An orally active formulation of angiotensin-(1-7) produces an antithrombotic effect

Abstract

Introduction and objective: The heptapeptide angiotensin-(1-7) is a component of the renin-angiotensin system, which promotes many beneficial cardiovascular effects, including antithrombotic activity. We have recently shown that the antithrombotic effect of angiotensin-(1-7) involves receptor Mas-mediated NO-release from platelets. Here, we describe an orally active formulation based on angiotensin-(1-7) inclusion in cyclodextrin [Ang-(1-7)- CyD] as an antithrombotic agent. Cyclodextrins are pharmaceutical tools that are used to enhance drug stability, absorption across biological barriers and gastric protection.

Method: To test the antithrombotic effect of Ang-(1-7)-CyD, thrombus formation was induced in the abdominal vena cava of spontaneously hypertensive rats that were pretreated either acutely or chronically with Ang-(1-7)-CyD. Male Mas-knockout and wild-type mice were used to verify the role of the Mas receptor on the effect of Ang-(1-7)-CyD.

Results: Acute or chronic oral treatment with Ang-(1-7)-CyD promoted an antithrombotic effect (measured by thrombus weight; all values are, respectively, untreated vs. treated animals) in spontaneously hypertensive rats (acute: 2.86 ± 0.43 mg vs. 1.14 ± 0.40 mg; chronic: 4.27 ± 1.03 mg vs. 1.39 ± 0.68 mg). This effect was abolished in Mas-knockout mice (thrombus weight in Mas wild-type: 0.76 ± 0.10 mg vs. 0.37 ± 0.02 mg; thrombus weight in Mas-knockout: 0.96 ± 0.11 mg vs. 0.87 ± 0.14 mg). Furthermore, the antithrombotic effect of Ang-(1-7)-CyD was associated with an increase in the plasma level of Angiotensin-(1-7).

Conclusion: These results show for the first time that the oral formulation Ang-(1-7)-CyD has biological activity and produces a Mas-dependent antithrombotic effect.

Figure 1

Ang-(1-7)-CyD promoted antithrombotic effects in SHRs. A) Acute treatment with Ang-(1-7)-CyD (five hours before thrombus induction) inhibited thrombus formation when administered in doses of the equivalent of 10 µg/kg or 30 µg/kg of Ang-(1-7). B) Chronic administration of Ang-(1-7)-CyD [equivalent of 30 µg/kg of Ang-(1-7) per day] over the course of eight weeks promoted a strong inhibition of thrombus formation in SHRs. *p<0.05 significantly different from the respective control group treated with CyD (one-way ANOVA for panel A and unpaired Student's t-test for panel B). Each column represents the mean ± SEM from 7-10 experiments.

Figure 2

The antithrombotic effect of Ang-(1-7)-CyD is dependent on Mas. A) Ang-(1-7)-CyD promoted a reduction in FeCl₃ solution-induced thrombus weight in Mas^+/+ mice; this effect was absent in Mas^-/- mice. B) Oral administration of Ang-(1-7)-CyD [equivalent of 100 µg of Ang-(1-7)] increases the plasma level of Ang-(1-7) in C57Bl/6 mice. *p<0.05 and **p<0.01 significantly different from the control group treated with CyD (unpaired Student's t-test). Each column represents the mean ± SEM from 6-8 experiments.