Sodium-depleted Marmosets Research Articles

Structural Modification of the P2‘ Position of 2,7-Dialkyl-Substituted 5(S)-Amino-4(S)-hydroxy-8-phenyl-octanecarboxamides: The Discovery of Aliskiren, a Potent Nonpeptide Human Renin Inhibitor Active after Once Daily Dosing in Marmosets

Due to its function in the rate limiting initial step of the renin-angiotensin system, renin is a particularly promising target for drugs designed to control hypertension, a growing risk to health worldwide. Despite vast efforts over more than two decades, no orally efficacious renin inhibitor had reached the market. As a result of a structure-based topological design approach, we have identified a novel class of small-molecule inhibitors with good oral blood-pressure lowering effects in primates. Further lead optimization aimed for improvement of in vivo potency and duration of action, mainly by P2' modifications at the hydroxyethylene transition-state isostere. These efforts resulted in the discovery of aliskiren (46, CGP060536B, SPP100), a highly potent, selective inhibitor of renin, demonstrating excellent efficacy in sodium-depleted marmosets after oral administration, with sustained duration of action in reducing dose-dependently mean arterial blood pressure. Aliskiren has recently received regulatory approval by the U.S. Food and Drug Administration for the treatment of hypertension.

Novel 2,7-Dialkyl-Substituted 5(S)-Amino-4(S)-hydroxy-8-phenyl-octanecarboxamide Transition State Peptidomimetics Are Potent and Orally Active Inhibitors of Human Renin

The action of renin is the rate-limiting step of the renin-angiotensin system (RAS), a key regulator of blood pressure. Effective renin inhibitors directly block the RAS entirely at source and, thus, may provide a vital weapon for hypertension therapy. Our efforts toward identifying novel small-molecule peptidomimetic renin inhibitors have resulted in the design of transition-state isosteres such as 1 bearing an all-carbon 8-phenyl-octanecarboxamide framework. Optimization of the extended P3 portion of 1 and extensive P2' modifications provided analogues with improved in vitro potencies in the presence of plasma. X-ray resolution of rh-renin/38a in the course of SAR work surprisingly unveiled the exploitation of a previously unexplored pocket (S3sp) important for strong binding affinities. Several inhibitors demonstrated oral efficacy in sodium-depleted marmosets. The most potent, 38a, induced dose-dependently a pronounced reduction in mean arterial blood pressure, paralleled by complete blockade of active plasma renin, up to 8 h post-dose. Oral bioavailability of 38a was 16% in marmosets.

Novel therapy in the management of hypertension: Preliminary experience with renin inhibitors

