WO2004075892A2 - Combination therapy for hypertension using lercanidipine and an angiotensin ii receptor blocker - Google Patents

Abstract

Combination Therapy for Hypertension using Lercanidipine and an Angiotensin II Receptor Blocker Lercanidipine is used in the preparation of a medicament for the treatment of hypertension in combination with the prior, concurrent or post-administration of an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan, and optionally in further combination with the prior, concurrent or post­administration of a diuretic such as hydrochlorothiazide. Compositions containing lercanidipine and the ARB (or lercanidipine, the ARB and a diuretic) are claimed.

Description

Combination Therapy for Hypertension using Lercanidipine and an Angiotensin II Receptor Blocker

DESCRIPTION

The invention relates to the use of lercanidipine and an angiotensin II receptor blocker (ARB) in combination therapy for the freatment of hypertension, including that in patients also suffering from tachycardia. A diuretic may also be included in the combination therapy.

Hypertension

Hypertension is one of the most common cardiovascular disease states. In the United States, over 50 million people have been diagnosed with hypertension. Elevated arterial pressure can cause pathological changes in the vasculature and hypertrophy of the left ventricle. Due to the damage that can be produced by hypertension, it is proposed to be the principal cause of stroke, myocardial infarction, and sudden cardiac death. Additionally, it is believed to be a major contributor to cardiac failure, renal insufficiency, and dissecting aneurysm of the aorta.

Tachycardia

Tachycardia refers to an increased heart rate, i.e., above the normal range of about 60-100 beats per minute electrically regulated by the sino-afrial (SA) node of the right atrium. Abnormal heart rates can range from about 100-400 beats per minute and can be life threatening. Tachycardia can arise from atrial fibrillation (AF) in which the heartbeat is irregular and rapid due to the upper chambers, or atria, beating about four times faster than normal. AF can lead to other rhythm problems, chronic fatigue and congestive heart failure. By contrast, ventricular tachycardia (VT) refers to sudden rapid heartbeats originating in the ventricles. When VT occurs, the ventricles may not be able to fill with enough blood to supply the body with sufficient amounts of oxygen rich blood. Symptoms of VT include feeling faint, passing out, dizziness, or a pounding in the chest. The most common electrical therapy for VT is implantation of a device known as an Implantable Cardioverter Defibrillator or ICD. The ICD applies an electric shock to the heart muscle to interrupt or disrupt the fast rhythm. The electric shock may be in the form of specially timed pacemaker pulses (unfelt by the patient) or by high voltage shock.

VT can lead to ventricular fibrillation, or VF, which is characterized by irregular and chaotic rapid heartbeats. Because the fibrillating ventricular muscle cannot contract and pump blood to the brain and vital organs, VF is the number one cause of sudden cardiac death. Without immediate emergency treatment of an electric shock to restore normal rhythm, an individual loses consciousness within seconds and dies within minutes. Hypertension and cardiac arrhythmias commonly coexist in many patients, and both need to be managed appropriately. Furthermore, hypertensive left ventricular hypertrophy could cause a wide variety of ventricular arrhythmias, which could end in sudden cardiac arrest (Greenberg et al., J Clin Hypertens (Greenwich) 2000; 2(1): 14-19; Hennersdorf et al., J Hypertens. 2001; 19(2): 167-77. A side effect of some dihydropyridine calcium channel blockers can be reflex tachycardia. Tachycardia occurred in about 0-9% of patients who were administered lercanidipine, and the tachycardia manifested either at the beginning of treatment, or during dose escalation, disappearing upon cessation of medication. However, of the calcium channel blockers, lercanidipine showed one of the lowest, if not the lowest, incidence of tachycardia as a side effect. Sustained-release administration, such as in a depot formulation, was also shown to abrogate this side effect (Sada et al., Nippon Yakurigaku Zasshi. 2003; 122(6): 539-47; Harada et al., Circ. J. 2003; 67(2): 139- 45; and van Zweiten, Blood Press. Suppl. 1998; 2:5-9).

Accordingly, there is a need in the art to mitigate the tachycardia associated with hypertension, or associated with dihydropyridine calcium channel blockers used to treat hypertension, since tachycardia or other arrhythmia can result in cardiac events and death.

Angiotensin II receptor blockers The renin-angiotensin-aldosterone system is an important regulator of arterial pressure. The inactive angiotensinogen peptide is converted to the pro-peptide angiotensin I by the enzyme renin. Angiotensin I then is converted to the active angiotensin II form by the angiotensin converting enzyme. Angiotensin II then acts through a variety of receptor mediated mechanisms, such as increasing the total peripheral resistance and inhibiting the excretion of sodium and water by the kidneys, to increase arterial pressure. Angiotensin II receptor blockers (ARBs) are a class of active agents used in the treatment of hypertension. ARBs block the vasoconstrictor and aldosterone-secreting effects of angiotensin II by selectively blocking the binding of angiotensin II to the AT_t receptor in many tissues, such as smooth muscle and the adrenal gland. ARBs include irbesartan, valsartan, telmisartan, olmesartan medoxomil ("olmesartan"), losartan potassium ("losartan") and eprosartan mesylate ("eprosartan"). (Physicians' Desk Reference, 57^th edition, 2003).

The recommended daily dosages of ARBs when used as monotherapy for the freatment of hypertension are as follows: irbesartan - 150 to 300 mg; valsartan - 80 to 320 mg; telmisartan - 40 to 80 mg; olmesartan - 20 to 40 mg; losartan - 25 to 100 mg, and eprosartan - 400 to 800 mg.

ARBs are commercially available. Irbesartan is available from Bristol-Myers Squibb and Sanofi-Synthelabo under the trade name Avapro^® in dosages of 75, 150 and 300 mg. Valsartan is available from Novartis under the trade name Diovan^® in dosages of 40, 80, 160 and 320 mg. Telmisartan is available from Boehringer Ingelheim under the trade name Micardis^® in dosages of 20, 40 and 80 mg. Olmesartan is available from Sankyo under the trade name Benicar^® in dosages of 5, 20 and 40 mg. Losartan is available from Merck under the trade name Cozaar^® in dosages of 25, 50 and 100 mg. Eprosartan is available from Bioavail under the trade name Teveten^® in dosages of 400 and 600 mg. Olmesartan, 2,3-dihydroxy-2-butenyl 4-(l -hydroxy- l-methylethyl)-2-propy 1-1-

[p-(o-lH-tetrazol-5-ylphenyl)-benzyl]-imidazole-5-carboxylate cyclic 2,3 -carbonate, is an angiotensin II receptor (AT! subtype) antagonist described in US 5616599. Following oral administration, peak serum concentrations of olmesartan occur within about 1-2 hours. There is virtually no metabolism of olmesartan, approximately 35 to 50% is cleared in the urine and the remainder is eliminated in faeces. Olmesartan is a specific competitive antagonist of the ATj receptor with a much greater affinity (more than 12,500-fold) for the AT] receptor compared to the AT₂ receptor and no agonist activity. Blockade of the ATj receptor removes the negative feedback of angiotensin II on rerun secretion. The resulting increase in plasma renin activity and circulating angiotensin II, however, does not overcome the positive effects of olmesartan for treating hypertension. Olmesartan does not inhibit ACE or renin or affect other hormone receptors or ion channels known to be involved in regulation of blood pressure.

The recommended starting dosage of olmesartan as monotherapy for essential hypertension is 20 mg once per day, with drug tifration to 40 mg per day. Several weeks of therapy may be required to achieve optimal blood pressure reduction for a patient. A lower initial dose is recommended in patients with depletion of infravascular volume or salt. No dosage adjustment is necessary in elderly patients or in patients with hepatic impairment or mild to severe renal impairment. Safety and effectiveness have not been established in children. Treatment with olmesartan is well tolerated with an incidence of adverse events similar to placebo.

Irbesartan, 2-butyl-3-[p-(o-lH-tetrazol-5-ylphenyl)-benzyl]-l,3-diazaspiro[4.4] non-l-en-4-one, is an angiotensin II receptor (AT] subtype) antagonist described in US 5270317. Following oral administration, peak serum concentrations of irbesartan occur within about 1.5-2 hours. Irbesartan is metabolized via glucuronide conjugation and oxidation. Irbesartan is a specific competitive antagonist of the ATj receptor with a much greater affinity (more than 8500-fold) for AT] receptor compared to AT₂ receptor and no agonist activity. Blockade of the AT] receptor removes the negative feedback of angiotensin II on renin secretion. The resulting increase in plasma renin activity and circulating angiotensin II, however, does not overcome the positive effects of irbesartan for treating hypertension. Irbesartan does not inhibit ACE or renin or affect other hormone receptors or ion channels known to be involved in regulation of blood pressure.

