Combinatorial Chemistry: A New Approch for Drug Discovery
Parimal M Prajapati*, Yatri Shah, DJ Sen and CN Patel
ABSTRACT:
Combinatorial chemistry is a technology for
creating a multitude of different compounds by reacting different combinations
of interchangeable chemical "building blocks." The compounds are then
screened for their ability to carry out a specified function, most commonly to
act as drugs to treat a disease. Combinatorial chemistry allows the rapid
synthesis and testing of many related compounds, greatly speeding the pace of
drug discovery. Combinatorial chemistry is a method for reacting a small number
of chemicals to produce simultaneously a very large number of compounds, called
libraries, which are screened to identify useful products such as drug
candidates.
combinatorial chemistry can be explained simply, its application can take a
variety of forms, each requiring a complex interplay of classical organic
synthesis techniques, rational drug design strategies, robotics, and scientific
information management.
Combinatorial technologies offer significant advances over traditional
scientific research methodologies. In particular, their high-speed approach
promises faster results at considerably lower costs than conventional
techniques. combinatorial chemistry libraries requires
an application that understands the science behind combinatorial chemistry
while managing the chemical and biological data generated by combinatorial
chemistry programs.
INTRODUCTION:
Combinatorial chemistry is one of the important new methodologies developed by academics and researchers in the pharmaceutical, agrochemical, and biotechnology industries to reduce the time and costs associated with producing effective, marketable, and competitive new drugs. Is an approach that provides efficient synthesis of a large collection of molecules. Screening of libraries of related compounds to isolate the molecule of desirable property. Used in both academia and industries to generate huge libraries of compounds that have important biological properties. Combinatorial technologies offer significant advances over traditional scientific research methodologies. In particular, their high-speed approach promises faster results at considerably lower costs than conventional techniques1.
FIGURE 1. COMBINATORIAL CHEMISTRY
combinatorial chemistry to create large populations of molecules, or libraries, that can be screened efficiently . By producing larger, more diverse compound libraries, companies increase the probability that they will find novel compounds of significant therapeutic and commercial value. The field represents a convergence of chemistry and biology, made possible by fundamental advances in miniaturization, robotics, and receptor development. And not surprisingly, it has also captured the attention of every major player in the pharmaceutical, biotechnology, and agrochemical arena. While combinatorial chemistry can be explained simply, its application can take a variety of forms, each requiring a complex interplay of classical organic synthesis techniques, rational drug design strategies, robotics, and scientific information management2. This article will provide a basic overview of existing approaches to combinatorial chemistry, and will outline some of the unique information management problems that it generates.
COMBINATORIAL CHEMISTRY LIBRARY:
It requires an application that understands the science behind combinatorial chemistry while managing the chemical and biological data generated by combinatorial chemistry programs. To meet these ends, MDL plans to release Project Library, a complete, ready-to-use desktop software application that supports the multiple combinatorial chemistry research methods in use today, including functions such as:
· Storage of both oligomeric and non-oligomeric structures.
· Tracking of mixture and discrete compound libraries.
· Elucidation of mixtures or discrete compounds from a library derived from any of the active identification strategies in use today3.
FIGURE 2. COMBINATORIAL CHEMISTRY PROGRAMS
In addition, information concerning the components or building blocks of the library must be processed, stored, and tracked. And because very large numbers of novel structures have to be considered when designing new combinatorial libraries, researchers must be able to enumerate experimental or virtual libraries (groups of either subgeneric or fully specified structures derived from a single generic structure) for investigation and planning. MDL's Project Library is an application that not only helps researchers to manage combinatorial libraries, their building blocks, and their associated data at the project level, it also allows them to plan and refine combinatorial libraries by making it easy to build, store, and export "virtual" libraries4. Combinatorial synthesis can be processed to form building blocks. Once the building blocks are saved in the database, they can easily be incorporated into a generic structure representing a library. The building blocks can be organized by compound class, and data such as name or encoding information can be associated with individual building blocks and is automatically incorporated into the generic structure5.
FIGURE 3. COMBINATORIAL SYNTHESIS
TOOLS FOR COMBINATORIAL CHEMISTRY:
Screening of libraries of related compounds to isolate the molecule of desirable property. It is also Used in both academia and industries to generate huge libraries of compounds that have important biological properties
· Solid-phase synthesis
· Resins
· Reagents (Monomers)
· Linkers
· Screening methods
Solid Phase synthesis concept, first developed for biopolymer, has spread in every field where organic synthesis is involved. Many laboratories and companies focused on the development of technologies and chemistry suitable to SPS. This resulted in the spectacular outburst of combinatorial chemistry, which profoundly changed the approach for new drugs, new catalyst or new natural discovery. The use of solid support for organic synthesis relies on three interconnected requirements:
FIGURE 4. SOLID PHASE SYNTHESIS
1) A cross linked, insoluble polymeric material that is inert to the condition of synthesis;
2) Some means of linking the substrate to this solid phase that permits
selective cleavage of some or all of the product from the solid support during
synthesis for analysis of the extent of reaction(s), and ultimately to give the
final product of interest;
3) A chemical protection strategy to allow selective protection and deprotection of reactive groups6.