Suppressing the renin-angiotensin system (RAS) has become a crucial strategy for combating cardiovascular and renal disease—one that has been validated by the clinical success of angiotensin-converting enzyme inhibitors and angiotensin II (Ang II) receptor blockers (ARBs). Yet these agents may be limited in their ability to completely inhibit the RAS long-term. Because renin acts early in the RAS cascade, inhibiting the RAS at this step has long been considered a logical therapeutic target. Efforts to develop renin inhibitors have been underway for decades, but poor bioavailability, short duration of action, and high production costs have impeded their clinical development. Designed using molecular modeling techniques, aliskiren is the first agent in a new class of nonpeptide, low-molecular weight, orally active renin inhibitors. Aliskiren has demonstrated potent in vitro affinity for human renin, and has exhibited dose-dependent reductions in plasma renin activity (PRA), Ang II levels, and blood pressure (BP) in preclinical models. In sodium-depleted marmosets, aliskiren showed dose-dependent reductions in BP, and greater BP reductions than the previous-generation renin inhibitors, remikiren and zankiren. In spontaneously hypertensive rats, aliskiren 30 mg/kg/d yielded modest BP reductions; when administered in combination with benazepril or valsartan, however, BP reductions appeared synergistic. In the double-transgenic rat model, which expresses human renin and angiotensinogen, and becomes severely hypertensive and dies of organ failure if left untreated, 9 weeks of therapy with either aliskiren or valsartan significantly reduced left ventricular wall thickness and cardiac hypertrophy. Furthermore, both agents similarly reduced albuminuria and renal and cardiac fibrosis, as well as kidney and heart macrophage and T-cell accumulation. All untreated rats died by Week 8. The pharmacokinetics and pharmacodynamics of aliskiren were examined in 18 volunteers in a double-blind, 3-way crossover trial. During 3 periods of 8 days, each followed by a 6-day washout, each volunteer was randomized to receive 2 dosages of aliskiren (40 mg and 80 mg/d, or 160 mg and 640 mg/d) and either placebo or 20 mg/d enalapril. Aliskiren reduced PRA and angiotensin I (Ang I) and Ang II levels, while PRA and Ang I were increased with enalapril. The half-life of aliskiren was 29 hours. Aliskiren efficacy and safety were evaluated in a randomized, double-blind trial in 226 patients, aged 21–70 years, with mild-to-moderate hypertension. Patients received aliskiren (37.5 mg, 75 mg, 150 mg, or 300 mg) or losartan (100 mg) once daily for 4 weeks. Aliskiren yielded dose-dependent reductions in systolic and diastolic BP; at the 300-mg/d dose, the mean BP reduction was comparable to that of high-dose losartan. Losartan and all aliskiren doses were equally well tolerated. Aliskiren was also studied in a double-blind, placebo-controlled, 8-week trial of 652 patients with mild-to-moderate hypertension. Patients were randomized to once-daily aliskiren (150 mg, 300 mg, or 600 mg), irbesartan 150 mg, or placebo. The antihypertensive efficacy of aliskiren 150 mg/d was comparable to that of irbesartan; the reduction in diastolic BP with aliskiren 300 mg/d was significantly greater than with irbesartan. Tolerability with aliskiren up to 600 mg/d was similar to that with irbesartan. A double-blind, randomized, placebo-controlled, 4-period crossover study examined the effects of single doses of aliskiren 300 mg, valsartan 160 mg, and their combination (aliskiren 150 mg plus valsartan 80 mg) in 12 mildly sodium-depleted, normotensive, male subjects. As expected, PRA, Ang I, and Ang II were significantly reduced with aliskiren and increased with valsartan. However, the ARB-associated increases in PRA and Ang I and Ang II levels were eliminated with the addition of aliskiren to valsartan. Aliskiren thus appears to be a useful addition for the management of hypertension. Further investigation of the effects of aliskiren on end-organ protection is needed.

Analysis of Recent Papers in Hypertension Jan Basile, MD, Senior Editor

Analysis of Recent Papers in Hypertension Jan Basile, MD, Senior Editor

Aliskiren, a novel, orally effective renin inhibitor, lowers blood pressure in marmosets and spontaneously hypertensive rats

Aliskiren is a new renin inhibitor of a novel structural class that has recently been shown to be efficacious in hypertensive patients after once-daily oral dosing. We report the results of animal experiments performed in marmosets and rats in order to characterize aliskiren before its recent investigation in humans. The effects of aliskiren were investigated in sodium-depleted marmosets (oral dosing) and in spontaneously hypertensive rats (dosing via subcutaneous osmotic minipumps). Blood pressure (BP) and heart rate were measured by radiotelemetry. In sodium-depleted marmosets, single oral doses of aliskiren (1-30 mg/kg) dose-dependently lowered BP. At a dose of 3 mg/kg, peak effects were observed 1 h after dosing (-30 +/- 4 mmHg, n = 6) and the response persisted for more than 12 h. A single oral dose of 3 mg/kg aliskiren was more effective than the same dose of either remikiren or zankiren, two orally active renin inhibitors previously tested in humans. Aliskiren (10 mg/kg) was at least as effective as equal doses of the AT1-receptor blocker valsartan or the angiotensin-converting enzyme inhibitor benazepril. In spontaneously hypertensive rats, aliskiren dose-dependently (10-100 mg/kg per day) decreased BP. Aliskiren also potentiated the antihypertensive effects of low doses of valsartan or benazeprilat (1 or 3 mg/kg per day). Aliskiren is an orally effective, long-lasting renin inhibitor that shows antihypertensive efficacy in animals superior to previous renin inhibitors and at least equivalent to angiotensin-converting enzyme inhibitors and AT1-receptor blockers. Aliskiren may therefore represent an effective, novel approach to the treatment of hypertension and related disorders, alone or in combination with other antihypertensive agents.