The recommended starting dosage of irbesartan as monotherapy for essential hypertension is 150 mg once per day, with drug tifration to 300 mg per day. Several weeks of therapy may be required to achieve optimal blood pressure reduction for a patient. A lower initial dose of 75 mg is recommended in patients with depletion of infravascular volume or salt. No dosage adjustment is necessary in elderly patients or in patients with hepatic impairment or mild to severe renal impairment. Safety and effectiveness have not been established in children under 6 years old. Children 6-12 years old may reasonably be started on a dosage of 75 mg once daily, with tifration to 150 mg daily. Treatment with irbesartan is well tolerated with an incidence of adverse events similar to placebo.

Losartan, 2-butyl-4-chloro- 1 - p-(o- 1 H-tetrazol-5-ylphenyl)-benzyl]-imidazole- 5-methanol monopotassium salt, is an angiotensin II receptor (type AT]) antagonist, described in US 5138069. Oxidation of the 5-hydroxymethyl group on the imidazole ring results in the active metabolite of losartan. Mean peak concentrations of losartan and its active metabolite are reached in 1 hour and in 3-4 hours, respectively. Losartan is an orally active agent that undergoes substantial first-pass metabolism by cytochrome P450 enzymes. It is converted, in part, to an active carboxylic acid metabolite that is responsible for most of the angiotensin II receptor antagonism that follows losartan treatment. The terminal half-life of losartan is about 2 hours and of the metabolite is about 6-9 hours. Both losartan and its principal active metabolite do not exhibit any partial agonist activity at the AT] receptor and have much greater affinity (about 1000- fold) for the AT] receptor than for the AT₂ receptor. In vitro binding studies indicate that losartan is a reversible competitive inhibitor of the AT] receptor. The active metabolite is 10 to 40 times more potent by weight than losartan and appears to be a reversible, non-competitive inhibitor of the AT] receptor. Neither losartan nor its active metabolite inhibits ACE ; nor do they bind to or block other hormone receptors or ion channels known to be important in cardiovascular regulation. The usual starting dose of losartan is 50 mg once daily. The effect of losartan is substantially present within one week but in some studies the maximal effect occurred in 3-6 weeks. No initial dosage adjustment is necessary for elderly patients or for patients with renal impairment, including patients on dialysis. Safety and effectiveness in pediatric patients have not been established. The overall incidence of adverse experiences reported with losartan potassium was similar to placebo.

Valsartan, N-(l-oxopentyl)-N-[p-(o-lH-tefrazol-5-ylphenyl)-benzyl]-L-valine, is a nonpeptide, orally active, and specific angiotensin II antagonist acting on the ATj receptor subtype, described in US 5399578. Following oral administration, peak plasma concentration is reached 2 to 4 hours after dosing. . The recovery is mainly in feaces as unchanged, drug, with only about 20% of dose recovered as metabolites. The primary metabolite, accounting for about 9% of dose, is valeryl 4-hydroxy valsartan. The enzyme(s) responsible for valsartan metabolism have not been identified but do not seem to be CYP 450 isozymes. Valsartan has much greater affinity (about 20,000 fold) for the AT] receptor than for the AT₂ receptor.. Because valsartan does not inhibit ACE, it does not affect the response to bradykinin. Valsartan does not bind to or block other hormone receptors or ion channels known to be important in cardiovascular regulation. The recommended starting dose of valsartan is 80 mg or 160 mg once daily when used as monotherapy in patients who are not volume-depleted. The antihypertensive effect is substantially present within 2 weeks and maximal reduction is generally attained after 4 weeks. No initial dosage adjustment is required for elderly patients, for patients with mild or moderate renal impairment, or for patients with mild or moderate liver insufficiency. Safety and effectiveness in pediatric patients have not been established. The overall incidence of adverse experiences reported with valsartan was similar to placebo.

Telmisartan, 4' - { [2-propyl-4-methyl-6-( 1 -methyl-benzimidazol-2-yl)- benzimidazol- 1-yl] -methyl }-biphenyl-2-carboxylic acid, is a nonpeptide angiotensin II receptor (type AT]) antagonist, described in US 5591762. Following oral administration, peak concentrations of telmisartan are reached in 0.5-1 hours after dosing. Following either intravenous or oral administration, most of the administered dose (>97%) was eliminated unchanged in faeces via biliary excretion. Telmisartan is metabolized by conjugation to form a pharmacologically inactive acylglucuronide; the glucuronide of the parent compound is the only metabolite that has been identified in human plasma and urine. Telmisartan has much greater affinity (>3000 fold) for the AT] receptor than for the AT₂ receptor. Because telmisartan does not inhibit ACE, it does not affect the response to bradykinin. Telmisartan does not bind or block other hormone receptors or ion channels known to be important in cardiovascular regulation. The usual starting dose of the tablets is 40 mg once a day. Blood pressure response is dose related over the range of 20-80 mg. Most of the antihypertensive effect is apparent within two weeks and maximal reduction is generally attained after four weeks. No initial dosing adjustment is necessary for elderly patients or patients with mild-to-moderate renal impairment. Safety and effectiveness in pediatric patients have not been established. The overall incidence of adverse events similar to that of placebo was observed.

Eprosartan, (E)-3-[2-butyl-l-(4-carboxybenzyl)-imidazol-5-yl]-2-(2- thienylmethyl)-2-propenoic acid, is described in US 5185351. Eprosartan is a reversible inhibitor of the AT_j receptor. The affinity of eprosartan for the AT_j receptor is about 1000 times more than for the AT₂ receptor. Eprosartan is eliminated by biliary and renal excretion, primarily as an unchanged compound. There are no known active metabolites. Plasma concentrations peak at about 1-2 hours after oral administration.

The usual recommended starting dose of eprosartan is 600 mg once a day when used as monotherapy, but daily doses can range from 400-800 mg. No initial dosing adjustment is needed for the elderly or those with renal impairment if the maximum dose does not exceed 600 mg per day. Eprosartan is well-tolerated by most people up to 1200 mg per day.

Lercanidipine - a Calcium Antagonist

Another class of active agents that is used for the freatment of hypertension is calcium antagonists. These active agents influence the influx of calcium ions into cells, especially smooth muscle cells. Inhibition of calcium influx produces a relaxation of smooth muscles, such as those around the arteries and veins, which leads to a decrease in the observed hypertension. Such active agents as well as their hypotensive activity are described in a number of publications and patent applications.

Lercanidipine, methyl 1 , 1 ,N-trimethyl-N-(3 ,3 -diphenylpropyl)-2-aminoethyl l,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-pyridine-3,5-dicarboxylate, is a highly lipophilic dihydropyridine calcium antagonist with long duration of action and high vascular selectivity. Its mechanism of antihypertensive activity is due to a direct relaxant effect on vascular smooth muscle, thus lowering total peripheral resistance. The recommended starting dose of lercanidipine HC1 ("lercanidipine") immediate release tablets as monotherapy is 10 mg daily by oral route, with a drug tifration to 20 mg daily (for immediate release), and up to 80 mg per day for modified release formulations. Lercanidipine is rapidly absorbed following oral administration with peak plasma levels occurring 2-3 hours following dosing. Elimination is essentially via the hepatic route. By virtue of its high lipophilicity and high membrane distribution, lercanidipine combines a short plasma half life with a long duration of action. In fact, the preferential distribution of the drug into membranes of smooth muscle cells results in membrane-controlled pharmacokinetics which is characterized by a prolonged pharmacological effect. In comparison to other calcium antagonists, lercanidipine is characterized by gradual onset and long-lasting duration of action despite decreasing plasma levels. In vitro studies show that isolated rat aorta response to high K⁺ may be attenuated by lercanidipine, even after the drug has been removed for 6 hours. Lercanidipine is commercially available from Recordati S.p.A. (Milan, Italy) and has been described along with methods for making it and resolving it into individual enantiomers in US 4705797, US 5767136, US 4968832, US 5696139, EP 0153016 and EP 0824517.

