Evaluating the ligandability of a protein target is a key component when defining hit-finding strategies or when prioritize among drug targets. Computational as well as biophysical approaches based on nuclear magnetic resonance (NMR) fragment screening are powerful approaches but suffer from specific constraints that limit their usage. Here, we demonstrate the applicability of high-throughput thermal scanning (HTTS) as a simple and generic biophysical fragment screening method to reproduce assessments from NMR-based screening. By applying this method to a large set of proteins we can furthermore show that the assessment is predictive of the success of high-throughput screening (HTS). The few divergences for targets of low ligandability originate from the sensitivity differences of the orthogonal biophysical methods. We thus applied a new strategy making use of modulations in the solvent structure to improve assay sensitivity. This novel approach enables improved ligandability assessments in accordance with NMR-based assessments and more importantly positions the methodology as a valuable option for biophysical fragment screening.
The
MEK1 kinase plays a critical role in key cellular processes,
and as such, its dysfunction is strongly linked to several human diseases,
particularly cancer. MEK1 has consequently received considerable attention
as a drug target, and a significant number of small-molecule inhibitors
of this kinase have been reported. The majority of these inhibitors
target an allosteric pocket proximal to the ATP binding site which
has proven to be highly druggable, with four allosteric MEK1 inhibitors
approved to date. Despite the significant attention that the MEK1
allosteric site has received, chemotypes which have been shown structurally
to bind to this site are limited. With the aim of discovering novel
allosteric MEK1 inhibitors using a fragment-based approach, we report
here a screening method which resulted in the discovery of multiple
allosteric MEK1 binders, one series of which was optimized to sub-μM
affinity for MEK1 with promising physicochemical and ADMET properties.
Two structurally distinct series of novel, MAPK-activated kinase-2 prevention of activation inhibitors have been discovered by high throughput screening. Preliminary structure-activity relationship (SAR) studies revealed substructural features that influence the selective inhibition of the activation by p38α of the downstream kinase MK2 in preference to an alternative substrate, MSK1. Enzyme kinetics, surface plasmon resonance (SPR), 2D protein NMR, and X-ray crystallography were used to determine the binding mode and the molecular mechanism of action. The compounds bind competitively to the ATP binding site of p38α but unexpectedly with higher affinity in the p38α-MK2 complex compared with p38α alone. This observation is hypothesized to be the origin of the substrate selectivity. The two lead series identified are suitable for further investigation for their potential to treat chronic inflammatory diseases with improved tolerability over previously studied p38α inhibitors.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.