Synthesis, Biological Evaluation and Molecular Modelling Studies of Thiophene Piperazine-Carbamate Derivatives as Multi-Target Agents for Alzheimer’s Disease

Abstract

Alzheimer’s disease is a progressive neurodegenerative condition marked by cognitive deterioration, memory deficits and behavioural changes, underscoring the pressing need for innovative therapeutic strategies. While acetylcholinesterase (AChE) inhibitors remain a cornerstone in managing cholinergic dysfunction in AD, the multifaceted nature of the disease, which also involves oxidative stress, necessitates the development of multi-targeted therapeutic agents. In response to this challenge, a series of novel thiophene piperazine-carbamate hybrids (8a-g) was designed and synthesized to simultaneously inhibit AChE and butyrylcholinesterase (BChE), while also possessing potent antioxidant properties, as evidenced by ABTS radical scavenging activity. In vitro analysis revealed robust inhibition of AChE and BChE across all compounds, with a clear preference for AChE inhibition. Among these hybrids, compound 8e exhibited exceptional potency, achieving AChE inhibition (IC₅₀ = 0.12 ± 0.001 µM), BChE inhibition (IC₅₀ = 12.29 ± 0.02 µM) and antioxidant activity (IC₅₀ = 0.192 ± 0.001 µM). Biophysical kinetic studies confirmed that compound 8e operates via mixed-type inhibition of AChE, with inhibition constants (K_i1 = 0.158 µM, K_i2 = 0.347 µM). Molecular docking studies substantiated that both compounds bind effectively to key residues in the catalytic active site (CAS) and peripheral anionic site (PAS) of AChE, supporting their dual inhibition mechanism. Significantly, compounds 8e, 8d, 8g and 8a stand out as promising candidates for further development due to their dual-target inhibition and antioxidant properties. Structure-activity relationship (SAR) analysis highlighted that shorter, unbranched alkyl chains enhance binding affinity and inhibitory potency, while bulkier or branched groups introduce steric hindrance, reducing efficacy. Collectively, these findings position the thiophene piperazine-carbamate hybrids, particularly compounds 8e and 8d, as potent multi-target agents with significant potential for addressing both cholinergic dysfunction and oxidative stress in Alzheimer’s disease therapy.