A combinatorial approach to the identification of dipeptide aldehyde inhibitors of β-amyloid production

In an effort to rapidly identify potent inhibitors of A beta production and to probe the amino acid sequence specificity of the protease(s) responsible for the production of this peptide, a large number of dipeptide aldehydes were combinatorially synthesized and manually evaluated for their inhibitory properties. The starting point for this study was the dipeptide aldehyde carbobenzoxyl-valinyl-phenylalanal previously shown to inhibit the production of A beta in CHO cells stably transfected with the cDNA encoding beta APP695. Pools of related dipeptide aldehydes were combinatorially synthesized, and the most active pool was deconvoluted, resulting in the identification of the most active inhibitor of this pool. Systematic optimization of this inhibitor resulted in a series of dipeptide aldehydes with enhanced potencies relative to carbobenzoxylvalinyl-phenylalanal. The most active dipeptide aldehydes were those that possessed hydrophobic amino acids at both the P1 and P2 positions. The most potent compound identified in this study was 3,5-dimethoxycinnamamide-isoleucinyl-leucinal with an IC50 of 9.6 mu M, approximately 10-fold more active than carbobenzoxyl-valinyl-phenylalanal. In immunoprecipitation experiments using antibodies directed toward either A beta 1-40 or A beta 1-42, 3,5-dimethoxycinnamamide-isoleucinyl-leucinal, like carbobenzoxyl-valinyl-phenylalanal, preferentially inhibited the shorter 1-40 form of A beta, whereas the longer 1-42 form was not as strongly inhibited. These results suggest that dipeptide aldehydes related to carbobenzoxyl-valinyl-phenylalanal inhibit A beta through similar mechanisms and demonstrate the utility of a combinatorial synthesis approach to rapidly identify potent inhibitors of A beta production.