The direction of synthesis is opposite to that used in the cell. The intended carboxy terminal amino acid is anchored to a solid support. Then, the next amino acid is coupled to the first one. In order to prevent further chain growth at this point, the amino acid, which is added, has its amino group blocked. After the coupling step, the block is removed from the primary amino group and the coupling reaction is repeated with the next amino acid. The process continues until the peptide or protein is completed. Then, the molecule is cleaved from the solid support and any groups protecting amino acid side chains are removed. Finally, the peptide or protein is purified to remove partial products and products containing errors 7-8.
BECKMAN 96 DEEP-WELL TITER PLATE
Use multiple parallel synthesis in a matrix format - 20 reagents with 2 reactions gives 96 products.
FIGURE 5. TITER PLATE
Polyacrylamide polymers for peptide synthesis as it was expected that these polymers would more closely mimic the properties of the peptide chains themselves and have greatly improved solvation properties in polar, aprotic solvents (e.g. DMF, or N-methyl pyrrolidinone)9.
FIGURE 6(A). POLYAMIDE CHAIN
The protecting group finally chosen was the fluorenylmethoxycarbonyl (Fmoc) which can be removed by base (usually piperidine).
(a) The simple case where no by-products are generated. (b) A more complex case where excess coupling components have been used or by-product removal is needed.
In spite of some successes arising from the use of combinatorial chemistry it has also been responsible for a certain dumbing down of chemistry by its tendency to rely on straightforward and reliable reactions with accepted concomitant compromises in both yield and complexity of the structures that are synthesized 10-13. This is no longer acceptable and we must make compounds designed for purpose rather than simply making compounds because we can. In a research laboratory for example, there might be a requirement for greater diversity in the molecules that are made and high attrition rates may be acceptable. In the process environment however, greater reaction versatility and reliability are priorities thus lower attrition rates are essential.
The group that joins the substrate to the resin bead is an essential part of solid phase synthesis. The linker is a specialised protecting group, in that much of the time, the linker will tie up a functional group, only for it to reappear at the end of the synthesis. The linker must not be affected by the chemistry used to modify or extend the attached compound. And finally the cleavage step should proceed readily and in a good yield. The best linker must allow attachment and cleavage in quantitative yield14.
The first linking group used for peptide synthesis bears the name of the father of solid phase synthesis. Resin is cross-linked polystyrene functionalised with a chloromethyl group. The carbonyl group is attached by the nucleophilic displacement of the chloride with a cesium carboxylate salt in DMF. Cleavage to regenerate the carboxylic acid is usually achieved by hydrogen fluoride15.
The second class of linker used for carboxylic acid is the linker. This linker is generally attached to cross-linked polystyrene, TentaGel and polyacrylamide to form Wang resin. It was designed for the synthesis of peptide carboxylic acids using the Fmoc-protection strategy, and due to the activated benzyl alcohol design, the carboxylic acid product can be cleaved with TFA.
A more acid-labile form of the resin has been developed. The resin has the same structure as the linker but with the addition of a methoxy group to stabilise the carbonium ion formed during acid catalysed cleavage 16-17.
HIGH THROUGHPUT SCREENING AND COMBINATORIAL CHEMISTRY:
High-throughput screening (HTS) is an approach to drug discovery that has gained widespread popularity over the last three or four years. HTS is the process of assaying a large number of potential effectors of biological activity against targets (a biological event). The methods of HTS are applied to the screening of combinatorial chemistry, genomics, protein, and peptide libraries. The goal of HTS is to accelerate drug discovery by screening large libraries often composed of hundreds of thousands of compounds (drug candidates) at a rate that may exceed 20,000 compounds per week. This paper will focus on assay adaptations, robotic equipment, and implementation strategies that allow HTS programs to be successful.Traditional lead discovery approaches often rely on the synthesis of building blocks, their transformation in larger combinatorial libraries followed by the identification of compounds showing desired activity via high-throughput screening18.
In the last decade, a handful of studies have been reported in which the target protein is involved in the synthesis of its own inhibitor. These approaches, also termed target-guided synthesis (TGS), are relatively unexplored and poorly understood despite the reported success at this time. Our laboratory proposes to create a shortcut in drug discovery processes by developing TGS techniques combining synthesis and screening of low-molecular weight compound libraries in the very same step. Reactive building blocks are directly assembled to bigger, multivalent compounds in the presence of the target protein. Mainly for entropic reasons, multivalent compounds display much higher affinity to their biological targets than the individual monovalent building blocks. Thus, even fragments with only modest affinity to a targets individual binding pocket can ultimately provide compounds with strong binding properties when coupled together in the correct way 19-20.
