Synthesis, in vitro biological evaluation and molecular modelling of new 2-chloro-3-hydrazinopyrazine derivatives as potent acetylcholinesterase inhibitors‏ on PC12 cells

Background: The loss of cholinergic neurotransmission in Alzheimer's disease (AD) patients' brain is accompanied by a reduced concentration of Acetylcholine (ACh) within synaptic clefts. Thus, the use of acetylcholinesterase inhibitors (AChEIs) to block the cholinergic degradation of ACh is a promising approach for AD treatment. In the present study, a series of 2-chloro-3-hydrazinopyrazine derivatives (CHP1-5) were designed, synthesized, and biologically evaluated as potential multifunctional anti-AD agents.

Methods: In addition, the chemical structures and purity of the synthesized compounds were elucidated through using IR, ¹H and ¹³C NMR, and elemental analyses. Further, the intended compounds were assessed in vitro for their AChE inhibitory and neuroprotective effects. Furthermore, DPPH, FRAP and ABTS assays were utilized to determine their antioxidant activity. The statistical analysis was performed using one-way ANOVA.

Results: Based on the results, CHP4 and CHP5 exhibited strong AChE inhibitory effects with the IC₅₀ values of 3.76 and 4.2 µM compared to the donepezil (0.53 µM), respectively. The study examined the effect and molecular mechanism of CHP4 on the Ab1-42-induced cytotoxicity in differentiated PC12 cells. At concentrations of 0-100 μM, CHP4 was non-toxic in PC12. Additionally, Ab1-42 significantly stimulated tau hyperphosphorylation and induced differentiated PC12 cell death. Further, CHP4 resulted in diminishing the Ab1-42-induced toxicity in PC12 cell significantly. CHP4 at 30 μM concentration significantly increased the Ab1-42-induced HSP70 expression and decreased tau hyperphosphorylation.

Conclusions: According to the results of our studies CHP4 can be considered as safe and efficient AChEI and employed as a potential multifunctional anti-AD agent.