Conclusions

In summary, this study successfully synthesized and characterized a novel series of peptidomimetics demonstrating reversible, micromolar inhibition of both human acetylcholinesterase (hAChE) and butyrylcholinesterase (hBChE). The biological evaluation of the synthesized compounds revealed a distinct preference for hBChE over hAChE. This indicates that the studied scaffold possesses an inherent selectivity for the BChE active site, which may be attributed to the larger active site of BChE compared to the more constricted gorge of AChE. Notably, compounds 5 and 8 exhibited significant selectivity for hBChE, displaying 279- and 169-fold higher preference, respectively. However, the lipophilicity of those compounds is high and the introduction of polar functional groups or substituents to decrease log P while maintaining or improving potency is necessary. Therefore, compound 1 can be considered the primary lead for both enzymes in this series, as it has a good balance of potency and lipophilicity.

Principal component analysis effectively differentiated compounds based on their enzyme selectivity or joint inhibition, and these findings were supported by quantum-chemical docking simulations, which revealed key driving interactions. This integrated approach, combining experimental enzyme inhibition assays with computational modelling, not only elucidates the structural basis of peptidomimetic scaffold binding to cholinesterases but also establishes a strong foundation for the rational design of more potent and selective cholinesterase inhibitors with potential therapeutic applications in neurological disorders such as Alzheimer’s disease.