Structure-based design of aliskiren, a novel orally effective renin inhibitor

Hypertension is a major risk factor for cardiovascular diseases such as stroke, myocardial infarction, and heart failure, the leading causes of death in the Western world. Inhibitors of the renin–angiotensin system (RAS) have proven to be successful treatments for hypertension. As renin specifically catalyses the rate-limiting step of the RAS, it represents the optimal target for RAS inhibition. Several peptide-like renin inhibitors have been synthesized previously, but poor pharmacokinetic properties meant that these compounds were not clinically useful. We employed a combination of molecular modelling and crystallographic structure analysis to design renin inhibitors lacking the extended peptide-like backbone of earlier inhibitors, for improved pharmacokinetic properties. This led to the discovery of aliskiren, a highly potent and selective inhibitor of human renin in vitro, and in vivo; once-daily oral doses of aliskiren inhibit renin and lower blood pressure in sodium-depleted marmosets and hypertensive human patients. Aliskiren represents the first in a novel class of renin inhibitors with the potential for treatment of hypertension and related cardiovascular diseases.

Preclinical and clinical profile of Valsartan, a selective angiotensin II type-1 receptor blocker

Valsartan (Diovan) is a potent, orally active, specific, and highly selective blocker for the AT1-receptor subtype. The antihypertensive effects of oral treatment with valsartan were preclinically demonstrated in the sodium-depleted marmosets, renal hypertensive rats (2K1C), spontaneous hypertensive rats (SHR) and stroke-prone SHR. Moreover, valsartan had protective effects against hypertensive end-organ damage such as cardiac hypertrophy and renal disease. In an investigation of pharmacokinetics and pharmacodynamics in normotensive male volunteers, valsartan was rapidly absorbed with the maximal plasma concentration occurring 2-3 h after oral administration. The elimination half-life was about 4-6 h, valsartan was poorly metabolized, and most of the drug was excreted via feces. Valsartan produced persistent reductions of blood pressure in patients with mild to moderate essential hypertension and had a good safety profile with a wide therapeutic window between the effective pharmacological doses and the toxic doses. Therefore, valsartan is expected to be a safe and effective antihypertensive agent for the treatment of essential and severe hypertension.

Novel low molecular renin inhibitors which show good oral blood pressure lowering effects in marmosets

A series of low molecular renin inhibitors (molecular weight ∗< 600) were prepared and their biological activities were evaluated. The inhibitors containing indole-2-carbonyl moieties at the P3 position and 2-amino-3,5- anti -diol fragment at the P1-P1′ position showed renin inhibitory activities with IC 50 of 10 −8 M order and good blood pressure lowering effects after oral administration to sodium-depleted marmosets. A series of low molecular renin inhibitors (molecular weight ∗< 600) were prepared and their biological activities were evaluated.

Pharmacological profile of valsartan: a potent, orally active, nonpeptide antagonist of the angiotensin II AT1‐receptor subtype

