Yannick Marc

The role of the brain renin-angiotensin system in hypertension: implications for new treatment.

Hypertension affects 26% of adults and is in constant progress related to increased incidence of obesity and diabetes. One-third of hypertensive patients may be successfully treated with one antihypertensive agent, one-third may require two agents and in the remaining patients will need three agents for effective blood pressure (BP) control. The development of new classes of antihypertensive agents with different mechanisms of action therefore remains an important goal. Brain renin-angiotensin system (RAS) hyperactivity has been implicated in hypertension development and maintenance in several types of experimental and genetic hypertension animal models. Among the main bioactive peptides of the brain RAS, angiotensin (Ang) II and Ang III have similar affinities for type 1 (AT1) and type 2 (AT2) Ang II receptors. Following intracerebroventricular (i.c.v.) injection, Ang II and Ang III similarly increase arginine-vasopressin (AVP) release and BP. Blocking the brain RAS may be advantageous as it simultaneously (1) decreases sympathetic tone and consequently vascular resistance, (2) decreases AVP release, reducing blood volume and vascular resistance and (3) blocks angiotensin-induced baroreflex inhibition, decreasing both vascular resistance and cardiac output. However, as Ang II is converted to Ang III in vivo, the exact nature of the active peptide is not precisely determined. We summarize here the main findings identifying AngIII as one of the major effector peptides of the brain RAS in the control of AVP release and BP. Brain AngIII exerts a tonic stimulatory effect on BP in hypertensive rats, identifying brain aminopeptidase A (APA), the enzyme generating brain Ang III, as a potentially candidate target for hypertension treatment. This has led to the development of potent orally active APA inhibitors, such as RB150--the prototype of a new class of centrally acting antihypertensive agents.

Orally Active Aminopeptidase A Inhibitors Reduce Blood Pressure A New Strategy for Treating Hypertension

Overactivity of the brain renin-angiotensin system has been implicated in the development and maintenance of hypertension. We reported previously that angiotensin II is converted to angiotensin III by aminopeptidase A in the mouse brain. We then used specific and selective aminopeptidase A inhibitors to show that angiotensin III is one of the main effector peptides of the brain renin-angiotensin system, exerting tonic stimulatory control over blood pressure in hypertensive rats. Aminopeptidase A, the enzyme generating brain angiotensin III, thus represents a potential candidate central nervous system target for the treatment of hypertension. Given this possible clinical use of aminopeptidase A inhibitors, it was, therefore, important to investigate their pharmacological activity after oral administration. We investigated RB150, a dimer of the selective aminopeptidase A inhibitor, EC33, generated by creating a disulfide bond. This chemical modification allows prodrug to cross the blood-brain barrier when administered by systemic route. Oral administration of RB150 in conscious DOCA-salt rats inhibited brain aminopeptidase A activity, resulting in values similar to those obtained with the brains of normotensive rats, demonstrating the central bioavailability of RB150. Oral RB150 treatment resulted in a marked dose-dependent reduction in blood pressure in DOCA-salt but not in normotensive rats, with an ED50 in the 1-mg/kg range, achieved in <2 hours and lasting for several hours. This treatment also significantly decreased plasma arginine-vasopressin levels and increased diuresis, which may participate to the blood pressure decrease by reducing the size of fluid compartment. Thus, RB150 may be the prototype of a new class of centrally active antihypertensive agents.

Central Antihypertensive Effects of Orally Active Aminopeptidase A Inhibitors in Spontaneously Hypertensive Rats

Brain renin-angiotensin system hyperactivity has been implicated in the development and maintenance of hypertension. We reported previously in the brain that aminopeptidase A and aminopeptidase N are involved in the metabolism of angiotensin II and angiotensin III, respectively. By using in vivo specific and selective aminopeptidase A and aminopeptidase N inhibitors, we showed that angiotensin III is one of the main effector peptides of the brain renin-angiotensin system, exerting a tonic stimulatory control more than blood pressure in hypertensive rats. Aminopeptidase A, the enzyme generating brain angiotensin III, thus represents a potential target for the treatment of hypertension. We demonstrated here the antihypertensive effects of RB150, a prodrug of the specific and selective aminopeptidase A inhibitor, EC33, in spontaneously hypertensive rats, a model of human essential hypertension. Oral administration of RB150 in conscious spontaneously hypertensive rats inhibited brain aminopeptidase A activity, demonstrating the central bioavailability of RB150 and its ability to generate EC33 into the brain. Oral RB150 treatment dose-dependently reduced blood pressure in spontaneously hypertensive rats with an ED50 of 30 mg/kg, lasting for several hours. This decrease in blood pressure is partly attributed to a decrease in sympathetic tone, reducing vascular resistance. This treatment did not modify systemic renin-angiotensin system activity. Concomitant oral administration of RB150 with a systemic renin-angiotensin system blocker, enalapril, potentiated the RB150-induced blood pressure decrease achieved in <2 hours. Thus, RB150 may be the prototype of a new class of centrally active antihypertensive agents that might be used in combination with classic systemic renin-angiotensin system blockers to improve blood pressure control.

