rapidly. In the Irori system, small radio-frequency tags or two-dimensional optical bar codes are placed in or on microreactors, which are then used to synthesize compounds. The tags or bar codes can then be used to track compounds subjected to multiple split-and-mix synthesis steps. The technology combines the efficiency advantages of split-and-mix synthesis with the ability to recover and identify individual products. The methodology is used at GlaxoSmithKline to prepare large arrays of diverse compounds as well as focused libraries designed to interact with specific molecular targets. A recent change at the company is an increased use of microwave heating for library synthesis. GlaxoSmithKline researchers are currently using the Smith Synthesizer from Personal Chemistry, Uppsala, Sweden, which permits precise control of reaction temperature and pressure and has a typical throughput of more than 10 reactions per hour. All library compounds produced at the Harlow center are analyzed for purity by liquid chromatography-mass spectrometry (LC/MS), using the LCT with MUX technology from Micromass, Manchester, England. This instrument connects a time-of-flight mass analyzer with as many as eight LC columns--permitting an eightfold increase in throughput compared with conventional LC/MS. Company researchers also analyze the purity of compounds in focused libraries and representative compounds in large libraries by nuclear magnetic resonance (NMR) spectroscopy using the BEST-NMR system from Bruker, Rheinstetten, Germany--an automated-flow NMR instrument that makes it possible to analyze samples directly from microtiter plates. Synthesis products are weighed utilizing an automated weighing station, the Bohdan Balance Automator. After purity analysis, the results are imported into a custom-designed computer system called RADICAL (the Registration, Analysis & Design Interface for Combinatorial & Array Libraries). RADICAL categorizes products into three groups: those that have passed quality control, those that require further purification, and those that are beyond hope and must be discarded. RADICAL also handles data regarding library design, reagent management, library synthesis, purification, quality control, and product registration in the corporate compound database, and it's used to communicate essential information about compounds to the high-throughput screening group. THERE'S A GROWING interest in avoiding the need for product purification by generating pure compounds in the first place, Hunter notes. This can be done, for example, by carrying out combinatorial syntheses with solid support-bound reagents, which make it possible to recover pure individual compounds by simple filtration and removal of solvent. But for compounds generated in other ways, purification is often necessary. Initial cleanup can involve liquid-liquid extraction (with the Allex system from Mettler-Toledo Myriad) or scavenging of impurities (by ion-exchange-based extraction or covalent bond formation with solid-phase-supported reagents). Subsequent purification is carried out by automated flash chromatography--using the FlashMaster II from Jones Chromatography, Hengoed, Wales, or the Quad3 from Biotage, Charlottesville, Va.--and by high-throughput, preparative high-performance LC (HPLC) using Biotage Parallex purification systems. The FlashMaster II system can purify 10 compounds sequentially, the Quad3 is a parallel 12-column instrument, and the Parallex is a fully automated system with four HPLC columns that run in parallel and can handle 200 to 300 samples per day, Hunter says. Postpurification processing is then carried out with a series of instruments from Biotage; Micromass; Tecan, Männedorf, Switzerland; Zymark; Genevac, Suffolk, England; and Bohdan. In this step, purified compounds are put in bar-coded vials and transferred to the high-throughput screening group for testing. Combinatorial chemistry and high-throughput screening have become an integral part of daily operations in medicinal chemistry groups at GlaxoSmithKline in the past few years, Hunter says. Has the changeover from the traditional one-compound-at-a-time approach been worth it? As a result of the changeover, the productivity of biochemists and medicinal chemists at the company has been enhanced, Hunter says, while the quality of compounds going into the drug development pipeline has been maintained. DYNAMIC COMBICHEM In a preliminary study of binding specificity, Thomas and coworkers found that iminium products of two aldehydes (R5CHO and R8CHO) in a library of eight aldehydes bound with highest affinity to the enzyme -galactosidase. "HIGH-THROUGHPUT synthesis has had a substantial impact on the progress of research programs, with numerous examples of instant SARs, reduced cycle time, and more potent leads," Hunter says. SARs (structure-activity relationships), which measure the influence of compound structural changes on biological activity, can help researchers identify more highly active agents. GlaxoSmithKline's combinatorial program, Hunter says, has also led to "increased serendipity"--the generation of compounds of unexpectedly high potency and the discovery in libraries of leads for unexpected targets. Most importantly, he adds, more than 50% of drug leads identified at the company in 2000 originated from high-throughput chemistry--more than twice as many as in 1999. One attendee asked Hunter when he thought people would see case studies of compounds from high-throughput chemistry that had actually gone into human clinical trials. "It's taking longer than anticipated," Hunter replied. "But eventually the majority of drugs going into clinical trials will originate from high-throughput chemistry." |
| UPDATE | 08.01 |
| COMPANY | GlaxoSmithKline |
| LITERATURE REF. | This data is not available for free |
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