Cardiotonic and coronary vasodilator responses to milrinone, forskolin, and analog P87-7692 in the anesthetized dog.

Forskolin and milrinone both increase cyclic AMP concentrations to enhance cardiac contractility and cause vascular dilation in vitro and in vivo. However, forskolin acts via direct stimulation of adenylate cyclase while milrinone inhibits phosphodiesterase (PDE-III) activity. The forskolin analog, 7-desacetyl-7-(O-propionyl)-hydroxyl-aminocarbonyl-forskolin (P87-7692) has also been shown to directly stimulate adenylate cylase and increase cyclic AMP production in isolated cardiac tissue; however, the in vivo activity of this compound has not been described. Thus, the purpose of this study was to compare the cardiovascular effects of equivalent doses of these compounds and to further characterize the cardiotonic activity of P87-7692 in the anesthetized dog. It was found that both i.v. (3-30 micrograms/kg) and intracoronary (0.1-30 micrograms) administration of milrinone, forskolin, and P87-7692 caused dose-related positive inotropic, coronary, and peripheral vasodilator effects in anesthetized dogs; however, P87-7692 produced significantly greater and more sustained cardiotonic activity following a single 30-micrograms/kg, i.v., bolus injection when compared to the same dose of milrinone and forskolin. Analysis of the dose-response relationship between the changes in contractile force and heart rate for these compounds revealed that a 50% augmentation in contractile force was associated with increases in heart rate of 2.1% for milrinone, 6.4% for P87-7692, and 13.7% for forskolin. These data indicate an improved separation between the chronotropic and inotropic effects for P87-7692 as compared to forskolin. All three compounds also produced coronary vasodilation in vivo and in vitro; however, P87-7692 consistently showed greater activity relative to the same doses of milrinone and forskolin. Moreover, P87-7692 was significantly (p less than 0.05) more potent at relaxing KC1-precontracted canine coronary rings, with an EC50 of 2.1 x 10(-7) M as compared to 1.1 x 10(-6) M for forskolin and 3.2 x 10(-6) M for milrinone. The results of these studies indicate that structural modification of the forskolin molecule can increase the separation between positive inotropic and chronotropic effects, improve the overall hemodynamic profile, and prolong the duration of cardiotonic activity for this class of compounds.

(Aminoalkyl)carbamates of forskolin: intermediates for the synthesis of functionalized derivatives of forskolin with different specificities for adenylyl cyclase and the glucose transporter.

(Aminoalkyl)carbamates of forskolin were synthesized at the 6- and 7-hydroxyl positions of forskolin with the length of the alkyl chain varying from ethyl to heptyl. Two of these derivatives, 7-[[(2-aminoethyl)amino]carbonyl]-7-desacetylforskolin (2) and 6-[[(2-aminoethyl)amino]carbonyl]forskolin (3), were used to synthesize iodinated derivatives of forskolin that bind with high affinity to adenylyl cyclase in bovine brain membranes and the glucose transporter in human erythrocyte membranes, respectively. Hydroxyphenyl derivatives of forskolin were prepared from the (aminoalkyl)carbamates and tested for their ability to bind to adenylyl cyclase in bovine brain membranes and the glucose transporter in human erythrocyte membranes. The 6-derivative (18) of forskolin had a Kd of 9 nM at adenylyl cyclase and was more potent than either the 7-derivatives or the 6-derivatives of 7-desacetylforskolin. The 7-derivatives were more potent at binding to the glucose transporter than forskolin. In contrast, the 6-derivatives had Kd's greater than 100 microM at the glucose transporter. Isothiocyanates and N-bromoacetyl derivatives were synthesized from 2 and 3 as potential alkylating agents for forskolin binding sites. The alkylating agents produced an irreversible loss of forskolin binding to adenylyl cyclase. In contrast, the alkylating agents bound reversibly to the glucose transporter.

Cardiac adenylate cyclase activity, positive chronotropic and inotropic effects of forskolin analogs with either low, medium or high binding site affinity.

A series of in vitro studies were conducted examining the adenylate cyclase stimulation, positive chronotropic and inotropic effects of forskolin and nine analogs which exhibited a range of [3H]forskolin binding site affinities (K1) from 0.020 to 3.174 microM. A significant (P less than .001) linear correlation (r = 0.94) was found between binding site affinity and adenylate cyclase stimulation (EC50) for forskolin and the nine structural analogs. Adenylate cyclase activity was also significantly correlated with the positive chronotropic and inotropic effects of these substances on isolated guinea pig atria. Compounds with K1 values between 0.020 and 1.136 microM produced concentration-dependent increases in heart rate and contractile force in isolated spontaneous and electrically paced guinea pig atria, respectively. In contrast, an analog with a K1 of 3.174 microM caused significant (P less than .05) negative chronotropic and inotropic effects at concentrations above 10 microM. The optimal separation between positive inotropic and chronotropic activity was found with compounds displaying potent [3H]forskolin binding site affinity but moderate adenylate cyclase stimulation, i.e., K1 and EC50 values of approximately 0.05 to 0.10 and 3 microM, respectively. The results of this study show that the forskolin analog, P87-7692 [7-desacetyl-7-(O-propionyl)-hydroxyl amino-carbonyl-forskolin], has marked activity with a wide separation between positive inotropic (248 +/- 41%) and chronotropic effects (43 +/- 13%) at 6.2 microM and may serve as a prototype for a forskolin-based cardiotonic.