1. The pharmacological profile of valsartan, (S)-N-valeryl-N-([2'-(1H-tetrazol-5-yl)biphenyl-4-yl]-methyl)-vali ne, a potent, highly selective, and orally active antagonist at the angiotensin II (AII) AT1-receptor, was studied in vitro and in vivo. 2. Valsartan competed with [125I]-AII at its specific binding sites in rat aortic smooth muscle cell membranes (AT1-receptor subtype) with a Ki of 2.38 nM, but was about 30,000 times less active in human myometrial membranes (AT2-receptor subtype). 3. In rabbit aortic rings incubated for 5 min with valsartan, at concentrations of 2, 20 and 200 nM, the concentration-response curve of AII was displaced to the right and the maximum response was reduced by 33%, 36% and 40%, respectively. Prolongation of the incubation time with valsartan to 1 h or 3 h, further reduced the maximum response by 48% or 59% (after 20 nM) and by 59% or 60% (after 200 nM) respectively. After 3 h incubation an apparent pKb value of 9.26 was calculated. Contractions induced by noradrenaline, 5-hydroxytryptamine, or potassium chloride were not affected by valsartan. No agonistic effects were observed in the rabbit aorta at concentrations of valsartan up to 2 microM. 4. In bovine adrenal glomerulosa, valsartan inhibited AII-stimulated aldosterone release without affecting the maximum response (pA2 8.4). 5. In the pithed rat, oral administration of valsartan (10 mg kg-1) shifted the AII-induced pressor response curves to the right, without affecting responses induced by the electrical stimulation of the sympathetic outflow or by noradrenaline. Animals treated with valsartan 24 h before pithing also showed significant inhibition of the response to AII. 6. In conscious, two-kidney, one-clip renal hypertensive rats (2K1C), valsartan decreased blood pressure in a dose-dependent manner after single i.v. or oral administration. The respective ED30 values were 0.06 mg kg-1 (i.v.) and 1.4 mg kg-1 (p.o.). The antihypertensive effect lasted for at least 24 h after either route of administration. After repeated oral administration for 4 days (3 and 10 mg kg-1 daily), in 2K1C renal hypertensive rats, systolic blood pressure was consistently decreased, but heart rate was not significantly affected. 7. In conscious, normotensive, sodium-depleted marmosets, valsartan decreased mean arterial pressure, measured by telemetry, after oral doses of 1-30 mg kg-1. The hypotensive effect persisted up to 12 h after 3 and 10 mg kg-1 and up to 24 h after 30 mg kg-1. 8. In sodium-depleted marmosets, the hypotensive effect of valsartan lasted longer than that of losartan(DuP 753). In renal hypertensive rats, both agents had a similar duration (24 h), but a different onset of action (valsartan at 1 h, losartan between 2 h and 24 h).9. These results demonstrate that valsartan is a potent, specific, highly selective antagonist of AII at theAT1-receptor subtype and does not possess agonistic activity. Furthermore, it is an efficacious, orally active, blood pressure-lowering agent in conscious renal hypertensive rats and in conscious normotensive,sodium-depleted primates.

The rennin???angiotensin system

We investigated the effects of the human renin inhibitor, ES-8891, and captopril on renal renin secretion and storage in the marmoset. Either ES-8891 (30mg/kg) or captopril (2 mg/kg) was given orally twice a day for 1 week to conscious, sodium-depleted marmosets (n=6 for each group). The ES-8891-treated group displayed a significant reduction in mean arterial pressure (MAP), plasma renin activity (PRA) and plasma immunoreactive renin (PIR) compared with the control group. Kidney renin content was significantly increased compared with the control group and enlarged renin granules containing heterogenous internum were observed in juxtaglomerular cells after treatment with ES-8891. Treatment with captopril significantly increased PRA and PIR compared with the control day as well as increasing kidney renin content and the number of renin granules with crystalline content in juxtaglomerular cells compared with the control group. These results suggest that ES-8891 inhibits both PRA and renin secretion from the kidney, resulting in an increase in renal renin storage

Kidney renin gene expression after renin inhibition in the marmoset.

