Current Medicinal Chemistry - Volume 9, Issue 11, 2002

A considerable body of evidence has accumulated in recent years implicating the ß-amyloid protein (Aß) in the etiology of Alzheimer's disease (AD). The highly hydrophobic Aß can nucleate and form neurotoxic fibrils that are the principal components of the cerebral plaques characteristic of AD. Aß is formed from the amyloid-ßprecursor protein (APP) through two protease activities. First, ß-secretase cleaves APP at the Aß N-terminus, resulting in a soluble, secreted APP derivative (ß-APPs) and a 12 kDa membrane-retained C-terminal fragment. The latter is further processed to Aß by γ secretases, which cleave within the single transmembrane region. Other APP molecules can be cleaved by α-secretase within the Aß region, thus precluding Aß formation. Both β- and γ- secretase have become prime targets for the development of therapeutic agent that reduce Aβ production. β-Secretase has recently been identified as a new membrane-anchored aspartyl protease in the cathepsin D family. Inhibitor profiling, site-directed mutagenesis, and affinity labeling together have suggested that the multi-pass presenilins are γ-secretases, novel intramembrane-cleaving aspartyl proteases activated through autoproteolysis. In this article, we review the current knowledge of γ-secretase biochemistry and cell biology and the development of inhibitors of this important therapeutic target.

In the nonamyloidogenic processing pathway the Alzheimer's amyloid precursor protein (APP) is proteolytically cleaved by α-secretase. As this cleavage occurs at the Lys16-Leu17 bond within the amyloid β domain, it prevents deposition of intact amyloidogenic peptide. In addition, the large ectodomain (sAPPα) released by the action of α-secretase has several neuroprotective properties. Studies with a range of hydroxamic acid-based compounds, such as batimastat, indicate that α-secretase is a zinc metalloproteinase, and members of the adamalysin family of proteins, TACE, ADAM10 and ADAM9, all fulfil some of the criteria required of α-secretase. APP is constitutively cleaved by α-secretase in most cell lines. However, on stimulation with muscarinic agonists or activators of protein kinase C, such as phorbol esters, the α-secretase cleavage of APP is up-regulated. The constitutive α- secretase activity is primarily at the cell surface, while the regulated activity is predominantly located within the Golgi. The beneficial action of cholinesterase inhibitors may in part be due to activation of muscarinic receptors, resulting in an up-regulation of α-secretase. Other agents can also increase the nonamyloidogenic cleavage of APP including estrogen, testosterone, various neurotransmitters and growth factors. As the α- secretase cleavage of APP both precludes the deposition of the amyloid β peptide and releases the neuroprotective sAPPα, pharmacological up-regulation of α-secretase may provide alternative therapeutic approaches for Alzheimer's disease.

The discovery of small molecules that inhibit β-peptide and other amyloid fibril formation has been impeded by featureless structure-activity-relationships, low apparent efficacy of inhibition, and by the perception that protein-protein interactions are too diffuse to be proper medicinal chemistry targets. Atomic resolution structural information on the critical target species are lacking. Despite these difficulties, substoichiometric inhibitors of fibrillogenesis have been reported. By carefully defining assay systems and considering a spectrum of data from different types of measurements, medicinal chemistry can improve molecular structures and their measured effectiveness can be better understood. Compounds with good pharmacokinetic and toxicologic profiles that persist in the target tissue at useful concentrations may be as useful as would extremely high affinity inhibitors with less favorable biological properties.

Recent identification of β-scretasse being a membrane-associated aspartic protease has stimulated strong interest on this enzyme as a therapeutic target for Alzheimer's disease. Here we review the current understanding in the structure-function relationship as well as the status in the design of its inhibitors of this protease, memapsin 2 (BACE, ASP-2). The development in the basic tools, such as the preparation of recombinant memapsin 2, the assay method for kinetic measurements of inhibition, the determination of the subsite specificity and the crystal structure of memapsin 2 complexed to a tight-binding inhibitor, has made the structural based inhibitor design possible. More recent inhibitors, having Ki values in the nanomolar range and molecular size in low 700 Da, show some promise that clinically useful inhibitors of β-scretasse may be attainable.