G protein-coupled receptor (GPCR) kinases (GRKs) are key regulators of GPCR signaling cascades. GRKs phosphorylate activated GPCRs and initiate the recruitment of arrestins which promotes GPCR desensitization, endocytosis and degradation. Specific GRK family members have been proposed as drug targets for various disease states, most notably GRK2 overexpression being strongly implicated in the progression of heart failure. Using two novel X-ray crystal structures of GRK5 bound to small-molecules (AMP-PNP and sangivamycin), virtual screening was performed to identify small molecule inhibitors with improved GRK5 affinity and selectivity. All virtual screening was performed using a novel molecular docking algorithm with a cheapest-path side chain optimization. All virtual screening compounds were experimentally characterized using a Surface Plasmon Resonance (SPR) biophysical binding assay for GRK5 and GRK2 providing an experimental assessment of the selectivity of compounds between GRK2/GRK5. Virtual screening hits confirmed by SPR are additionally characterized using differential scanning fluorescence thermal shift assays and an <i>in vitro</i> kinase activity assay using [γ<sup>32</sup>P] ATP. Overall virtual screening results are presented as well as data for three structural hit classes (pyrrolo-pyrimidines, quinolin-2-ones, and indole-2-ones). Detailed computational modeling and virtual screening was utilized to select derivatives of these structural classes specifically aimed at interrogating the structural determinants of selectivity for GRK2 and GRK5.

Virtual screening and biophysical characterization of GRK2 and GRK5 inhibitors

251st National Meeting (2016)

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