Clinical studies have shown that lercanidipine 10 mg daily (typically titrated to 20 mg daily in patients not responding or responding inadequately to the 10 mg dose) provides a sustained pharmacological action and a significant antihypertensive effect. In hypertensive patients the onset of lercanidipine action is gradual and the drug has a consistent and sustained blood pressure lowering effect throughout the dosage interval. The gradual and smooth antihypertensive effect has been recently confirmed also by using the "Smoothness Index", as described in Omboni and Zanchetti, Hypertension, 1998, 16:1831-8. The analysis of a large population of hypertensive patients has documented that lercanidipine is a very well tolerated drug, for the most part, with few and/or moderate side effects, including tachycardia, palpitations and oedema. However, only about 0-9% of patients have reported tachycardia as a side effect. Since lercanidipine is mebabolized by CYP3A4, co-administration with other drugs that strongly inhibit CYP34A could lead to excessively high levels of lercanidipine. Thus, interactions with lercanidipine with CYP34A substrates such as Midazolam, sildenafil, Simvastatin and cyclosporine should be avoided. Moreover, any CYP34A inhibitors such as ketoconazole, itraconazole, ritonavir, and erythromycin in combination with lercanidipine is not recommended. In man, lercanidipine is confraindicated (as are all dihydropyridines) in patients with unstable angina or recent (<1 month) myocardial infarction.

Diuretics Diuretics are often added as adjunct therapy to ARBs for the treatment of hypertension. In fact, telmisartan is co-formulated with the diuretic hydrochlorothiazide (HCT) for the treatment of hypertension. Diuretics reduce the amount of fluid in the blood stream by lowering the amount of salt and water in the body, which helps to reduce blood pressure. There are three main types of diuretic drugs, thiazide diuretics, potassium-sparing diuretics, and loop diuretics. However, while it is known that co-administration of an ARB and a diuretic is effective therapy for the prevention and treatment of hypertension (see WO 03/087045, which claims a combination of valsartan, amlodipine and HCT), and WO 2003/09704 discloses triple a combination of an ARB, a calcium channel blocker and a diuretic, neither of these published applications disclose lercanidipine as the calcium channel blocker, nor do they disclose specific combinations with superior results. Thiazide diuretics directly inhibit sodium and calcium reabsorbtion and augment calcium absorption in the early distal convoluted tubule of the kidney. Excess sodium, calcium, and water reduce extracellular volume in mild to moderate congestive heart failure. Thiazide diuretics potentiate anti-hypertensive agents by about a third to a half in an effort to reduce blood pressure and overall fluid volume. The increased serum concentrations of calcium allow for the relaxation of arterial smooth muscle that, in turn, reduces peripheral vascular resistance. Thiazide diuretics possess anti-hypertensive properties because of the direct vasodilatation of arterioles, altered sodium balance, and the reduction in fluid volume. Thiazide diuretics are not the drug of choice if massive amount of diuresis are necessary or if the patient has a history of intolerance to sulfa containing medications.

Commonly used thiazide diuretics are bendroflumethiazide, chlorthalidone, chlorothiazide, hydrochlorothiazide, hydroflumethiazide, methyclothiazide, metolazone, poly thiazide, quinethazone and trichlormethiazide, Additional thiazide or thiazide-like diuretics include benzylhydrochlorothiazide, cyclopenthiazide, cyclothiazide, ethiazide and indapamide.

Potassium-sparing diuretics are mild diuretics that act upon the distal convoluting tubule to inhibit sodium exchange for potassium. Gradually, sodium and water are excreted in the urine, and potassium is conserved. Aldactone (spironolactone) is a synthetic steroid that is similar to aldosterone and acts as an antagonist by competing for aldosterone binding sites. Inhibition of aldosterone leads to the excretion of sodium and the retention of potassium in the distal portion of the nephron. Other members of the potassium-sparing diuretic group do not alter aldosterone binding, but work primarily by impairing the exchange of potassium and sodium in the distal convoluting tubule. These agents act as a slight anti-hypertensive and potentiate anti-hypertensive medications. Potassium-sparing diuretics can be given in conjunction with potassium-pitching diuretics in an attempt to prevent hypokalemia and the complications involved with that particular electrolyte imbalance.

Commonly used potassium-sparing diuretics include spironolactone, triamterene and amiloride.

Loop diuretics relieve excess extracellular fluid volume and regulate vascular osmolarity. Loop diuretics are the most potent and expedient diuretics available, and they inhibit the reabsorbtion of sodium, chloride, and potassium ions in the ascending loop of Henle in the kidney. Loop diuretics also cause renal vasodilatation and a transient rise in glomerular filtration rate. The combination of increased renal blood flow and the prevention of the sodium-potassium-chloride co-transport system permits secretion of large volumes of fluid and electrolytes. Loop diuretics have systemic hemodynamic effects: increased venous capacitance (which reduces left ventricular filling pressures or preload), increased ejection fraction (an indicator of improved ventricular function), decreased systemic and peripheral vascular resistance (reduced pulmonary, organ, and extracellular edema or afterload) which all allow for a reduction in blood pressure and cardiac workload.

Commonly used loop diuretics include bumetanide, torsemide, ethacrynic acid and furosemide.

Since they are not very effective alone, the potassium-sparing diuretics are often typically combined into a single tablet or capsule with another diuretic, usually HCT. In addition, diuretics are often put together into a single tablet or capsule with drugs from other classes of antihypertensives.

Several pharmacological rationales can be advanced for combining any ARB with a calcium antagonist to treat hypertension. For example, the fact that multiple physiologic systems participate in blood pressure control has been proposed as a major reason why individual active agents decrease in efficacy over time. Pharmacological intervention on one of these systems is believed to trigger counterregulatory mechanisms. A combination of treatments increases the number of mechanisms potentially capable of reducing an elevated blood pressure and reduces the rate and magnitude of the adverse events produced by each drug. Further, the addition of one agent may counteract some deleterious effects of the other. Therefore a low-dose combination of two different agents reduces the risk of dose-related adverse reaction while still allowing sufficient blood pressure reduction.

In addition to pharmacological advantages, combination therapy has been requested to meet evolving guidelines that look for more aggressive treatment of blood pressure. For example, recent World Health Organisation guidelines recommend a diastolic blood pressure lower than 85 mm Hg and a systolic blood pressure lower than 130 mm Hg in younger patients and in diabetic patients.

Fixed combinations offer the possibility of administering a combination of active agents in a single dosage form. Such a form will likely increase the patient compliance. That is, such a dosage form will likely increase a patient's adherence to a therapeutic scheme and will increase the success of such a treatment therapy. Additionally, a number of patents are nonresponsive to one or more of the available monotherapies, and some patients are not responsive to known combination therapies. There is no way at present to predict whether these patients will be responsive to therapy using a new combination of active ingredients. It has been calculated that, overall, 30-50% of patients are non-responders to monotherapy (this average does not include data of patients taking lercanidipine).

The combined therapy of lercanidipine with candesartan cilexetil, an ARB available from AstraZeneca under the trade name Atacand^®, has been reported (Aranda, et al., J. Hypertension 18 (Suppl. 2), June 2000, SI 52).

There is a continuing need for safe and effective combination of anti- hypertensivefreatments that have a long lasting, selective mechanism of action with few side effects.

Definitions

International Non-Proprietary Names are used herein to mean not just the base of the compound concerned, but pharmaceutically acceptable salts of the base and any other derivative form of the base which has received or which does in the future receive a marketing authorization or product license. For example, "lercanidipine" is used to mean not just the base methyl l,l,N-trimethyl-N-(3,3-diphenylpropyl)-2-aminoethyl l,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-pyridine-3,5-dicarboxylate but also any of its pharmaceutically acceptable salts, e.g., its salts with inorganic or organic acids such as HC1, HBr, H₂SO₄, maleic acid, fumaric acid, tartaric acid and citric acid.

"Hypertension" as used herein refers to abnormally high arterial blood pressure, when compared to prior blood pressure readings, maintained over a specified time period. Conventionally, the time period is 3-6 months. The increase may be observed in systolic pressure, diastolic pressure, or both. Typically, blood pressure is expressed as two numbers separated by a slash, where the first number is the systolic pressure (the pressure induced by the contraction of the heart by which the blood is forced onward and the circulation kept up) and the second number is the diastolic pressure (the pressure induced by the dilatation of the cavities of the heart during the period in which they fill with blood). Conventionally, hypertension is defined as a blood pressure of equal to or greater than 140/90 mm Hg.

