Visualization of time-dependent inactivation of human tumor cathepsin B isozymes by a peptidyl fluoromethyl ketone using a fluorescent print technique.

A peptidyl fluoromethyl ketone (Z-Phe-Ala CH2F) was found to be an effective compound in a time dependent inactivation of cathepsin B isozymes from a number of tissues including human tumors. The effect was visualized by employing an activity-specific fluorescent print technique preceded by isoelectric focusing. The technique could yield additional information of selective inhibition of isozymes as observed with rat pancreas. The fluoromethyl ketone is 30-fold more potent than the known inhibitor of cathepsin B, Z-Phe-AlaCHN2 in parallel evaluation. Furthermore, the fluoromethyl ketone may have in vivo potential in the inhibition of cathepsin B, in view of the results of toxicological studies. The findings demonstrate that the application of enzyme-directed overlay membranes, impregnated with specific substrates, following isoelectric focusing could be very useful in the study of proteases and their involvement in the oncogenic process.