PMC

Select PKD1 inhibitors were evaluated to determine if they were mixed, competitive or non-competitive inhibitors of ATP. Panels A and B, representative Michaelis-Menten and Lineweaver-Burk plots of CID 5086667. This compound was found to be an ATP-competitive inhibitor. Data presented were representative of three independent experiments.

New PKD1 inhibitory chemotypes were identified through HTS activities with twelve inhibitory chemotypes confirming in a radiometric PKD1 kinase assay. In vitro IC₅₀ values ranged from 0.2–6.1 µM. Data are represented as mean ± range or SD of 2–4 independent experiments.

Panel A shows the binding pose of BPKDi (red), a cell-based PKD inhibitor, with a known mechanism of action (i.e., ATP competitive) and ATP (ball-and-stick representation and colored by atom type). Panels B and C depict our two cellular PKD1 small molecule inhibitors and ATP docked to the conserved catalytic domain (Panel B, CID 5389142 (blue); Panel C, CID 2011756). Panel D displays all three cellular PKD1 small molecule inhibitors (CID 5389142-blue, CID 2011756-yellow and BPKDi-red) docked to the conserved ATP binding domain.

Panels A and B, compounds were evaluated initially for PKD1 inhibitory activity at 50 µM in LNCaP cells. Panel C, a representative Western blot shows that four compounds CID 2011756, CID 5389142, CID 1893668 and CID 755673 (control compound), displayed a concentration-dependent inhibition of PMA-induced, endogenous PKD1 activity. Quantification of Western blotting data was determined using densitometry analysis and is graphically represented as mean ± SEM. Data presented are representative of three to four independent experiments. ns, not statistically significant; *, p<0.05; **p<0.01; ***, p<0.001.