1. The effects of renin inhibitor ES-8891 on renin synthesis and its secretion by the kidney were investigated in normotensive sodium-depleted marmosets. We measured plasma renin activity, plasma immunoreactive renin concentration, plasma angiotensin II concentration and kidney renin mRNA content after oral administration of ES-8891 (60 mg day-1 kg-1) for 1 week. 2. The mean blood pressure was significantly decreased (P less than 0.01) on day 7 after oral administration of ES-8891. There was no significant change in heart rate during the administration. 3. Oral administration of ES-8891 for 1 week markedly decreased the plasma renin activity, the plasma immunoreactive renin concentration and the plasma angiotensin II concentration (to 18%, 41% and 24% of the corresponding control values; P less than 0.05 for each, n = 5). 4. The kidney renin mRNA content in ES-8891-treated marmosets was significantly lower than that in normal controls (4.2 +/- 3.5 versus 12.8 +/- 5.5 pg/micrograms of total RNA, means +/- SD, P less than 0.05, n = 5). 5. Oral administration of the renin inhibitor ES-8891 for 1 week not only inhibited plasma renin activity but also decreased renin synthesis and its secretion by the kidney.

Ro 42-5892 is a potent orally active renin inhibitor in primates.

The goal of the present study was to characterize the new renin inhibitor Ro 42-5892 in vitro and in vivo. In vitro, Ro 42-5892 inhibited purified human renin and human plasma renin specifically with an IC50 of 0.7 nM and 0.8 nM, respectively. In vivo, Ro 42-5892 reduced mean arterial blood pressure in sodium-depleted marmosets and squirrel monkeys with as low a dose as 0.1 mg/kg orally. Higher doses reduced pressure by 30-35 mm Hg in both species. The duration of blood pressure decrease with 3 mg/kg orally was more than 24 hours. Maximal changes of plasma renin activity, immunoreactive angiotensin I, and immunoreactive angiotensin II were observed at 15 minutes. Renin was reduced by 74 +/- 31%, angiotensin I by 85 +/- 14%, angiotensin II by 89 +/- 17%, and immunoreactive active renin was increased by 70 +/- 39%. However, unlike pressure, these maximal effects were only transient with complete recovery of renin at 60 minutes under still reduced levels of angiotensin I (61 +/- 24%) and angiotensin II (71 +/- 38%) and increased concentrations of active renin (86 +/- 30%). The blood pressure lowering was due to specific renin inhibition as exemplified by the influence of the kidney, sodium status, species, or stereoselectivity. Moreover, the reduction of arterial blood pressure was similar to the action of the angiotensin converting enzyme inhibitor cilazapril and was not associated with reflex tachycardia in contrast to the pure vasodilator minoxidil. We conclude that Ro 42-5892 is a potent orally active renin inhibitor acting mainly by inhibition of renin in an extraplasmatic compartment.

1,2,4-Triazolo[4,3-a]pyrazine derivatives with human renin inhibitory activity. 3. Synthesis and biological properties of aminodeoxystatine and difluorostatone derivatives

Two series of 1,2,4-triazolo[4,3-a]pyrazine derivatives with human renin inhibitory activity have been synthesized which incorporate the transition-state mimetics (3S,4S)- and (3R,4S)-5-cyclohexyl-3,4-diaminopentanoic acid ((S)- and (R)-CDAPA), and (4S)-4-amino-5-cyclohexyl-2,2-difluoro-3-oxopentanoic acid (ACDFOPA). Several compounds in these series, for example 13a, 19c, and 19f, were highly potent inhibitors of partially purified human renin (IC50 values of 3.9, 1.6, and 1.4 nM, respectively). The ACDFOPA-based compounds 19c and 19f contain no natural amino acid fragments and have molecular weights which compare well with those of previously reported inhibitors of nanomolar in vitro potency. When administered intravenously to anesthetized, sodium-depleted marmosets at doses of 3 mg/kg, compounds 13a and 19c caused a marked reduction in mean arterial pressure, but in the same animal model at 30 mg/kg, oral activity was not seen.

