A biomimetic polyketide-inspired approach to small-molecule ligand discovery - PubMed (original) (raw)

A biomimetic polyketide-inspired approach to small-molecule ligand discovery

Claudio Aquino et al. Nat Chem. 2011.

Abstract

The discovery of new compounds for the pharmacological manipulation of protein function often embraces the screening of compound collections, and it is widely recognized that natural products offer beneficial characteristics as protein ligands. Much effort has therefore been focused on 'natural product-like' libraries, yet the synthesis and screening of such libraries is often limited by one or more of the following: modest library sizes and structural diversity, conformational heterogeneity and the costs associated with the substantial infrastructure of modern high-throughput screening centres. Here, we describe the design and execution of an approach to this broad problem by merging principles associated with biologically inspired oligomerization and the structure of polyketide-derived natural products. A novel class of chiral and conformationally constrained oligomers is described (termed 'chiral oligomers of pentenoic amides', COPA), which offers compatibility with split-and-pool methods and can be screened en masse in a batch mode. We demonstrate that a COPA library containing 160,000 compounds is a useful source of novel protein ligands by identifying a non-covalent synthetic ligand to the DNA-binding domain of the p53 transcription factor.

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Conflict of interest statement

Competing Interests Statement

The authors declare no competing financial interests.

Figures

Figure 1

Figure 1. Natural and synthetic oligomers, polyketide-derived natural products, and a polyketide-inspired class of chiral and conformationally rigid synthetic oligomer

a, A selection of biological and biopolymer mimetics. b, A selection of polyketide-derived natural products (regions highlighted in red are substituted alkenes that impart conformational rigidification as a result of A1,3 strain). c, General structure of COPAs – chiral oligomers of _N_-substituted 5-amino-2,4-dialkyl-3-pentenoic amides. d, Structural features that lead to the rigidification of COPA oligomers.

Figure 2

Figure 2. Stereochemistry of COPA backbone is anticipated to have a substantial impact on skeletal shape and the disposition of side chains in space

Distribution of conformers found within 1.1 kcal/mol of the lowest energy conformation identified. These calculations were conducted with Spartan-08/MMFF model/Conformer Distribution option/Monte-Carlo algorithm. Colored spheres highlight the relative position of heteroatoms (green) and alkenes (blue). While these molecular mechanics calculations are not interpreted to predict the solution phase structure of these oligomers, the calculations provide a uniform lens through which to observe unique characteristics associated with this new class of synthetic oligomer.

Figure 3

Figure 3. Chemical development of COPA oligomers: From general oligomerization strategy, asymmetric synthesis and library construction

a , The “sub-monomer” style synthesis of peptoids. b, Asymmetric synthesis of 5-chloro-2,4-dimethyl-3-pentenoic acid 1. c, Use of 1 in solution phase oligomerization. d, Panel of monomers used in library synthesis. e, General information regarding resin and linker employed in solid-phase library synthesis. f, General structure of libraries prepared from building blocks depicted in Figure 3d – COPA and peptoid tetramers.

Figure 4

Figure 4. COPA library synthesis, screening, structure elucidation and validation

a , General scheme for on-bead screening of COPA library against DNA binding domain of p53 (p53-DBD, residues 94 to 312) expressed with an epitope tag FLAG. TentaGel beads bound to p53-DBD protein were visualized under fluorescent microscope by treating beads with anti-FLAG primary antibody and anti-IgG secondary antibody conjugated to Quantum dot emitting red fluorescent light at 655 nm. b, Sequence elucidation and identification of a COPA tetramer that binds to the p53-DBD. Sequence of the COPA tetramer was established by analysis of mass spectral data derived from ETD-based fragmentation. c, Fluorescence polarization assay for binding affinity of fluorescein conjugated COPA tetramer (14a) against p53-DBD, carbonic anhydrolase II (CAH II from bovine erythrocyte), platelet activating factor acetyl hydrolase (PAFAHIB3), and bromodomain containing 4 (BRD4) proteins. Error bars show the standard deviation (SD) of the data obtained from three independent experiments. The data analysis and curve fitting were performed with GraphPad Prism 5.0 (GraphPad Software, San Diego, CA) using a non-linear regression method (y=[_x_/(_KD_ + _x_)](_ymax_ − _ymin_) + ymin). A COPA tetramer with the same linker region and different side chains on the amide nitrogens was used as a control oligomer (co). The binding affinity of COPA tetramer to p53-DBD was determined as KD ~ 10μM. See Supporting Information for additional details.

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