Reprogramming Combinatorial Biology for Combinatorial Chemistry

Sean V. Taylor 35.1

Introduction

As the other chapters in this book have nicely illustrated, combinatorial chemistry has made dramatic technical advances since its inception, with significant improvements in the design, synthesis, purification, and evaluation of combinatorial libraries. It is now commonplace for companies focusing on drug discovery to have 0.5-1.5 million compounds arrayed throughout their combinatorial libraries, which can be screened in a matter of weeks or months for activity against any number of biological targets. The potential success of this strategy is underscored by reports from numerous pharmaceutical companies of compounds that were identified or optimized using combinatorial chemistry and that have now entered clinical drug trials. It is quite likely that, over the next 100 years, a significant percentage of pharmaceuticals will be derived in some manner from combinatorial libraries.

If we expand our definition of combinatorial libraries, however, the current importance of the combinatorial approach to drug discovery, as well as to most life processes, is more immediately clear. For instance, eight of the 20 top-selling pharmaceuticals are natural products or derived from natural products [1-3]. It is not a large leap in logic to consider these compounds as already being derived from combinatorial libraries - biological, or perhaps ''evolutionary'' combinatorial libraries. In contrast to combinatorial libraries produced by synthetic means, the combinatorial libraries that give rise to natural products are not produced at the ''compound'' level, but at the genetic level during the process of evolution, through mutation, gene duplication, and interspecies genetic transfer. The evaluation of these biological combinatorial libraries is simply the process of natural selection. Those compounds that provide some value toward the survival of the producing organism will be propagated and further improved upon through additional evolutionary rounds.

*Portions of this article were previously published (Angew. Chem. 113 (2001) 3408-3436; Angew. Chem. Int. Ed. 40 (2001) 3310-3335). [Ref. 100]

Handbook of Combinatorial Chemistry. Drugs, Catalysts, Materials. Vol. 2. Edited by K. C. Nicolaou, R. Hanko, and W. Hartwig Copyright © 2002 WILEY-VCH Verlag GmbH, Weinheim ISBN: 3-527-30509-2

Nature has utilized such ''evolutionary'' libraries over billions of years, but molecular biologists, biochemists, and chemists have only recently learned how to harness this combinatorial power, and for a number of different uses. In this chapter, the various efforts to engineer and apply biological combinatorial libraries for use in enzymology, drug discovery, bioprocessing, and also for addressing fundamental issues in chemistry and biology will be explored. Instead of an exhaustive review, specific examples will be used to highlight various combinatorial aspects of biology. In addition to asking the question ''Where are we now?'', I also hope to ask ''Where are we (or where could we be) going?''

35.2

Combinatorial Biosynthesis: Creation of Novel Small-molecule Natural Products

Many secondary metabolites are composed of polyketides, nonribosomally produced peptides, carbohydrates, or some combination of all three (Fig. 35.1) [4-6]. Polyketides are complex and highly stereogenic compounds that are constructed from acyl-coenzyme A building blocks, often by huge modular enzyme complexes called polyketide synthases (PKSs) [7]. Nonribosomal peptides (NRPs), as the name implies, are amino acid-derived molecules wherein the peptide bonds are not produced by the traditional translational machinery of the ribosome, but instead on large modular enzyme complexes called nonribosomal peptide synthases (NRPSs) [8]. Carbohydrate moieties, very commonly deoxysugars in secondary metabolites, are produced by more ''traditional'' nonmodular biosynthetic pathways [9-11]. Illustrations of the typical biosynthetic pathways are shown in Fig. 35.2.

The relaxed specificity demonstrated by some of the enzymes involved in producing these compounds, as well as the modular nature of some of the systems, have allowed researchers to produce new ''natural'' products by cleverly reengineering the biosynthetic pathways. Such engineering is usually called combinatorial biosynthesis, and is often achieved by substituting (in combinatorial fashion) different non-natural building blocks and the enzymes that can utilize them into a poly-ketide, nonribosomal peptide, or deoxysugar biosynthetic pathway [12-18]. The reprogramming of biosynthetic pathways has the potential to be enormously useful for combinatorial chemistry, since it creates a method for the parallel synthesis of a large number of complex molecules whose conventional organic syntheses are generally too complex to be easily adapted for combinatorial production. Another advantage is that these combinatorial biosynthetic libraries are often closely related to natural products that have already shown some useful activity, so the chances of harvesting pharmaceutical lead compounds from them may be higher than those from a synthetic combinatorial library. Finally, combinatorial biosynthesis could provide a convenient method for creating libraries that could be further manipulated by synthesis, reducing some of the time and cost associated with synthetic combinatorial chemistry.

For all of these reasons, many corporate and academic research groups are working to manipulate biosynthetic pathways for use in combinatorial biosynthesis [19].

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epothilone anticancer

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