Blood pressure in normal and hypertensive adults is typically categorized as follows:

Source: The Sixth Report of the Joint National Committee on Prevention, Detection, Evaluation, and treatment of High Blood Pressure

The term "antihypertensive activity" refers to the effect of an active agent to lower the blood pressure of a patient with hypertension by at least 20 mm Hg for systolic pressure and/or by at least 10 mm Hg for diastolic pressure. The active agent may or may not decrease the blood pressure in a person that does not have hypertension or may not decrease blood pressure in all persons with hypertension or may not achieve blood pressure regulation in all patients who experience a decrease in blood pressure. In a preferred embodiment, the active agent decreases a patient's blood pressure to below 140/90 mm Hg (i.e., regulates the blood pressure). The term "active agent" or "active ingredient" refers to a compound that produces a pharmacological effect that leads to a physiological change. As used herein, the active agents are antihypertensive agents, such as lercanidipine and the ARBs which are employed in the combination treatment of the invention. Conventionally, an active agent is considered as having an antihypertensive effect if it decreases either systolic or diastolic blood pressure by at least 10 mm Hg.

The term "predetermined increment" refers to the minimum reduction in blood pressure that is needed for a patient to decrease blood pressure to or below a predetermined limit, preferably 140/90. Thus, an active agent which at a dosage tolerated by the patient achieves reduction by a predetermined increment is considered effective to treat hypertension in the specific patient, and the patient is considered responsive to this agent (also known as a "responder"). In other words, if an active agent decreases blood pressure by a predetermined increment in one patient (i.e., has sufficient antihypertensive activity in the patient) but does not decrease blood pressure by the predetermined increment in another patient (i.e., does not have sufficient antihypertensive activity in the patient), then the first patient is responsive to the treatment (a "responder", as defined below) but the second patient is not (a "non- responder", as defined below). The decrease in blood pressure can be in the systolic pressure, diastolic pressure, or both. Reduction in diastolic blood pressure is a particularly desirable result.

As used herein, the term "responder" refers to a patient that has previously responded to a treatment for hypertension involving administration of a particular active agent (or combination of active agents) in a particular amount or amounts. In other words, the active agent or active agents have "antihypertensive activity" and reduce the patient's blood pressure by the "predetermined increment". A determination of responsiveness to an antihypertensive regimen may require administration of a particular agent in a particular amount and frequency for a period of time, usually 1 month for ARBs and calcium antagonists. Such treatments include, but are not limited to, administration of ARBs, calcium channel blockers, beta blockers, ACE inhibitors and diuretics. The phrase "responsive to monotherapy" refers to patients who are administered only one active agent (monotherapy) and the monotherapy achieves a reduction in blood pressure by the "predetermined increment" as that term is defined above. The term "non-responder" refers to a patient who has been determined not to have responded to freatment for hypertension with a particular agent or combination of agents, i.e., for whom the regimen has not achieved a reduction in blood pressure. In other words, the active agent or active agents do not have "antihypertensive activity" in the patient, and therefore the patient's blood pressure is not decreased by the "predetermined increment". The term encompasses patients who do not undergo any decrease in blood pressure upon treatment e.g. with lercanidipine alone or an ARB alone.

The term "partial responder" refers to a patient for whom a particular active agent (or combination of active agents), in a particular amount or amounts, produces "antihypertensive activity" in the patient but does not decrease blood pressure by the "predetermined increment". Increases in the amount of active agent (or combination of active agents) may or may not further decrease the blood pressure of these patients. The term encompasses patients that respond only insufficiently, i.e., exhibit some decrease in blood pressure, but short of the "predetermined increment" (to below 140/90 mm Hg). Generally, in those patients the amount of antihypertensive agent needs to be increased. But this may bring on or aggravate side effects.

The terms "suboptimal" or "sub-threshold" amounts of active agent for monotherapy refer to amounts of active agent that are insufficient to decrease blood pressure by the predetermined increment. A "suboptimal" amount is an amount that is effective to decrease the blood pressure but does not decrease it sufficiently to achieve regulation in a patient who would be a responder if administered an effective amount. A "sub-threshold" amount is an amount that provides no effect on individuals who would respond to an effective dose. "Suboptimal" or "sub-threshold" amounts may well vary from patient to patient. A patient who fails to achieve a decrease in blood pressure by the predetermined increment upon administration of a given dosage of active agent has either been administered a "suboptimal" or "sub-threshold" amount of active agent or may, alternatively, be a non-responder to the active agent. "Suboptimal" or "sub- threshold" amounts of active agent may be distinguished from the case of administration to a non-responder by increasing the administered dosage of active agent. In the case where a patient fails to achieve a decrease in blood pressure by the predetermined increment due to administration of a "suboptimal" or "sub-threshold" amount of active agent, administration of an increased dosage of active agent will cause the patient to achieve a decrease in blood pressure by the predetermined increment. In the case where a patient fails to achieve a decrease in blood pressure by the predetermined increment due to said patient being a non-responder, increasing the dosage of active agent will not cause the patient to achieve a decrease in blood pressure by the predetermined increment.

As used herein, the term "monotherapy" refers to the administration of a single active agent to treat hypertension.

The term "combination therapy" refers to the concomitant administration of two (or more) active agents for the treatment of a single disease state. The active agents may be combined and administered in a single dosage form, may be administered as separate dosage forms at the same time, or may be administered as separate dosage forms at different times, for example alternately or sequentially on the same or separate days.

The Invention

The invention provides the use of lercanidipine in the preparation of a medicament for the treatment of hypertension in combination with the prior, concurrent or post-administration of an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan, and optionally of a diuretic.

The invention also provides a composition comprising lercanidipine and an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan. Such a composition is preferably a pharmaceutical composition comprising lercanidipine, an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan and losartan, and a pharmaceutically acceptable diluent or carrier. The composition or pharmaceutical composition may optionally contain a diuretic.

A preferred diuretic is hydrochlorothiazide (HCT), 6-chloro-3,4-dihydro-2H- l,2,4,-benzothiadiazine-7-sulphonamide 1,1-dioxide, although any of the diuretics discussed above may be used.

The medicaments and pharmaceutical compositions of the invention may be useful in several classes of patients. A first class of patients is those that are responders to monotherapy with lercanidipine or with an ARB, but who suffer from side-effects and for whom it would be desirable to decrease the dosage amount of the active agent used in monotherapy.

A second class of patients is those who are non-responders to monotherapy. The medicaments of the invention may be particularly desirable for those patients that are resistant to lercanidipine monotherapy. Lercanidipine generally works quite well, so patients resistant to lercanidipine monotherapy can be difficult to treat.

A third class of patients is those who are partial responders to monotherapy and combination therapy. Monotherapy or combination therapy produces an antihypertensive effect in these patients, but the therapy does not decrease the blood pressure by the predetermined increment. Higher doses do not produce the desired effect of decreasing blood pressure by the predetermined amount, and may produce undesirable side effects.

A fourth class of patients is those that are responders to monotherapy but have been previously determined (or are expected) to become non-responders over time. Conventionally, patients in this class, upon becoming non-responders, would then require a monotherapy involving higher dosage amounts of the same active agent or would need a change of medication to another active agent to treat hypertension (i.e., reduce blood pressure by the predetermined increment). However, it should be noted that these patients may not further respond to increased dosages due to maximal efficacy of the compound having been reached. The cause for such a change in a patient's response also may be a compensation (counterregulatory) mechanism or another cause.

In principle, the medicaments of the invention could be employed with naive patients, although the regulatory authorities' guidelines do not encourage such a practice.

The medicaments and pharmaceutical compositions described herein have the potential advantages of allowing freatment with suboptimal amounts of one or both agents, sub-threshold amounts of at least one active agent, allowing greater tolerability in patients sensitive to the active agent, of allowing for synergism, i.e., superadditivity between active agents, of allowing for sustained long term efficacy of treatment and for sustained dosaging throughout a dosage period or for achieving regulation of blood pressure that was elevated, i.e., severe hypertension. Moreover, the compositions and methods described herein have potential of being of increased effectiveness in freatment or decreased side effects, compared to other combinations of lercanidipine and other classes of active agents or combinations of lercanidipine with other ARBs. In the invention lercanidipine, the ARB and the optional diuretic may be further combined with one or more additional active ingredients, e.g., a β-receptor blocker and/or an ACE inhibitor. The dosage amounts of the active agents may be adjusted when combined with other active agents to achieve desired effects (e.g., reduction of blood pressure by a predetermined increment, reduction or avoidance of a particular side-effect).