The effect of the renin inhibitor ES-1005 on the expression of the kidney renin gene in sodium-depleted marmosets

The effect of the renin inhibitor ES-1005 or captopril on the expression of the kidney renin gene was investigated in sodium-depleted marmosets. We measured the level of kidney renin messenger RNA (mRNA) after continuous administration of ES-1005 (48 mg/kg per day) or captopril (2 mg/kg per day) intraperitoneally, via an osmotic mini-pump, for one week. The level of kidney renin mRNA was measured by densitometric Northern blot analysis using an alpha-32P-labelled human renin cDNA fragment as the hybridization probe. Captopril treatment markedly increased plasma renin activity and the level of kidney renin mRNA by 4.7-fold and 6.3-fold, respectively. ES-1005 treatment completely inhibited plasma renin activity and significantly decreased the level of kidney renin mRNA (46% of the normal control P less than 0.01). However, plasma immunoreactive renin concentration was significantly increased by the treatment with ES-1005 (P less than 0.05). These results suggest that the treatment with the renin inhibitor ES-1005 for one week has a paradoxical effect on kidney renin gene expression and renin release from the kidney in sodium-depleted marmosets.

1,2,4-Triazolo[4,3-a]pyrazine derivatives with human renin inhibitory activity. 2. Synthesis, biological properties and molecular modeling of hydroxyethylene isostere derivatives

A series of inhibitors of human renin have been synthesized, derived from combination of a 2-(8-propyl-6-pyridin-3-yl-1,2,4-triazolo[4,3-a]pyrazin-3-yl)- 3-pyridin- 3-ylpropionic acid moiety 6c with the hydroxyethylene isostere of the scissile amide bond (2S,4S,5S)-5-amino-6-cyclohexyl-4-hydroxy-2-isopropylhexanoic acid (ChaOH--Val). The more potent members of this series showed good inhibitory activity against partially purified human renin, 7d, for example, having an IC50 of 0.2 nM. Structure-activity relationships for these compounds were consistent with their binding to the S4-S2' sites of human renin. Analogues 7e and 7h-k with a variety of substituents at the C-terminus all had in vitro IC50S less than 1 nM. In contrast with the majority of previously reported inhibitors of similar potency, these compounds contain no natural amino acid fragments. When administered intravenously to anesthetized, sodium-depleted marmosets at doses of 0.3-3.0 mg/kg, compound 7d caused a marked reduction in mean arterial pressure. Following oral administration at 30 mg/kg in the same animal model, 7d again elicited a significant fall in mean arterial pressure, accompanied by suppression of plasma renin activity lasting up to 3 h after dosing.

ES-8891, an orally active inhibitor of human renin.

A newly synthesized orally active renin inhibitor, N-morpholinoacetyl-(1-naphthyl)-L-alanyl-(4-thiazolyl)-L-alanyl (3S,4S)-4-amino-3-hydroxy-5-cyclohexylpentanoyl-n-hexylamide (ES-8891), was found to be a highly potent competitive inhibitor of human renin with an inhibition constant of 1.1 nM. This inhibitor was also active against monkey renin, although there was less inhibition of renin in pig, rabbit, and rat. ES-8891 did not inhibit cathepsin D, pepsin, trypsin, chymotrypsin, angiotensin converting enzyme, and urinary kallikrein at a concentration of 10(-5) M. A single oral administration of ES-8891 (10 or 30 mg/kg) to conscious, sodium-depleted marmosets caused a dose-related decrease in plasma renin activity and blood pressure. ES-8891 (30 mg/kg) produced an 80% inhibition of plasma renin activity, which lasted for more than 6 hours. Kidney renin messenger RNA was not significantly changed 6 hours after oral administration of ES-8891 (30 mg/kg). A single oral administration of 240 mg ES-8891 to healthy human volunteers (n = 6) produced a significant inhibition of plasma renin activity (75% inhibition at 0.5 and 1 hour, 50% inhibition at 2 hours) with a good correlation of plasma levels of ES-8891. There were no significant changes in blood pressure or heart rate, and no adverse effects were observed. These results suggest that ES-8891 is an orally active human renin inhibitor that may be clinically useful.

