Acta Pharmacologica Sinica (2009) 30: 1195-1203; doi: 10.1038/aps.2009.91; published online 6 July 2009

Aim: To design novel bifunctional derivatives of huperzine B (HupB) based on the concept of dual binding site of acetylcholinesterase (AChE) and evaluate their pharmacological activities for seeking new drug candidates against Alzheimer’s disease (AD).

Methods: Novel 16-substituted bifunctional derivatives of HupB were synthesized through chemical reactions.  The inhibitory activities of the derivatives toward AChE and butyrylcholinesterase (BuChE) were determined in vitro by modified Ellman’s method.  Cell viability was quantified by the reduction of MTT.

Results: A new preparative method was developed for the generation of 16-substituted derivatives of HupB, and pharmacological trials indicated that the derivatives were multifunctional cholinesterase inhibitors targeting both AChE and BuChE.  Among the derivatives tested, 9c , 9e, 9f , and 9i were 480 to 1360 times more potent as AChE inhibitors and 370 to 1560 times more potent as BuChE inhibitors than the parent HupB.  Further preliminary pharmacological trials of derivatives 9c and 9i were performed, including examining the mechanism of AChE inhibition, the substrate kinetics of the enzyme inhibition, and protection against hydrogen peroxide (H₂O₂)-induced cytotoxicity in PC12 cells.

Conclusion: Preliminary pharmacological evaluation indicated that 16-substituted derivatives of HupB, particularly 9c and 9i, would be potentially valuable new drug candidates for AD therapy, and further exploration is needed to evaluate their pharmacological and clinical efficacies.

Keywords: Alzheimer’s disease; huperzine B; acetylcholinesterase; butyrylcholinesterase; cholinesterase inhibitors