A new strategy for treating hypertension by blocking the activity of the brain renin-angiotensin system with aminopeptidase A inhibitors.

Hypertension affects one-third of the adult population and is a growing problem due to the increasing incidence of obesity and diabetes. Brain RAS (renin-angiotensin system) hyperactivity has been implicated in the development and maintenance of hypertension in several types of experimental and genetic hypertension animal models. We have identified in the brain RAS that APA (aminopeptidase A) and APN (aminopeptidase N), two membrane-bound zinc metalloproteases, are involved in the metabolism of AngII (angiotensin II) and AngIII (angiotensin III) respectively. The present review summarizes the main findings suggesting that AngIII plays a predominant role in the brain RAS in the control of BP (blood pressure). We first explored the organization of the APA active site by site-directed mutagenesis and molecular modelling. The development and the use in vivo of specific and selective APA and APN inhibitors EC33 and PC18 respectively, has allowed the demonstration that brain AngIII generated by APA is one of the main effector peptides of the brain RAS, exerting a tonic stimulatory control over BP in conscious hypertensive rats. This identified brain APA as a potential therapeutic target for the treatment of hypertension, which has led to the development of potent orally active APA inhibitors, such as RB150. RB150 administered orally in hypertensive DOCA (deoxycorticosteroneacetate)-salt rats or SHRs (spontaneously hypertensive rats) crosses the intestinal, hepatic and blood-brain barriers, enters the brain, generates two active molecules of EC33 which inhibit brain APA activity, block the formation of brain AngIII and normalize BP for several hours. The decrease in BP involves two different mechanisms: a decrease in vasopressin release into the bloodstream, which in turn increases diuresis resulting in a blood volume reduction that participates in the decrease in BP and/or a decrease in sympathetic tone, decreasing vascular resistance. RB150 constitutes the prototype of a new class of centrally acting antihypertensive agents and is currently being evaluated in a Phase Ib clinical trial.

Inhibition of brain angiotensin III attenuates sympathetic hyperactivity and cardiac dysfunction in rats post myocardial infarction

AIMS In rats post-myocardial infarction (MI), activation of angiotensinergic pathways in the brain contributes to sympathetic hyperactivity and progressive left ventricle (LV) dysfunction. The present study examined whether angiotensin III (Ang III) is one of the main effector peptides of the brain renin-angiotensin system controlling these effects. METHODS AND RESULTS After coronary artery ligation, Wistar rats were infused intracerebroventricularly for 4 weeks via minipumps with vehicle, the aminopeptidase A (APA) inhibitor RB150 (0.3 mg/day), which blocks the formation of brain Ang III, or losartan (0.25 mg/day). Blood pressure (BP), heart rate, and renal sympathetic nerve activity in response to air stress and acute changes in BP were measured, and LV function was evaluated by echocardiography and Millar catheter. At 4 weeks post-MI, brain APA activity was increased, sympatho-excitatory and pressor responses to air stress enhanced, and arterial baroreflex function impaired. LV end-diastolic pressure (LVEDP) was increased and ejection fraction (EF) and maximal first derivative of change in pressure over time (dP/dt(max)) were decreased. Central infusion of RB150 during 4 weeks post-MI normalized brain APA activity and responses to stress and baroreflex function, and improved LVEDP, EF, and dP/dt(max). Central infusion of losartan had similar effects but was somewhat less effective, and had no effect on brain APA activity. CONCLUSION These results indicate that brain APA and Ang III appear to play a pivotal role in the sympathetic hyperactivity and LV dysfunction in rats post-MI. RB150 may be a potential candidate for central nervous system-targeted therapy post-MI.

[OP.6D.03] MODE OF ACTION OF RB150, AN AMINOPEPTIDASE A INHIBITOR PRODRUG AS A CENTRALLY-ACTING ANTIHYPERTENSIVE AGENT IN DOCA-SALT HYPERTENSIVE RATS.

