Discovery of a series of aromatic lactones as ALDH1/2-directed inhibitors

Abstract

In humans, the aldehyde dehydrogenase superfamily consists of 19 isoenzymes which mostly catalyze the NAD(P)(+)-dependent oxidation of aldehydes. Many of these isoenzymes have overlapping substrate specificities and therefore their potential physiological functions may overlap. Thus the development of new isoenzyme-selective probes would be able to better delineate the function of a single isoenzyme and its individual contribution to the metabolism of a particular substrate. This specific study was designed to find a novel modulator of ALDH2, a mitochondrial ALDH isoenzyme most well-known for its role in acetaldehyde oxidation. 53 compounds were initially identified to modulate the activity of ALDH2 by a high-throughput esterase screen from a library of 63,000 compounds. Of these initial 53 compounds, 12 were found to also modulate the oxidation of propionaldehyde by ALDH2. Single concentration measurements at 10μM compound were performed using ALDH1A1, ALDH1A2, ALDH1A3, ALDH2, ALDH1B1, ALDH3A1, ALDH4A1, and/or ALDH5A1 to determine the selectivity of these 12 compounds toward ALDH2. Four of the twelve compounds shared an aromatic lactone structure and were found to be potent inhibitors of the ALDH1/2 isoenzymes, but have no inhibitory effect on ALDH3A1, ALDH4A1 or ALDH5A1. Two of the aromatic lactones show selectivity within the ALDH1/2 class, and one appears to be selective for ALDH2 compared to all other isoenzymes tested.

Keywords: Aldehyde dehydrogenase; High-throughput screening; Inhibition.

Figure 1

A) Primary screen results for the 53 lead compounds. The esterase activity of ALDH2 with 10 μM compound present was compared to the activity of ALDH2 in the absence of compound (- control). Selection criteria was either >130% activity or <65% activity. The positive control was 100 μM daidzin. B) Effect of the 53 lead compounds on the dehydrogenase activity of ALDH2. The oxidation of propionaldehyde by ALDH2 with 10 μM compound present was compared to the activity of ALDH2 in the absence of compound. Value is an average of 2 trials.

Figure 2

A) Structures of the aromatic lactones. B) Lineweaver-Burk representation of the fit to the tight binding mixed-type non-competitive inhibition equation for 2P4 versus varying propionaldehyde concentrations with ALDH2. K_i, K_m, and α values are presented with standard error. The plot is a representative single experiment from one of four independent experiments. C) % Dehydrogenase activity for ALDH2, ALDH1B1, ALDH1A1, ALDH1A2, ALDH1A3, ALDH3A1, ALDH4A1, and ALDH5A1 in the presence of 10 μM compound. Value is the average of at least three independent trials (n≥3) with standard error, except for the measurement of ALDH1B1 activity with 2P4 (n=2).