SYNTHESIS AND BIOLOGICAL-ACTIVITY OF A SERIES OF POTENT FLUOROMETHYL KETONE INHIBITORS OF RECOMBINANT HUMAN CALPAIN-I

Calpain I, an intracellular cysteine protease, has been implicated in the neurodegeneration following an episode of stroke. In this paper, w e report on a series of potent dipeptide fluoromethyl ketone inhibitor s of recombinant human calpain I (rh calpain I). SAR studies revealed that while calpain I tolerates a variety of hydrophobic groups at the P-1 site, Leu at Pa is preferred. However, the nature of the N-termina l capping group has a significant effect on the inhibitory activity of this series of compounds. Compound 4e rahydroisoquinolin-2-yl)carbony l-Leu-D,L-Phe-CH2F] , having a tetrahydroisoquinoline containing urea as the N-terminal capping group, is the most potent dipeptide fluorome thyl ketone inhibitor of calpain I (with a second-order rate constant for inactivation of 276 000 M-1 s(-1)) yet reported; tripeptide 4k (Cb z-Leu-Leu-D,L-Phe-CH2F) is equipotent. A number of compounds presented in this study displayed excellent selectivity for calpain I over cath epsins B and L, two related cysteine proteases. Compounds which exhibi ted good inhibitory activity in the assay against isolated rh calpain I also inhibited intracellular calpain I in a human cell line. Thus, i n an intact cell assay, compounds 4e and 4k inhibited calpain I with I C50 values of 0.2 and 0.1 mu M, respectively. Finally, we also disclos e the first example of fluorination of a dipeptide enol silyl ether to generate the corresponding dipeptide fluoromethyl ketone.