Objective: In the brain, angiotensin III (AngIII) generated from angiotensin II (AngII) by aminopeptidase A (APA) exerts a tonic stimulatory action on the control of blood pressure (BP) in hypertensive rats. Inhibition of brain APA by RB150 (also known as QGC001), an orally active prodrug of the specific and selective APA inhibitor EC33, blocks brain AngIII formation and normalizes BP in hypertensive rats. Thus, RB150 has emerged as the prototype of a new class of centrally-acting antihypertensive agents and is currently evaluated in a phase II clinical trial in hypertensive patients. Since following blockade of brain AngIII formation by RB150, no AngII accumulation was observed, we aimed to delineate if this treatment induces the activation of another metabolic pathway of brain AngII, in particular angiotensin converting enzyme type 2 (ACE2) which converts AngII into angiotensin 1–7 (Ang1-7). Design and method: For this purpose, RB150 and MLN4760, an ACE2 inhibitor, were administered by intracerebroventricular (ICV) route after insertion of an intracerebral cannula to alert DOCA-salt rats, a model of salt-dependent hypertension. Mean arterial blood pressure (MABP) was recorded via an arterial femoral catheter. Maximal MABP decrease and area under the curve (AUC) of the variations in MABP were calculated. Results: ICV administration of RB150 (100 &mgr;g) significantly decreased MABP compared to vehicle (−23 ± 3 mmHg vs −6 ± 2 mmHg; AUC: 17.6 ± 2 vs 2.1 ± 3). Administration of MLN4760 (10 &mgr;g) didn’t induce any significant change of MABP compared to vehicle (−10 ± 3 mmHg; AUC: 1.4 ± 2). Experiments in progress show that a co-administration of MLN4760 (10 &mgr;g) with RB150 (100 &mgr;g) partially inhibits the RB150-induced BP decrease. Conclusions: In conclusion, ICV administration of the ACE2 inhibitor alone has no effect on BP indicating that in DOCA-salt rats, brain Ang1-7 does not exert a tonic stimulatory effect on the control of BP. However, when conversion of brain AngII into AngIII is blocked by RB150, there is an activation of the conversion by ACE2 of brain AngII into Ang1-7, which by acting on the Mas receptor, may participate to the RB150-induced BP decrease in hypertensive rats.

[OP.8C.06] CENTRAL ANTIHYPERTENSIVE EFFECTS OF CHRONIC TREATMENT WITH RB150/QGC001, AN ORALLY ACTIVE AMINOPEPTIDASE A INHIBITOR PRODRUG IN DOCA-SALT HYPERTENSIVE RATS

Objective: Hyperactivity of the brain renin-angiotensin system (RAS) is implicated in the development and maintenance of arterial hypertension (HTA). We previously showed that angiotensin III (AngIII) is one of the main effector peptides of the brain RAS, exerting a tonic stimulatory control over blood pressure (BP) in alert hypertensive rats. Therefore, aminopeptidase A (APA) the enzyme generating brain AngIII, represents a new therapeutic target for the treatment of HTA. This led to the development of RB150 (also known as QGC001), a prodrug of the specific and selective APA inhibitor, EC33. When given orally, RB150 is able to cross the blood-brain-barrier, to inhibit brain APA activity and to decrease BP in two experimental models of HTA. We investigate here the effects of repeated oral administrations of RB150 (50 mg/kg/day) over 10 days on BP in conscious DOCA-salt rats to assess if a tolerance to the antihypertensive RB150 effect occurs after chronic treatment. Design and method: For this purpose, 5 hours after RB150 administration, we measured daily variations in systolic BP (sBP) by the tail-cuff method, plasma arginine-vasopressin (AVP) levels by radioimmunoassay and metabolic parameters Results: Chronic oral treatment with RB150 significantly decreased sBP compared with saline over the 10 days treatment period, without significantly altering heart rate. The maximal decrease in sBP (−29.6 ± 6 mmHg) was observed on the second day and the average RB150-induced decrease in sBP over 10 days was not significantly changed (−21 ± 5 mmHg), showing no tolerance to the antihypertensive effect of RB150 after repeated administrations. Chronic oral treatment with RB150 reduced by 59% the increase in plasma AVP levels observed in DOCA-salt rats compared with normotensive rats receiving saline. The administration of RB150 increased significantly diuresis by 39% and natriuresis by 34% without modification of plasma sodium and potassium levels when compared to DOCA-salt rats receiving saline over a 10 days analysis. This may participate to the blood pressure decrease by reducing the size of fluid compartment. Conclusions: Our results further confirm the interest of developing RB150 as a novel centrally-acting agent for the treatment of HTA.