FIGURE 8. HIGH THROUGHPUT SCREENING TECHNIQUE
In the last decade, a handful of studies have been reported in which the target protein is involved in the synthesis of its own inhibitor. These approaches, also termed target-guided synthesis (TGS), are relatively unexplored and poorly understood despite the reported success at this time. Our laboratory proposes to create a shortcut in drug discovery processes by developing TGS techniques combining synthesis and screening of low-molecular weight compound libraries in the very same step. Reactive building blocks are directly assembled to bigger, multivalent compounds in the presence of the target protein. Mainly for entropic reasons, multivalent compounds display much higher affinity to their biological targets than the individual monovalent building blocks. Thus, even fragments with only modest affinity to a targets individual binding pocket can ultimately provide compounds with strong binding properties when coupled together in the correct way 19-20.
NEW APPROCHES IN COMBINATORIAL CHEMISTRY
• Use biologically relevant building blocks
• Use branching networks of reactions
• Produce libraries of natural-product-like compounds
• Make all possible combinations of both core skeletal structures and peripheral groups
• Dynamic Combichem (DCC)
• Synthetic receptors that bind tightly to small molecules
• Uses equilibrium forces to amplify compounds that bind well to targets
As with traditional drug design, combinatorial chemistry relies on organic synthesis methodologies. The difference is the scope--instead of synthesizing a single compound, combinatorial chemistry exploits automation and miniaturization to synthesize large libraries of compounds. But because large libraries do not produce active compounds independently, scientists also need a straightforward way to find the active components within these enormous populations 21-22. Thus, combinatorial
organic synthesis (COS) is not random, but systematic and repetitive, using sets of chemical "building blocks" to form a diverse set of molecular entities. Scientists have developed several different COS strategies, each with the same basic philosophy--stop shooting in the dark and instead, find ways to determine active compounds within populations, either spatially, through chemical encoding, or by systematic, successive synthesis and biological evaluation (deconvolution). There are three common approaches to COS. During arrayed, spatially addressable synthesis, building blocks are reacted systematically in individual reaction wells or positions to form separated "discrete molecules." Active compounds are identified by their location on the grid. This method has been applied in scale (as in the Parke-Davis Pharmaceutical DIVERSOMER technique), as well as in miniature (as in the Affymax VLSIPS technique). The second technique, known as encoded mixture synthesis, uses nucleotide, peptide, or other types of more inert chemical tags to identify each compound. During deconvolution, the third approach, a series of compound mixtures is synthesized combinatorially, each time fixing some specific structural feature. Each mixture is assayed as a mixture and the most active combination is pursued. Further rounds systematically fix other structural features until a manageable number of discrete structures can be synthesized and screened. Scientists working with peptides, for example, can use deconvolution to optimize, or locate, the most active peptide sequence from millions of possibilities. You could say that combinatorial chemistry is a technologically advanced way of finding a needle in a haystack. The whole idea is to remove the guesswork and instead, to create and test as many compounds or mixtures as possible--logically and systematically--to obtain a viable set of active leads.
APPLICATION OF COMBINATORIAL CHEMISTRY
• Transition-state analog HIV protease inhibitors.
Extensive efforts toward the rational design of aspartyl protease inhibitors such as renin and HIV have led to the discovery of several transition-states analog mimics. These templates can serve as the central unit around which molecular diversity can be generated by application of appropriate chemistries. Recently, solid phase synthesis of hydroxyethylamine and 1,2-diol transition-state pharmacophore units and their utility for synthesis of HIV protease inhibitors have been reported by two different groups. The first instance, bifunctional linker are used by Wang to serve the dual purpose of protecting the hydroxyl group of these BBs and providing point for attachment on solid support23.
Thus, one linker possesses a vinyl ether group at one end and a free carboxylate group at the other. The vinyl ether moiety is reacted with diamino alcohol BB 1 under acid-catalysed conditions to form an acetal protecting group and the carboxylic acid group is used for ester-type linkage to the solid support24. The other linker possesses a methyl ketone and carboxylic groups at the two ends, with the ketone group forming a ketal with diol 3. Resulting intermediates 2 and 4 are now well suited for a bi-directional solid phase synthesis strategy for preparing C2 symmetric HIV protease inhibitors. The two terminal amino groups of 2 and 4 are deprotected and reacted with a variety of carboxylic acid, sulfonyl chlorides, isocyanates, and chloroformates to extend the core unit in both directions and generate a wide variety of aspartyl protease inhibitors. The authors claim that a library of 300 discrete analogs was prepared and screened against HIV protease to identify several potent inhibitors 25-26.
• Benefits to the Pharmaceutical Industry
Provides a stimulus for robot-controlled and immobilization strategies that allow high-throughput and multiple parallel approaches to drug discovery
• Speed
• Economics
CONCLUSION:
As with traditional drug design, combinatorial chemistry relies on organic synthesis methodologies. The difference is the scope--instead of synthesizing a single compound, combinatorial chemistry exploits automation and miniaturization to synthesize large libraries of compounds.
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Received on 30.12.2009 Modified on 02.02.2010
Accepted on 17.03.2010 © AJRC All right reserved
Asian J. Research Chem. 3(2): April- June 2010; Page 249-254