Synthesis, characterization, in silico and pharmacological investigation of new benzimidazole derivatives against neurodegenerative diseases

Abstract

Neurodegeneration is age-associated progressive decline of structural and functional features of neurons. Neurodegenerative diseases such as Alzheimer’s disease & Parkinson’s disease usually arise from unknown reasons and associated with pathological expressions that primarily affect specific neurons. The relentless progression rate in such disorders is involved with several unlocked mysteries. The biology of neurodegeneration include several genetic and molecular factors such as neuro-inflammation, oxidative stress, mitochondrial dysfunction, activation of microglia and many other biochemical events, that lead to apoptosis. The increased cellular manifestation of reactive oxygen species (ROS) and production of numerous inflammatory mediators such as nuclear factor κB (NF-κB), cyclooxygenase-2 (COX2), tumor necrosis factor (TNF-α), ionized calcium-binding adapter molecule-1 (Iba-1) & interleukins (IL-1β, IL-6) plays a critical role in the pathogenesis of neurodegeneration. This cascade not only explains the complexity of events taking place at different levels but also provide a potential therapeutic roadmap for the treatment of neurodegenerative disorders. Many synthetic derivatives containing amide linkage as a central scaffold have emerged as successful agents for the treatment of neurological disorders. In this study, ten novel benzimidazole acetamide derivatives (FP1-FP10) were synthesized and characterized to investigate its neuroprotective effects in ethanol-induce neurodegeneration in a rat model. Further, five derivatives (FP1, FP3, FP4, FP7 and FP8) were selected for in vivo molecular analysis based on in silico molecular docking, in silico ADMET prediction and preliminary in vitro antioxidant screening assay. Molecular analysis revealed elevated expression of neuroinflammatory cytokines (NF-κB, COX2, TNF-α, Iba-1 and IL-6), increased cellular oxidative stress, and reduced antioxidant enzymes (Glutathione (GSH) and glutathione s-transferase (GST)) in ethanol exposed rats brain which was further validated by enzyme-linked immunosorbent assay (ELISA). The behavioral (Y-maze and x Morris water maze test) results revealed that ethanol treated rats showed more escape latency and had lower alteration percentage as compared to the vehicle treated rats. Moreover, real time PCR was used for the detection and expression of neuroinflammatory markers (TNF-α) in the rat brain. Molecular docking analysis was performed to assess the affinity of synthesized benzimidazole acetamide derivatives against selected protein targets (COX2, TNF-α, Iba-1, IL-1β and IL-6). The ADME predictor tool predicted the drug-likeness, non carcinogenicity, non-mutagenicity and ability to cross blood brain barrier (BBB) of FP1, FP3, FP4, FP7 and FP8. Pretreatment with novel benzimidazole acetamide derivatives (FP1, FP3, FP4, FP7 and FP8) significantly ameliorated the ethanol-induced memory deficits, oxidative stress, and proinflammatory cytokines (NF-κB, COX2, TNF-α, Iba-1 and IL-6) in the cortex and hippocampus of rat brains. The morphological assessment (H&E staining and immunohistochemistry) further validated these findings. While, real time PCR results depicted that ethanol significantly down-regulated the TNF-α mRNA in brain cortical tissue as compared to ethanol treated group. The multipurpose nature of benzimidazole acetamide derivatives and its versatile affinity towards numerous receptors highlight its multistep targeting potential. These results indicated the neuroprotective potential of benzimidazole acetamide derivatives (FP1, FP3, FP4, FP7 & FP8) in ethanol-induced neurodegeneration which may potentially be due to inhibition of neuroinflammatory-oxidative stress vicious cycle.