Suitable β-receptor blockers include acebutolol HC1, atenolol, betaxolol HC1, carvedilol, esmolol HC1, labetalol HC1, levobunolol HC1, metoprolol, nadolol, pindolol, propranolol HC1, sotalol HC1, timolol and timolol maleate.

Suitable ACE inhibitors include captopril, enalapril, lisinopril, quinapril, fosinopril, ramipril, cilazapril, spirapril, benazepril and perindopril.

The medicament of the invention may be administered by any route that results in systematic availability of the active ingredients. Intravenous, intratracheal, subcutaneous, oral, parenteral, buccal, sublingual, ophthalmic, intranasal, pulmonary, transmucosal, transdermal, and intramuscular routes are possible. Dosage forms may be prepared as appropriate for the chosen administration route, and may include pharmaceutically acceptable additives such as carriers, diluents, flavourants, sweeteners, preservatives, dyes, binders, suspending agents, dispersing agents, colourants, disintegrants, excipients, lubricants, plasticisers, edible oils or any combination of two or more of the foregoing. Oral administration is preferred, especially for chronic treatment. Solid dosage forms, such as tablets, pills and capsules, are preferred for oral administration but liquid dosage forms can also be used. Solid unit dosage forms may be prepared by mixing the active agents with a pharmaceutically acceptable carrier and any other desired additives as described above. The mixture is typically mixed until a homogeneous mixture of the active agents of the invention and the carrier and any other desired additives is formed, i.e., until the active agents are dispersed evenly throughout the composition. In this case, the compositions can be formed as dry or moist granules. Tablets or pills can be coated or otherwise compounded to form a unit dosage form which has delayed and/or prolonged action, such as time release and sustained release unit dosage forms. For example, the tablet or pill can comprise an inner dosage and an outer dosage component, the latter being in the form of a layer or envelope over the former. The two components can be separated by an enteric layer which serves to resist disintegration in the stomach and permits the inner component to pass intact into the duodenum or to be delayed in release.

Biodegradable polymers for controlling the release of the active agents, include, but are not limited to, polylactic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydro-pyrans, polycyanoacrylates and cross-linked or amphipathic block copolymers of hydrogels.

For liquid dosage forms, the active substances or their physiologically acceptable salts are brought into solution, suspension or emulsion, optionally with the usually employed substances such as solubilizers and emulsifiers. Solvents for the active combinations and the corresponding physiologically acceptable salts, can include water, physiological salt solutions or alcohols, e.g. ethanol, propane-diol or glycerol. Additionally, sugar solutions such as glucose or mannitol solutions may be used. A mixture of the various solvents mentioned may be used.

Transdermal administration may also be used for timed release and sustained release of the active agents of the invention, dosage form also is contemplated by the invention. Transdermal dosage forms may be a diffusion-driven transdermal system (transdermal patch) using either a fluid reservoir or a drug-in-adhesive matrix system. Other transdermal dosage forms include, but are not limited to, topical gels, lotions, ointments, transmucosal systems and devices, and iontohoretic (electrical diffusion) delivery system.

Parenteral administration may be suitable in some cases. Dosage forms for parenteral administration, and in particular by injection, typically include a pharmaceutically acceptable carrier as described above. They may be in the form of solutions, suspensions or emulsions. A preferred liquid carrier is vegetable oil. Injection may be, for example, intravenous, epidural, intrathecal, intramuscular, intraluminal, intratracheal, or subcutaneous.

The active agents also can be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines.

The active agents of the invention may also be coupled with soluble polymers as targetable drug carriers. Such polymers include, but are not limited to, polyvinyl- pyrrolidone, pyran copolymer, polyhydroxypropylmethacryl-amidephenol, polyhydroxy-ethylaspartamidephenol, and polyethyl-eneoxideopolylysine substituted with palmitoyl residues.

The medicament of the invention may be administered according to a dosage and administration regimen defined by routine testing in order to obtain optimal antihypertensive activity (especially for patients who are partial responders or non- responders to conventional monotherapy or to other combination therapies) and a decreased in blood pressure by the predetermined increment while minimizing toxicity or side-effects for a particular patient. The exact dosage and administration regimen utilizing the combination therapies of the invention is selected in accordance with a variety of factors including type, species, age, weight, gender and medical condition of the patient; the severity and etiology of the hypertension to be treated; the route of adminisfration; the renal and hepatic function of the patient; the treatment history of the patient; and the responsiveness of the patient. Optimal precision in achieving concentrations of active agents within the range that yields efficacy without toxicity requires a regimen based on the kinetics of the drug's availability to target sites. This involves a consideration of the absorption, distribution, metabolism, excretion of a drug, and responsiveness of the patient to the dosage regimen.

For combination therapy according to the invention, the active agents may initially be provided as separate dosage forms until an optimum dosage combination and administration regimen is achieved. Therefore, the patient may be titrated to the appropriate dosages for his/her particular hypertensive condition. After the appropriate dosage of each of the active agents is determined to achieve a decrease of the blood pressure by the predetermined increment without untoward side effects, the patient may then be switched to a single dosage form containing the appropriate dosages of each of the active agents, or may continue with a dual dosage form.

As indicated above, dosages will be determined for each patient individually. As guidance, however, the daily dosages for oral administration to an adult human are likely to fall within the following ranges: Lercanidipine: 2.5 to 60 mg, preferably 5 to 40 mg,

Olmesartan: 5 to 40 mg, preferably 10 to 20 mg,

Irbesartan: 50 to 300 mg, preferably 150 to 300 mg,

Valsartan: 80 to 320 mg, preferably 160 to 320 mg,

Telmisartan: 20 to 80 mg, preferably 40 to 80 mg, and

Losartan: 25 to 100 mg, preferably 50 to 100 mg.

The following is an exemplary composition of a tablet for oral adminisfration:

Ingredient Amount (mg)

Lercanidipine HC1 10

Olmesartan 20

Lactose 102

Microcrystalline cellulose 40

Sodium bicarbonate 8

Sodium starch glycolate 20

Povidone K30 8

Magnesium stearate 2

The above composition may be used as the tablet or may be used as a core for a coated tablet. The following composition is exemplary of a coating for a coated tablet:

Ingredient Amount (mg)

Hypromellose 1.91

Talc 0.15

Titanium dioxide 0.60

Macrogol 6000 0.30

Ferric oxide 0.04

The following compositions are suitable for 5 and 10 mg dosage tablets of lercanidipine, in cases in which the lercanidipine is formulated separately from the selected ARB: Ingredient Amount (mg) Amount (mg)

Core

Lercanidipine HC1 5.0 10.0 Lactose monohydrate 35.0 30.0

Microcrystalline cellulose 39.0 39.0 Sodium starch glycolate 15.5 15.5 Povidone 4.5 4.5

Magnesium stearate 1.0 1.0 Core total weight 100.0 100.0

Coating

Opadry OY-SR-6497(*) 3.00 3.00

*Hypromellose 1.91 1.91

*Talc 0.15 0.15

* Titanium dioxide 0.60 0.60

*Macrogol 6000 0.30 0.30

*Ferric oxide 0.04 0.04

Total 103.00 103.00

Examples

The following Examples illustrate the invention.

Example 1 - Lercanidipine and Irbesartan

Male Sprague-Dawley rats, weighing 250-300 g, are anaesthetized with pentobarbital sodium (35 mg/kg, i.p.) and placed on a thermic blanket. The temperature is maintained at 37°C with a thermoregulator via a rectal probe. The animals are tracheotomized to facilitate spontaneous breathing. A polyethylene catheter is placed in the left jugular vein to allow for infusion of pentobarbital sodium to maintain anesthesia. The left femoral vein and artery are cannulated with polyethylene catheters to allow drugs adminisfration and to monitor blood pressure, respectively.

