Design, Synthesis, and Study of Protective Activity Against Stroke for Novel Water-Soluble Aldehyde Dehydrogenase 2 Activators

Abstract

Stroke poses a serious threat to human health, while there are very few drugs that can directly alleviate ischemia/reperfusion injury and improve the prognosis. Studies have shown that small-molecule activators of aldehyde dehydrogenase 2 (ALDH2) have the potential to become novel therapeutic drugs for ischemic stroke. In this study, through the systematic structural optimization of novel N-benzylaniline-based ALDH2 activators obtained from our previous virtual screening, ALDH2 activators with improved water solubility and activity were obtained. Among them, compound D10 exhibits the best activity, with a maximum activation fold reaching 114% relative to Alda-1. And the water solubility of its hydrochloride salt D27 was increased by more than 200-fold. The intravenous injection of this compound can significantly reduce the infarct area in the rat model of cerebral infarction compared with the model group. This study lays a good foundation for the future research on ALDH2 activators used in the treatment of stroke.

Keywords: activators; aldehyde dehydrogenase 2; cerebral infarction; stroke; water-soluble.

Figure 1

(a) Structures of representative activators; and (b) topological structure of ALDH2. The protein structure in the figure is shown using a ribbon model, with the two subunits represented in pink and red, respectively. Three regions of each subunit are labeled. The mutated amino acids in the ALDH2 variant are shown using a yellow CPK model, and the small-molecule activators are demonstrated using a purple stick model.

Figure 2

(a) Binding mode of N-benzylaniline-based activator C6 with ALDH2. The ALDH2 protein structure template is derived from the crystal structure (PDB ID: 3INJ). The protein is shown as a gray ribbon and a solvent-accessible surface, and the residues important for binding are shown as gray stick models. The activator is shown as a purple stick model, and the hydrogen bonds are shown as blue dashed lines. (b) The design of novel N-benzylaniline-based ALDH2 activators.

Figure 3

Binding mode of N-benzylaniline-based activator D10 with ALDH2 shown as solvent-accessible surface models (a) or gray ribbon models (b). The ALDH2 protein structure template is derived from the crystal structure (PDB ID: 3INJ). The residues important for binding are shown as gray stick models. The activators are shown as purple stick models, and the hydrogen bonds are shown as blue dashed lines.

Scheme 1

Four synthetic routes I–IV of novel N-benzylaniline-based ALDH2 activators. Reagents and conditions: (a) DMSO, K₂CO₃, 80 °C, reflux, 1–4.5 h, 79–99%; (b) DMAP, EDCI, DCM, r. t. 1.5–6 h, 67–95%; (c) EtOH, NH₄Cl, Fe, H₂O, 80 °C, reflux, 1–3 h, 71–97%; (d) DCE, NaBH(OAc)₃, 80 °C reflux, r. t., 2–12 h, 59–95%; (e) DCM, TFA, r. t., 0.5–2.5 h, 73–90%; (f) NaOH, MeOH, THF, H₂O, 1–2 h, 91%; (g) conc-HCl, EA, 1.5 h, r. t., 69%; (h) HATU, DIPEA, DMF, 80 °C, 12 h, 42.8%; (i) PdXPhosG₂, Pd/C, K₃PO₄, Dioxane/H₂O (4:1), 80 °C, 4 h; then MeOH, NH₄COOH, r. t. to 60 °C, 16 h, 75%; and (j) Pd(dppf)Cl₂, K₂CO₃, Dioxane/H₂O (3:1), 100 °C, 12 h, ~70%.

Figure 4

(a) Protective effect of D27 against cerebral infarction in a middle cerebral artery occlusion (MCAO) rat model. The infarct area in MCAO rats is shown as mean ± SD, * p < 0.05, n = 6. (b) Representative brain tissue sections of rats in the D27 group and the model group. The ischemic infarct region is white.