Figures & data
Figure 1. Structures of Alda-1, activator (1) and inhibitors (2) and (3) and their EC50 values in the DOPAL inactivation assay. Compounds (1–3) were among the 21 hits identified in the virtual screening.
Figure 2. Generic workflow from PD tissue to the discovery of small molecule modulators of ALDH1. (a) Expression profiling of PD tissues using the Genome-Wide SpliceArray™ led to identification of ALDH1A1 down-regulation together with published literature and pathway analysis formed the basis for the ALDH1 activation hypothesis. Values in parenthesis represent the number of each probe type monitoring the gene transcript deregulation. For the letter code definition of each probe type see supplementary material. (b) Virtual screening workflow from structure to hits. A hit from the virtual screening was validated as an activator in vitro and could serve as starting point for hit to lead medicinal chemistry optimization program.
Figure 3. Recombinant ALDH1A1 is rapidly inactivated by DOPAL. rALDH1A1 was preincubated with different concentrations of DOPAL (shown in mM) for 20 min and residual activity was followed after adding an excess of NAD+ and propanal. The residual activity was measured 12 times with an interval of 2 min between reads. Preincubation with DOPAL led to a concentration-dependent inactivation of rALDH1A1 with complete inactivation observed at 0.05 mM (black squares)and 0.10 mM (black circles) when compared to the solvent preincubated enzyme (open triangles).
Figure 4 (a). Alda-1 activates rALDH1A1 in the DOPAL inactivation assay with an EC50 of 45 µM. Percent activity was calculated as explained in Methods section. (b) Active hits from the virtual screening in the DOPAL inactivation assay. Compound 1 activated rALDH1A1 in this assay with an activation EC50 = 79 µM whereas, compounds 2 and 3 inhibited rALDH1A1 with inhibition EC50 values of 40 µM and 7 µM, respectively. Percentage activity was calculated as explained in the methods section.