Animals then undergo a left nephrectomy by excising the left kidney via a flank incision. The right kidney and renal vein, artery and ureter are then exposed via a right refroperitoneal incision, under a dissecting microscope. Silk threads are placed around both vessels and ureter. The cavity is then covered with Vaseline oil. See Recordati, et al. 2000, J. Hypertension, 18:1277-1287. After 30-60 minutes of basal recordings of arterial blood pressure and heart rate the threads around the renal vessels and ureter are tied close to the renal hilum to induce complete renal ischemia of the right kidney. After 2 hours of ischemia, the threads are removed to allow renal reperfusion and urine output. The reopening of the renal hilum and restoration of renal circulation, induces an increase in blood pressure that peaks after 5-10 minutes and lasts about 60 minutes. Drugs [vehicle, irbesartan (100 μg/kg), lercanidipine (7.5 μg/kg), or both lercanidipine (7.5 μg/kg) and irbesartan (100μg/kg)] are administered intravenously at 5 minutes after reperfusion is begun. Irbesartan is tested individually and in combination with lercanidipine. Statistical analysis is performed during the reperfusion time (time 125-180 minutes). To evaluate the effects of drugs administration on blood pressure and heart rate within each group, two-way ANOVA and Dunnet's test was used ("within treatment").

To evaluate the statistical differences among the treatment groups, data are analyzed using a three-way ANOVA (analysis of variance) with repeated measures on factor time and pre-planned multiple comparisons. The analysis was performed using, for each treatment, delta values (log transformation) Statistical analysis is performed by means of general linear model procedure (GLM) with SAS software version 6.12. ("between treatments"). In the vehicle-treated group, the hypertensive state was maintained without any statistically significant decrease in systolic and diastolic blood pressure up to 25 and 40 min, respectively, after the kidney reperfusion (Table 1; Figures 1 and 2). Lercanidipine (7.5 μg/kg) and irbesartan (100 μg/kg) induced a statistically significant decrease in both systolic and diastolic blood pressure (Tables 2 and 3; Figures 1 and 2). The antihypertensive effect was similar for both drugs. When lercanidipine and irbesartan were administered in association (7.5 μg/kg + 100 μg/kg, respectively) they induced a rapid and significant fall in blood pressure, reversing immediately the ischemia-induced hypertension and lowering blood pressure to normotensive pre-ischemic levels (Table 4; Figures 1 and 2). The combination of lercanidipine and irbesartan induced changes in SBP statistically different from those observed after injection of vehicle and irbesartan alone (Table 5). Furthermore, the decrease of SBP induced by the administration of the combination was long-lasting, as the decrease of SBP was significantly different from the vehicle treated group for the whole period of observation (Figure 3).

The decrease of DBP induced by administration of the two drugs tested alone was significantly different (on the whole) with respect to the vehicle, and the effect observed after adminisfration of the combination was significantly different from all the other treatments (Table 6).

. The antihypertensive effect induced by the combination of drugs on diastolic blood pressure was higher than each single drug and statistically different from control group from 5 min to the end of the experiment (Figure 4). The antihypertensive effect of the combination appeared additive, since the decrease in DBP induced by the drugs in combination was equal or higher than the sum of the decrease induced by each single drag.

With regard to the effect of treatments on HR, irbesartan induced no modification on heart rate respect to the vehicle treatment, whereas lercanidipine induced a positive chronofropic effect, probably due to the baroreflex-activation, that was different from that observed in the vehicle and irbesartan group. Interestingly, when lercanidipine was administered together with irbesartan the whole changes in heart rate were not different from those of the vehicle-treated group (Table 7).

As shown in Figure 6, the combination of lercanidipine and irbesartan reduces the tachycardic peak effect due acute administration of lercanidipine alone, at least after the first 20 min after admimstration. The increases of HR observed 5 min after adminisfration of lercanidipine alone and of the combination, in fact, were significantly different (p <0.05).

The results from this study demonstrate that the combination of lercanidipine and the ARB irbesartan immediately reduced, in an additive manner, the extent of renal hypertension for the entire duration of the experiment. The effects of combination resulted longer lasting in comparison with the effects of the compounds administered alone, in particular with regard to the decrease of SBP (Figure 3).

Furthermore, the combination of lercanidipine and irbesartan reduced the tachycardic effect of lercanidipine administered alone. TABLE 1

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous adminisfration of vehicle (1 ml/kg) in uninephrectomized anesthetized rats. Mean value + S.E.M. (n=6).

b= p <0.01 vs 125 min (within freatment) (Two way ANOVA and Dunnett's test)

TABLE 2

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous admimstration of lercanidipine (7.5 μg/kg) in uninephrectomized anesthetized rats. Mean value + S.E.M. (n=6).

Time SBP DBP HR min mmHg mmHg beats/min

0 120.7 + 3.5 75.7 + 1.7 383.3 ± 12.0

Ischemia

120 105.2 + 5.3 63.3 ± 2.9 381.7 ± 7.5

Reperfusion

125 160.7 + 8.5 117.2 + 4.4 346.7 ± 8.4

Drug

130 136.7 ± 7.4 b 92.0 + 5.4 b 434.2 ± 15.3 b

135 140.7 ± 7.1 b 94.3 ± 6.0 b 426.7 ± 14.3 b

150 143.2 + 7.2 b 96.3 + 5.7 b 411.7 ± 12.8 b

165 144.5 ± 9.1 b 94.3 ± 6.3 b 411.7 ± 15.6 b

180 136.8 + 8.4 b 91.8 ± 6.7 b 410.0 ± 18.3 b

b = p <0.01 vs 125 min (within treatment). (Two way ANOVA and Dunnett's test) TABLE 3

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of irbesartan (100 μg/kg) in uninephrectomized anesthetized rats. Mean value ± S.E.M. (n=6).

Time SBP DBP HR min mmHg mmHg beats/min

0 113.3 ± 6.7 63.8 + 3.5 373.3 ± 14.5

Ischemia

120 112.0 ± 5.6 59.0 ± 2.0 360.0 ± 13.4

Reperfusion

125 153.7 ± 5.1 107.0 ± 4.8 336.7 ± 14.1

Drug

130 134.3 ± 5.5 b 86.3 + 5.8 b 341.7 ± 13.8

135 138.7 + 5.3 b 90.0 + 5.6 b 346.7 ± 13.6

150 139.0 ± 5.7 b 91.7 + 6.1 b 361.7 ± 7.5 a

165 136.0 ± 7.6 b 85.7 + 6.6 b 363.3 ± 11.8 a

180 125.7 ± 8.3 b 79.3 + 8.1 b 375.0 ± 13.1 b a=p<0.05; b= p<0.01 vs 125 min (within treatment). (Two way ANOVA and Dunnett's test)

TABLE 4

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous adminisfration of lercanidipine (7.5 μg/kg) and irbesartan (100 μg/kg) in uninephrectomized anesthetized rats.

Mean value ± S.E.M. (n=6).

Time SBP DBP HR min mmHg mmHg beats/min

0 109.8 ± 4.0 70.0 ± 2.0 388.3 ± 4.0

Ischemia

120 96.3 ± 4.3 57.3 ± 2.1 368.3 ± 8.3

Reperfusion

125 148.7 ± 7.1 107.8 ± 5.0 346.7 ± 9.5

Drug

130 107.3 ± 5.9 b 62.3 + 4.4 b 390.0 ± 10.3 b

135 110.3 ± 6.3 b 66.7 ± 4.5 b 400.0 ± 8.9 b

150 113.3 + 7.8 b 70.7 ± 5.6 b 406.7 ± 6.7 b

165 113.2 ± 7.5 b 70.3 ± 5.6 b 400.0 ± 7.3 b

180 109.0 ± 8.6 b 68.7 ± 6.0 b 403.3 ± 8.0 b

b= p<0.01 vs 125 min (within treatment). (Two way ANOVA and Dunnett's test) TTTAAABBBLLLEEE 555

Systolic blood pressure: general differences among groups

TABLE 6

Diastolic blood pressure: general differences among groups

TABLE 7

Heart rate: general differences among groups

Example 2 - Lercanidipine and Valsartan

Male Sprague-Dawley rats, weighing 250-300 g, are anaesthetized with pentobarbital sodium (35 mg/kg, i.p.) and placed on a thermic blanket. The temperature is maintained at 37 EC with a thermoregulator via a rectal probe. The animals are tracheotomized to facilitate spontaneous breathing. A polyethylene catheter is placed in the left jugular vein to allow for infusion of pentobarbital sodium to maintain anesthesia. The left femoral vein and artery are cannulated with polyethylene catheters to allow drugs adminisfration and to monitor blood pressure, respectively. Animals then undergo a left nephrectomy by excising the left kidney via a flank incision. The right kidney and renal vein, artery and ureter are then exposed via a right retroperitoneal incision, under a dissecting microscope. Silk threads are placed around both vessels and ureter. The cavity is then covered with Vaseline oil. See Recordati, et al. 2000, J. Hypertension, 18:1277-1287.

