A family of phosphodiesterase inhibitors discovered by cocrystallography and scaffold-based drug design

Abstract

Cyclic nucleotide phosphodiesterases (PDEs) comprise a large family of enzymes that regulate a variety of cellular processes. We describe a family of potent PDE4 inhibitors discovered using an efficient method for scaffold-based drug design. This method involves an iterative approach starting with low-affinity screening of compounds followed by high-throughput cocrystallography to reveal the molecular basis underlying the activity of the newly identified compounds. Through detailed structural analysis of the interaction of the initially discovered pyrazole carboxylic ester scaffold with PDE4D using X-ray crystallography, we identified three sites of chemical substitution and designed small selective libraries of scaffold derivatives with modifications at these sites. A 4,000-fold increase in the potency of this PDE4 inhibitor was achieved after only two rounds of chemical synthesis and the structural analysis of seven pyrazole derivatives bound to PDE4B or PDE4D, revealing the robustness of this approach for identifying new inhibitors that can be further developed into drug candidates.

Scaffold-based discovery of indeglitazar, a PPAR pan-active anti-diabetic agent

“…The target-naive screening library was designed by using compounds from a relatively narrow and low molecular weight range (150-350D), selected for diversity at both the putative ''scaffold'' core and at the whole molecule level (6). Compounds capable of modulating receptor signaling were identified based on a minimal signal for significant response relative to the background by using proximity-based co-activator recruitment assays with the ligand-binding domain (LBD) for each of the 3 PPARs and a compound concentration of 100 M. Candidates with weak activity against all 3 receptors, against any 2 of the pair-wise combinations, or containing chemotypes related to those in the first 2 groups but active against only one of the receptors, were selected for co-crystallization with at least 1 of the 3 receptors.…”

“…We have recently described an approach where low-molecular-weight (150-350Da) compounds, weakly active in an initial biochemical screen against a panel of structurally related targets, are subject to a second filtering process based on high-throughput co-crystallography (6)(7)(8). Of the compounds in which binding orientations in the active sites of target molecules can be unambiguously determined by X-ray diffraction analysis, we selected those offering the most efficient access to chemistry as the starting points (or scaffolds) for discovery programs.…”

Structural characterization of autoinhibited c-Met kinase produced by coexpression in bacteria with phosphatase

“…Conservation in the active site residues within the protein kinase gene family makes the development of selective kinase inhibitors challenging. Structural biology offers valuable information useful in the design of new inhibitors (2), but a limitation in its application to kinases can often be the inability to produce highly purified proteins in amounts suitable for cocrystallography. Inhibitor binding sometimes can be sensitive to the specific conformation state of a kinase (3), or to changes in the kinase sequence caused by mutations, such as those occurring during cancer progression (4)(5)(6)(7).…”

Novel inhibitors of anthrax edema factor

“…For the former, biophysical methods such as X-ray crystallography 26 , NMR spectroscopy 25, 27, 30 , and surface plasmon resonance 38 have been used to design and synthesize high-affinity ligands, based on fragments with good binding properties 25,31,32,[35][36][37] . Some compounds identified using fragment-based approaches have entered clinical trials 36 , and fragment based discovery can identify quality leads for targets where HTS has not succeeded 31, 32, 39 .…”