Identification of a novel ALDH1A3-selective inhibitor by a chemical probe with unrelated bioactivity: An approach to affinity-based drug target discovery.

The identification of biologically active target compounds and their binding proteins is important in mechanism-of-action studies for drug development. Additionally, the newly discovered binding proteins provide unforeseen ideas for novel drug discovery and for subsequent structural transformation to improve target specificity. Based on the lead and final candidate compounds related to the type 5 phosphodiesterase (PDE5) inhibitor E4021, we designed chemical probes and identified their target proteins by the affinity chromatography approach. Aldehyde dehydrogenase family 1 member A3 (ALDH1A3), currently reported as a cancer stem cell target, was clearly isolated as a binding protein of the lead 'immature' inhibitor probe against PDE5. In the early derivatization to the closely related structure, Compound 5 (ER-001135935) was found to significantly inhibit ALDH1A3 activity. The discovery process of a novel ALDH1A3-selective inhibitor with affinity-based binder identification is described, and the impact of this identification method on novel drug discovery is discussed.

Symmetry-Driven Strategy for the Assembly of the Core Tetracycle of (+)-Ryanodine: Synthetic Utility of a Cobalt-Catalyzed Olefin Oxidation and α-Alkoxy Bridgehead Radical Reaction.

Ryanodine (1) is a potent modulator of intracellular calcium release channels, designated as ryanodine receptors. The exceptionally complex molecular architecture of 1 comprises a highly oxygenated pentacyclic system with eleven contiguous stereogenic centers, which makes it a formidable target for organic synthesis. We identified the embedded C2 -symmetric tricyclic substructure within 1. This specific recognition permitted us to design a concise synthetic route to enantiopure tricycle 9 by utilizing a series of pairwise functionalizations. The four tetrasubstituted carbon centers of 9 were effectively constructed by three key reactions, a dearomatizing Diels-Alder reaction, the kinetic resolution of the obtained racemic 14 through asymmetric methanolysis, and the transannular aldol reaction of the eight-membered diketone 10. A new combination of cobalt-catalyzed hydroperoxidation and NfF-promoted elimination enabled conversion of the hindered olefin of 9 into the corresponding ketone, thus realizing the desymmetrization. Finally, the tetrasubstituted carbon was stereospecifically installed by utilizing the α-alkoxy bridgehead radical to deliver the core tetracycle 7 with the six contiguous tetrasubstituted carbon centers. Consequently, the present work not only accomplishes efficient assembly of four out of the five fused rings of 1, but also develops two new powerful methodologies: two-step ketone formation and bridgehead radical reaction.

Functional expression of full-length TrkA in the prokaryotic host Magnetospirillum magneticum AMB-1 by using a magnetosome display system.

Tropomyosin receptor kinase A (TrkA), a receptor tyrosine kinase, is known to be associated with various diseases. Thus, TrkA has become a major drug-screening target for these diseases. Despite the fact that the production of recombinant proteins by prokaryotic hosts has advantages, such as fast growth and ease of genetic engineering, the efficient production of functional receptor tyrosine kinase by prokaryotic hosts remains a major experimental challenge. Here, we report the functional expression of full-length TrkA on magnetosomes in Magnetospirillum magneticum AMB-1 by using a magnetosome display system. TrkAwas fused with the magnetosome-localized protein Mms13 and expressed on magnetosome surfaces. Recombinant TrkA showed both nerve growth factor (NGF)-binding and autophosphorylation activities. TrkA expressed on magnetosomes has the potential to be used, not only for further functional analysis of TrkA, but also for ligand screening.