New milrinone analogues: in vitro study of structure-activity relationships for positive inotropic effect, antagonism towards endogenous adenosine, and inhibition of cardiac type III phosphodiesterase

Abstract

Two mechanisms are responsible for the positive inotropic effect of the cardiotonic drug milrinone, i.e., inhibition of type III cAMP phosphodiesterase (PDE III), and displacement of endogenous adenosine from A(1) inhibitory receptor. Since PDE III inhibition may increase the likelihood of cardiac arrhythmias by increasing cAMP content, our attention focused on the synthesis of new compounds with more pronounced characteristics as adenosine antagonists. In this work, four new milrinone analogues were studied, in comparison with the parent drug, for their effects on the contractility of guinea pig isolated atrial preparations, their ability to antagonize endogenous adenosine at the level of A(1) receptor, and to inhibit the activity of PDE III partially purified from guinea pig heart. The new compounds present various chemical substitutions with respect to the parent drug: in compounds SF397 (methyl 5-cyano-2-methyl-6-oxo-1,6-dihydropyridine-3-carboxylate) and SF399 (benzyl 5-cyano-2-methyl-6-oxo-1,6-dihydropyridine-3-carboxylate), the 4-pyridil moiety of milrinone was replaced with a methoxycarbonyl and a benzyloxycarbonyl group, respectively; the same structural modifications were also associated with the replacement of the cyano-group in 5-position with an acetyl group in compounds SF416 (methyl 5-acetyl-2-methyl-6-oxo-1,6-dihydropyridine-3-carboxylate) and SF419 (benzyl 5-acetyl-2-methyl-6-oxo-1,6-dihydropyridine-3-carboxylate). All the new compounds had a marked positive inotropic effect, most of them also being more active and more potent than milrinone. When their affinity for A(1) receptor was assessed as the displacement of [(3)H] 8-cyclopentyl-1,3-dipropylxanthine ([(3)H]DPCPX) from cardiac membranes, SF397 and SF399 showed affinity (K(i) of about 600 nM) similar to that of milrinone (K(i) 550 nM). By contrast, SF416 and SF419 had very low (K(i) of about 10000 nM) or scarce (K(i) of about 2000 nM) anti-adenosine component, respectively. All the new compounds inhibited PDE III activity, their K(i) values proceeding in the following order: milrinone (3.80 microM) <SF397 (7.00 microM) <SF399 (8.80 microM) <SF416 (35.00 microM) SF419< (155.00 microM). To better characterize the mechanisms responsible for the positive inotropic response of the new compounds, we also investigated the effects of new analogues on some systems (ATP-dependent Ca(2+) uptake, Ca(2+)ATPase, Na(+)/K(+)ATPase, Na(+)/Ca(2+) exchange carrier) or a receptor (beta-adrenoceptor) mainly involved in the control of cardiac contractility. None of the tested compounds inhibited enzyme or transport systems; however, SF397, SF399 and SF416, although to different extents, had a direct beta-adrenergic action. Indications about structure-activity relationships are tentatively discussed, in order to obtain useful information for the design of new analogues with better pharmacological profiles.