Development of Novel Alaninamide Derivatives with Anticonvulsant Activity and Favorable Safety Profiles in Animal Models

Abstract

In our current study, we developed a focused series of original ((benzyloxy)benzyl)propanamide derivatives that demonstrated potent activity across in vivo mouse seizure models, specifically, maximal electroshock (MES) and 6 Hz (32 mA) seizures. Among these derivatives, compound 5 emerged as a lead molecule, exhibiting robust protection following intraperitoneal (i.p.) injection, as follows: ED₅₀ = 48.0 mg/kg in the MES test, ED₅₀ = 45.2 mg/kg in the 6 Hz (32 mA) test, and ED₅₀ = 201.3 mg/kg in the 6 Hz (44 mA) model. Additionally, compound 5 displayed low potential for inducing motor impairment in the rotarod test (TD₅₀ > 300 mg/kg), indicating a potentially favorable therapeutic window. In vitro toxicity assays further supported its promising safety profile. We also attempted to identify a plausible mechanism of action of compound 5 by applying both binding and functional in vitro studies. Overall, the data obtained for this lead molecule justifies the more comprehensive preclinical development of compound 5 as a candidate for a potentially broad-spectrum and safe anticonvulsant.

Keywords: amino acids derivatives; anticonvulsant activity; binding/functional assays; hybrid compounds; multimodal/multitarget compounds; tox studies.

Figure 1

Lead compound (R)-AS-1, [11] and design approach of hybrid compounds obtained in the current studies. The common structural elements forming the new hybrids are highlighted in color.

Scheme 1

Synthesis of intermediates 1–4 and target compounds 5–23.

Scheme 2

Synthesis of intermediates 24–35 and target pyrrolidin-2-on analogs 36–39.

Figure 2

The viability of HepG2 cells incubated in the presence of 5, 17, and 39. HepG2 cells were exposed to growing concentrations (1–50 µM) of tested compounds for 24 h. Cell viability was measured using an MTT assay. Bars represent mean percent of cell viability normalized to non-treated cells (100%) ± SEM. The doxorubicin was tested as a positive control.

Figure 3

The viability of SH-SY5Y cells incubated in the presence of compounds 5, 17, and 39. SH-SY5Y cells were exposed to growing concentrations (1–50 µM) of tested compounds for 24 h. Cell viability was measured using an MTT assay. Bars represent mean percent of cells viability normalized to non-treated cells (100%) ± SEM. The doxorubicin was tested as a positive control.