Differential inhibition of cardiac cyclic nucleotide phosphodiesterase isozymes by cardiotonic drugs

Several new cardiotonic drugs are postulated to act as potent inhibitors of cyclic nucleotide phosphodiesterase activity. Unfortunately, the presence of multiple phosphodiesterase isozymes in cardiac muscle makes it difficult to determine whether any of these enzymes are specific targets for the cardiotonic agents. We have developed a method for rapid isolation and assay of bovine cardiac muscle phosphodiesterases using monoclonal antibodies that distinguish between isozymes without inhibiting catalytic activities. By this method, milrinone, amrinone, and MDL 17,043 were tested for potency as inhibitors of soluble bovine heart phosphodiesterases. All three drugs were highly selective for a low-Km, cyclic GMP (cGMP)-inhibited phosphodiesterase. LC5Os (half-maximal inhibitory concentrations) for cGMP-inhibited phosphodiesterase were 0.5μM (milrinone), 30μM (amrinone), and 2 gM (MDL 17,043) when measured at 0.35μM cyclic AMP (cAMP). Milrinone and MDL 17,043 had greater than 50-fold lower potencies for the other heart phosphodiesterases (and arnrinone 20-fold). These data suggest the cGMP-inhibited phosphodiesterase as a probable site of action for these new cardiotonic drugs. In addition, the method described here should be useful for screening new drugs, and for studying physical and chemical properties of this phosphodiesterase/drug receptor in cardiac and other tissues.