β-Secretase (BACE1) Inhibitors from Pomegranate (Punica granatum) L. Husk

Alzheimer's disease (AD) is a neurodegenerative disorder clinically characterized by progressive dementia that inevitably leads to incapacitation and death. Two characteristic brain lesions define AD at the microscopic level: (1) amyloid plaques, extracellular deposits primarily composed of 4 kDa, 40–42 amino acid Aβ peptide, a product of APP proteolysis, and (2) neurofibrillary tangles, and intracellular aggregates of the microtubule associated protein tau. The relationships between amyloid plaques, neurofibrillary tangles, and the pathogenic mechanisms of AD are controversial. Evidence, however, suggests that Aβ is critically involved at an early stage in AD pathology. Two proteolytic cleavage events are required to generate Aβ from its precursor, one at the N-terminus by an enzyme termed β-secretase and one at the C-terminus by an enzyme termed γ-secretase. Among the secretases, a novel transmembrane aspartic protease BACE1 (for β-site APP, cleaving enzyme 1), also known as Asp2 (for novel aspartic protease 2) and memapsin 2 (for membrane aspartic protease/pepsin 2), is at present the most attractive target for the inhibition of amyloid production.

In the course of screening anti-dementia agents from natural products, two β-secretase (BACE1) inhibitors were isolated from the ethyl acetate soluble fraction of pomegranate husk. Chromatographic separation including silica gel, Sephadex LH-20, and RP-HPLC afforded two active principles. They were identified as ellagic acid (1) and punicalagin (2) and were shown to non-competitively inhibit β-secretase (BACE1) with the IC₅₀ values of 3.9×10^-6 M and 4.1×10^-7 M, respectively. The Ki values of 1 and 2 were 2.4×10^-5 M and 5.9×10^-7 M. They were less inhibitory to α-secretase (TACE) and other serine proteases such as chymotrypsin, trypsin, and elastase, suggesting that they were relatively specific inhibitors of BACE1.