After 30-60 minutes of basal recordings of arterial blood pressure and heart rate the threads around the renal vessels and ureter are tied close to the renal hilum to induce complete renal ischemia of the right kidney. After 2 hours of ischemia, the threads are removed to allow renal reperfusion and urine output. The reopening of the renal hilum and restoration of renal circulation, induces an increase in blood pressure that peaks 5-10 minutes and lasts about 60 minutes. Drugs [vehicle, valsartan 30 g/kg), lercanidipine (10 g/kg), or both lercanidipine (10 g/kg) and valsartan (30 μg/kg)] are administered intravenously at 5 minutes after reperfusion is begun. The

ARB, valsartan, is tested individually and in combination with lercanidipine. Results are expressed as mean values + S.E (in the tables) and in % changes

(in the figures).

To evaluate the effects of drugs administration on blood pressure within each group ("within treatment") and the statistical differences among the treatments

("between treatments"), the data were analysed using a three-way ANOVA with repeated measures on factor time and pre-planned multiple comparisons, using SAS software version 8.2.

The analysis "between treatments" was performed using delta values for each freatment (parameters value at time "t" minus basal value for each animal).

Effects of treatments on BP

Following reperfusion, a marked and fast increase of blood pressure, that had its peak at 5 min after the reopening of vascular renal hilum, was observed (Tables 8 to 11). In order to compare the effects of single drags and their combination on the reperfusion-induced hypertension, the compounds were intravenously administered at the peak effect of hypertension (5 min) and statistical analysis ("within groups") was performed from the adminisfration time to the end of experiment (time 125-180 in the Tables and Figures). In the vehicle-treated group, the hypertensive state was maintained without statistically significant decrease in both systolic and diastolic blood pressure up to 55 min after the kidney reperfusion (Table 8; Figures 7 and 8); lercanidipine (10 μg/kg) induced a statistically significant decrease in diastolic blood pressure form 5 to 60 min after administration and systolic blood pressure from 10 to 60 min (Table 9; Figures 7 and 8).

Valsartan at 30 μg/kg, induced a statistically significant decrease in diastolic blood pressure from 5 to 60 min after administration, whereas the effects on systolic blood pressure were statistically significant at 10 and 60 after administration (Table 10; Figures 7 and 8). The antihypertensive effect of valsartan was similar to that induced by lercanidipine.

When lercanidipine and valsartan were administered together (10 μg/kg + 30 μg/kg, respectively) they induced a rapid and significant fall in blood pressure, reversing immediately the ischemia-induced hypertension and lowering blood pressure to normotensive pre-ischemic levels (Table 11; Figures 7 and 8).

In order to compare the effects of the different treatments on BP, statistical analysis was performed on Δ SBP and Δ DBP values.

On the whole, the changes in SBP induced by injection of lercanidipine and valsartan alone were not statistically different from those observed in the vehicle- treated animals, whereas the combination of both drags induced changes statistically different from those observed after injection of vehicle alone from 5 to 25 min after administration (Table 12). Furthermore, the antihypertensive effect of the combination appeared additive at 5 min after the admimstration, since the decrease in SBP was higher than the sum of the decrease induced by each single drag, as shown in Figure 9.

The decrease of DBP induced by administration of the two drugs tested alone and in combination was significantly different (on the whole) with respect to the vehicle (Table 13). The antihypertensive effect induced by the combination of drags on diastolic blood pressure was higher than each single drag and statistically different from control group from 5 min to the end of the experiment (Figure 10). Effects of treatments on HR

Valsartan induced no statistically significative modification on heart rate (Table 10), whereas lercanidipine is characterized by a positive chronofropic effect, probably due to the baroreflex-activation, that was more marked than that observed in the vehicle group.

Interestingly, when lercanidipine was administered together with valsartan the whole changes in heart rate were not different from those of the vehicle- and valsartan-treated groups and statistically different from lercanidipine group (Table 14): as shown in Figures 11 and 12, the combination of lercanidipine and valsartan reduced the tachycardic effect due acute admimstration of lercanidipine alone.

TABLE 8

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous adminisfration of vehicle (1 ml/kg) in uninephrectomized anesthetized rats. Mean value + S.E.M. (n=6).

a=p <0.05; b= p<0.01 vs 125 min (within treatment) TABLE 9

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous adminisfration of lercanidipine (10 μg/kg) in uninephrectomized anesthetized rats. Mean value + S.E.M. (n=5).

b= p<0.01 vs l25 min (within treatment).

TABLE 10

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of valsartan (30 μg/kg) in uninephrectomized anesthetized rats. Mean value ± S.E.M. (n=7).

a=p<0.05; b= pO.Ol vs 125 min (within treatment). TABLE 11

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of lercanidipine

(10 μg/kg) and valsartan (30 μg/kg) in uninephrectomized anesthetized rats.

Mean value ± S.E.M. (n=5).

a=p <0.05; b= p <0.01 vs 125 min (within treatment).

TABLE 12

Systolic blood pressure: general differences among groups

TABLE 13

Diastolic blood pressure: general differences among groups

TABLE 14

Heart rate: general differences among groups

Example 3 - Lercanidipine and Olmesartan

Male Sprague-Dawley rats, weighing 250-300 g, were anaesthetized with pentobarbital sodium (35 mg/kg, i.p.) and placed on a thermic blanket. The temperature was maintained at 37°C with a thermoregulator via a rectal probe. The animals were tracheotomized to facilitate spontaneous breathing. A polyethylene catheter was placed in the left jugular vein to allow for infusion of pentobarbital sodium to maintain anesthesia. The left femoral vein and artery were cannulated with polyethylene catheters to allow drugs adminisfration and to monitor blood pressure, respectively.

Animals then underwent a left nephrectomy by excising the left kidney via a flank incision. The right kidney and renal vein, artery and ureter were then exposed via a right retroperitoneal incision, under a dissecting microscope. Silk threads were placed around both vessels and ureter. The cavity was then covered with Vaseline oil. See Recordati, et al. 2000, J. Hypertension, 18:1277-1287.

After 30-60 minutes of basal recordings of arterial blood pressure and heart rate the threads around the renal vessels and ureter were tied close to the renal hilum to induce complete renal ischemia of the right kidney. After 2 hours of ischemia, the threads were removed to allow renal reperfusion and urine output. The reopening of the renal hilum and restoration of renal circulation, induced an increase in blood pressure that peaked at 5-10 min and lasted about 60 min. Due to the intravenous mode of administration, olmesartan free acid was used in these treatments. Control (vehicle) or drug [olmesartan (10 μg/kg), lercanidipine (7.5 μg/kg), or both lercanidipine (7.5 μg/kg) and olmesartan (10 μg/kg)] were administered intravenously at 5 minutes after reperfusion is begun. Results were expressed as mean values ± S.E (in the tables) and in percent change observed.

To evaluate the effects of drug administration on blood pressure within each group ("within treatment") and the statistical differences among the treatments ("between treatments"), the data were analyzed using a three-way ANOVA with repeated measures on factor time and pre-planned multiple comparisons, using SAS software version 8.2. The analysis "between treatments" was performed using delta values for each treatment (parameters value at time "t" minus basal value for each animal).

Effects of treatments on BP

Following reperfusion, a marked and fast increase of blood pressure was observed that had its peak at 5 min after the reopening of vascular renal hilum. (Tables 15-18). In order to compare the effects of single drugs and their combination on the reperfusion-induced hypertension, the compounds were intravenously administered at the peak of induced hypertension (5 min) and statistical analysis ("within groups") was performed from the time of drug or vehicle administration to the end of experiment (125-180 min). In the control vehicle-treated group, the hypertensive state was maintained without statistically significant decrease in diastolic blood pressure up to 60 min (and in systolic blood pressure up to 45 min) after the kidney reperfusion (Table 15; Figures 13 and 14). Lercanidipine administration induced a statistically significant decrease in diastolic blood pressure from 5 to 55 min after adminisfration (Table 16; Figures 13 and 14).