Effect of renin inhibitor (ES-1005) on expression of the kidney renin gene in sodium-depleted marmosets

Effect of renin inhibitor ES-1005 or captopril on the renin synthesis by the kidney was investigated in sodium-depleted marmosets. The level of kidney renin mRNA was measured after 2-hour (acute study) and one-week (chronic study) administrations of the two agents. Relative amounts of kidney renin mRNA were measured by densitometric Northern blot analysis using an alpha-32P-labelled human renin cDNA fragment as a hybridization probe. In the acute study, treatment with captopril significantly increased plasma renin activity (PRA) (p less than 0.05), but did not change the level of kidney renin mRNA. In the chronic study, treatment with captopril markedly increased PRA as well as the level of kidney renin mRNA (4.7-fold and 6.3-fold increases, respectively). In contrast, treatment with ES-1005 completely inhibited PRA and significantly suppressed the level of kidney renin mRNA in both acute and chronic studies (about one-third of the normal control, p less than 0.05). These results suggest that renin inhibitor ES-1005 not only inhibits plasma renin activity but also suppresses the synthesis of renin by the kidney.

Syntheses and biological activities of renin inhibitors containing statine analogues.

Syntheses and biological activities of dipeptide renin inhibitors that contain statine analogues are described. The key steps of the synthetic approach to dipeptide renin inhibitors are the asymmetric synthesis of 2(R)-substituted-3-aminocarbonylpropionic acids and the diastereoselective syntheses of (3S,4S)-statine analogues. These inhibitors (2,14-40) inhibited human renin in the 3-140 nM range. Inhibitor ES 6864 (2) was found to be a highly potent inhibitor of human renin (IC50: 4.6 x 10(-9) M) and showed high enzyme specificity. Oral administration of ES 6864 at 3 mg/kg to conscious, sodium-depleted marmosets inhibited plasma renin activity (PRA) more than 80% after 1 h.

Effects of renin inhibitors on the expression of kidney renin gene and tissue renin-like activity.

The effect of renin inhibitor ES-1005 on renin gene expression was investigated in sodium-depleted marmosets. The kidneys were removed after continuous infusion of ES-1005 (12 mg/kg per h) for 2 h. The relative amount of kidney renin messenger (m)RNA was measured by densitometric Northern blot analysis using an alpha-32P-labelled human renin complementary (c)DNA fragment as a hybridization probe. Plasma renin activity was completely inhibited by ES-1005. The level of kidney renin mRNA decreased significantly to about one-third of the normal control value. We also investigated the inhibitory potency of the renin inhibitor ES-6864 on the renin-like activity in dog tissues (adrenal glands, aorta and brainstem). ES-6864 inhibited the tissue renin-like activity with an IC50 of 10(-7) to 10(-8) mol/l in vitro. Renin inhibitors not only inhibit the activity of plasma and tissue renin, but also suppress the synthesis of renin in the kidney.

Haemodynamic effects of acute and chronic renin inhibition in marmosets

We have developed marmoset models for the in vivo evaluation of primate-specific inhibitors of human renin. After acute intravenous administration to normotensive sodium-depleted marmosets, renin inhibitors of different structural types induced a maximum hypotensive response of a magnitude similar to that induced after angiotensin converting enzyme (ACE) inhibition. The response was prevented by pretreatment with an ACE inhibitor. A close relationship between the inhibition of plasma renin activity (PRA) and the fall in blood pressure was observed with most of the inhibitors. CGP 29,287, a synthetic renin inhibitor, and R-3-36-16, a monoclonal antibody, both induced a selective increase in renal blood flow similar to that induced by an ACE inhibitor. A sustained reduction in blood pressure was observed during continuous administration of CGP 29,287 or R-3-36-16 over 14 days, despite an increase in immunoreactive renin and an apparent recovery of PRA. A similar blood pressure fall and an increase in plasma renin was observed during continuous administration of an ACE inhibitor. The renin inhibitor CGP 29,287 also lowered blood pressure after acute administration to hypertensive marmosets with normal PRA. Our studies demonstrate that renin inhibitors have similar haemodynamic effects to ACE inhibitors, and indicate that they may have a similar antihypertensive efficacy.