Principles of Small Molecule Library Construction

Small-molecule libraries usually comprise approx 106 organic compounds (molecular weights < 600 amu) that have been prepared by combinatorial and/or traditional medicinal chemistry techniques (classic synthesis). Because both traditional chemistry and combinatorial chemistry offer distinct advantages, they frequently are combined. Libraries also may contain natural product extracts that are isolated from native environmental sources.

Using traditional medicinal chemistry, one molecule at a time is synthesized through a series of solution-phase organic reactions. The advantages of this method include easy isolation and identification of individual molecules, creativity of molecule design, and ability to synthesize complex molecules that may be unavailable from other sources. The major disadvantage of using this method to construct small-molecule libraries is the extensive amount of time required.

In combinatorial chemistry, large numbers of structurally diverse molecules are produced by synthesizing all possible combinations from a set of smaller chemical entities (10). Combinatorial chemical reactions are usually performed on solid phase, in which part of the target molecule is covalently attached to an insoluble support or resin. Then, split synthesis (see below) is used to generate the small-molecule library. Solid-supported chemistry was first optimized in peptide and oligonucleotide synthesis and subsequently applied to small-molecule synthesis. One advantage of generating combinatorial arrays in solid phase is that compounds are easily isolated by washing away reagents from the solid supports. Furthermore, reactions can be driven to completion using excess reagents. Another advantage of solid-phase combinatorial chemistry is that nucleotide, chemical, or radio-encoded tags can be attached to the solid support, which simplifies compound identification (11).

Combinatorial chemistry can also be performed in solution using parallel array synthesis (12). Dynamic combinatorial chemistry, in which the "building blocks" are assembled through reversible chemical reactions in the presence of the target ligand or receptor, is a recent method of preparing small-molecule libraries and offers a new dimension for combinatorial chemistry (10,13).

2.3.1. Split Synthesis

The split synthesis strategy generates libraries of equimolar mixtures of compounds by using repetitive "split and mix" operations (14). A quantity of polymer resin is first split into equal portions and placed into separate reaction vessels. Different chemical entities are attached to each portion of resin, the individual portions are recombined, and a second set of chemical entities is added. After mixing thoroughly, the polymer resin is reapportioned into the requisite number of reaction vessels (11). This procedure allows all the possible combinations of the two sets of chemical entities to be incorporated. The split, react, and mix operations are repeated for each new set of chemical entities added. The total number of compounds generated by this method is obtained by multiplying together the number of chemical entities that are used in each synthesis step.

By incorporating automated systems into the split synthesis process, the time involved in executing the repetitive split and mix operations is greatly reduced (15-17). The disadvantage of split synthesis is that pools (mixtures of compounds) are created; thus, deconvolution (i.e., identifying individual compounds from the mixtures) of the synthesized library is necessary. Fortunately, many solutions to the deconvolution problem have been developed, such as the positional scanning approach (18), the orthogonal library method (19), structural determination by automated analytical methods (20,21), and various encoding strategies (22-24).

2.3.2. Parallel Synthesis

In contrast to the mixtures of compounds that are produced by the split and mix procedure, individual compounds are generated through parallel synthesis (25). In this technique, many compounds are synthesized in parallel in spatially separate reaction vessels. The advantage of this method is that the identity of each compound in a particular location is known and can be confirmed directly. Furthermore, when the library is assayed, individual compounds that demonstrate the desired activity are easily isolated.

Many automated systems for performing parallel synthesis have been reported (26). The first system was introduced by Geyson and coworkers for the mapping of peptide epitopes (27) and later applied to the synthesis of small molecules. Subsequently, many other systems designed exclusively for small molecules were developed (28,29).

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