Olmesartan induced a statistically significant decrease in diastolic blood pressure from 5 to 55 min after administration, whereas the effects on systolic blood pressure were statistically significant at 40 and 55 min after administration (Table 17; Figures 13 and 14). The antihypertensive effect of olmesartan was similar to that induced by lercanidipine.

The combination of lercanidipine and olmesartan administered together induced a rapid and significant fall in blood pressure, immediately reversing the ischemia- induced hypertension and lowering blood pressure to normotensive pre-ischemic levels (Table 18; Figures 13 and 14). In order to compare the effects of the different treatments on BP, statistical analysis was performed on ΔSBP and ΔDBP values.

On the whole, the changes in SBP induced by injection of lercanidipine and olmesartan alone were not statistically different from those observed in the vehicle- treated animals, whereas the combination of both drugs induced changes statistically different from those observed after injection of vehicle alone (Table 19). The antihypertensive effect of the lercanidipine-olmesartan combination appeared additive at 5, 10 and 25 min after the administration, since the decrease in SBP was higher or equal than the sum of the decrease induced by each single drug (Figure 15).

The decrease of DBP induced by administration of lercanidipine and olmesartan tested alone and in combination was significantly different (on the whole) with respect to the vehicle (Table 20). The effect of olmesartan alone was statistically different from control group from 5 to 40 min after administration. The antihypertensive effect induced by the combination of lercanidipine and olmesartan on diastolic blood pressure was higher than each drug alone and statistically different from control group from 5 min to the end of the experiment (Figure 16).

Effects of treatments on HR Treatment with olmesartan alone induced no statistically significant change in heart rate (Table 17), whereas treatment with lercanidipine alone was characterized by an increase in heart rate. The observed increase in heart rate was probably due to the baroreflex-activation, that was more marked than that observed in the vehicle group. Interestingly, when lercanidipine was administered together with olmesartan the observed change in heart rate was not significantly different from the changes heart rate observed in the vehicle- and olmesartan-freated groups (Table 21 and Figures 17 and 18). Hence, the combination of olmesartan and lercanidipine completely inhibited the tachycardic effect observed following administration of lercanidipine alone. The observed differences in heart rate observed following administration of lercanidipine alone versus administration of the combination of lercanidipine and olmesartan were significant (p<0.01) throughout the observed freatment period.

TABLE 15

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of vehicle (1 ml/kg) in uninephrectomized anesthetized rats. Mean value + S.E.M. (n=6).

α=p<0.05; b= p<0.01 v 125 min (within treatment)

TABLE 16 Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of lercanidipine (7.5 μg/kg) in uninephrectomized anesthetized rats. Mean value ± S.E.M. (n=6).

b= p<0.01 vs 125 min (within freatment) TABLE 17

Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) after intravenous administration of olmesartan (10 μg/kg) in uninephrectomized anesthetized rats. Mean value ± S.E.M. (n=6).

b= p<0.01 v 125 min (within freatment).

TABLE 18 Time course of the effects on SBP (systolic blood pressure), DBP (diastolic blood pressure) and HR (heart rate) following infravenous admimstration of lercanidipine (7.5 μg/kg) and olmesartan (10 μg/kg) in uninephrectomized anesthetized rats.

Mean value ± S.E.M. (n=6).

b= p<0.01 v 125 min (within freatment). TABLE 19

Systolic blood pressure differences among freatment groups

TABLE 20

Diastolic blood pressure differences among treatment groups

TABLE 21

Heart rate differences among treatment groups

Claims

1. Use of lercanidipine in the preparation of a medicament for the treatment of hypertension in combination with the prior, concurrent or post-administration of an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan.

2. Use of lercanidipine in the preparation of a medicament for the treatment of hypertension in combination with the prior, concurrent or post-administration of an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan and in further combination with the prior, concurrent or post-administration of a diuretic.

3. Use according to claim 2 wherein the diuretic is hydrochlorothiazide.

4. Use according to claim 3 wherein the medicament is formulated to allow the oral administration of from 20 to 50 mg of hydrochlorothiazide per day.

5. Use according to any one of claims 1 to 4 wherein the medicament is formulated to allow the oral adminisfration of lercanidipine.

6. Use according to claim 5 wherein the medicament is formulated to allow the administration of from 2.5 to 60 mg of lercanidipine or a pharmaceutically acceptable salt thereof per day.

7. Use according to claim 5 wherein the medicament is formulated to allow the administration of from 5 ^' to 40 mg of lercanidipine or a pharmaceutically acceptable salt thereof per day.

8. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is olmesartan.

9. Use according to claim 8 wherein the medicament is formulated to allow the oral administration of olmesartan.

10. Use according to claim 9 wherein the medicament is formulated to allow the admimstration of from 5 to 40 mg of olmesartan per day.

11. Use according to claim 9 wherein the medicament is formulated to allow the admimstration of from 10 to 20 mg of olmesartan per day.

12. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is irbesartan.

13. Use according to claim 12 wherein the medicament is formulated to allow the oral adminisfration of irbesartan.

14. Use according to claim 13 wherein the medicament is formulated to allow the admimstration of from 50 to 300 mg of irbesartan per day.

15. Use according to claim 13 wherein the medicament is formulated to allow the administration of from 150 to 300 mg of irbesartan per day.

16. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is valsartan.

17. Use according to claim 16 wherein the medicament is formulated to allow the oral adminisfration of valsartan.

18. Use according to claim 17 wherein the medicament is formulated to allow the administration of from 80 to 320 mg of valsartan per day.

19. Use according to claim 17 wherein the medicament is formulated to allow the adminisfration of from 160 to 320 mg of valsartan per day.

20. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is telmisartan.

21. Use according to claim 20 wherein the medicament is formulated to allow the oral administration of telmisartan.

22. Use according to claim 21 wherein the medicament is formulated to allow the admimstration of from 20 to 80 mg of telmisartan per day.

23. Use according to claim 21 wherein the medicament is formulated to allow the adminisfration of from 40 to 80 mg of telmisartan per day.

24. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is losartan.

25. Use according to claim 24 wherein the medicament is formulated to allow the oral admimstration of losartan.

26. Use according to claim 25 wherein the medicament is formulated to allow the administration of from 25 to 100 mg of losartan per day.

27. Use according to claim 25 wherein the medicament is formulated to allow the admimstration of from 50 to 100 mg of losartan per day.

28. Use according to any one of claims 1 to 7 wherein the angiotensin II receptor blocker is eprosartan.

29. Use according to claim 28 wherein the medicament is formulated to allow the oral administration of eprosartan.

30. Use according to claim 29 wherein the medicament is formulated to allow the admimstration of from 400 to 800 mg of eprosartan per day.

31. Use according to claim 29 wherein the medicament is formulated to allow the administration of from 600 to 800 mg of eprosartan per day.

32. A composition comprising lercanidipine and an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan.

33. A composition according to claim 32, further comprising a diuretic.

34. A composition according to claim 33 in which the diuretic is hydrochlorothiazide .

35. A pharmaceutical composition comprising lercanidipine, an angiotensin II receptor blocker selected from olmesartan, irbesartan, valsartan, telmisartan, losartan and eprosartan, and a pharmaceutically acceptable diluent or carrier.

36. A pharmaceutical composition according to claim 35, further comprising a diuretic.

37. A pharmaceutical composition according to claim 36 in which the diuretic is hydrochlorothiazide .

38. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercamdipme and from 5 to 40 mg of olmesartan.

39. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercanidipine and from 50 to 300 mg of irbesartan.

40. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercanidipine and from 40 to 320 mg of valsartan.

41. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercanidipine and from 20 to 80 mg of telmisartan.

42. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercanidipine and from 25 to 100 mg of losartan.

43. A pharmaceutical composition according to any one of claims 35 to 37, which composition is formulated as a single oral dosage and contains from 2.5 to 60 mg of lercanidipine and from 400 to 800 mg of eprosartan.

44. A pharmaceutical composition according to any one of claims 38 to 44, which composition contains from 20 to 50 mg of hydrochlorothiazide.