WO2018090151A1 - Pharmaceutical combination for the treatment and prevention of arterial hypertension and vascular dysfunction - Google Patents

Pharmaceutical combination for the treatment and prevention of arterial hypertension and vascular dysfunction Download PDF

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Publication number
WO2018090151A1
WO2018090151A1 PCT/CL2016/050062 CL2016050062W WO2018090151A1 WO 2018090151 A1 WO2018090151 A1 WO 2018090151A1 CL 2016050062 W CL2016050062 W CL 2016050062W WO 2018090151 A1 WO2018090151 A1 WO 2018090151A1
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abh
nac
cih
combination
rats
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PCT/CL2016/050062
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Spanish (es)
French (fr)
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Marcelo PREITE
Rodrigo DEL RÍO
Rodrigo ITURRIAGA
Paola CASANELLO
Bernardo KRAUSE
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Pontificia Universidad Catolica De Chile
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Priority to PCT/CL2016/050062 priority Critical patent/WO2018090151A1/en
Publication of WO2018090151A1 publication Critical patent/WO2018090151A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/13—Amines
    • A61K31/131—Amines acyclic
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/13—Amines
    • A61K31/145—Amines having sulfur, e.g. thiurams (>N—C(S)—S—C(S)—N< and >N—C(S)—S—S—C(S)—N<), Sulfinylamines (—N=SO), Sulfonylamines (—N=SO2)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/191—Carboxylic acids, e.g. valproic acid having two or more hydroxy groups, e.g. gluconic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
    • A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid or pantothenic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/69—Boron compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/12—Antihypertensives

Definitions

  • the invention relates to a pharmaceutical combination, useful for the treatment and prevention of arterial hypertension and vascular dysfunction comprising the active components 2 (S) -amino-6-boronohexanoic acid (ABH) and N-acetylcysteine (NAC). And to the method of treatment or prevention for arterial hypertension and / or vascular dysfunction with said combination.
  • Arterial hypertension is the main cardiovascular risk factor in the world adult population, with a high prevalence (40%). Despite the development of drugs, the effectiveness of current treatments is limited. In 50% of patients, drug treatment fails to control it.
  • Pharmacological treatment is generally applied to patients who have a moderate CVRF onwards, opting in the case of those who present a low CVRF to lifestyle interventions.
  • hypotensive drugs There are several families of hypotensive drugs, the most commonly used are angiotensin II receptor antagonists (ARAN), angiotensin converting enzyme (ACEI) inhibitors, calcium channel blockers, diuretics, ⁇ -blockers and the adrenergic blockers.
  • ARAN angiotensin II receptor antagonists
  • ACEI angiotensin converting enzyme
  • the main contribution of this invention is to propose a new antihypertensive treatment, which seeks to intervene simultaneously, on the predominant mechanisms in hypertensive pathology (vasodilator deficit such as nitric oxide (NO), increased oxidative stress and vascular remodeling).
  • vadilator deficit such as nitric oxide (NO)
  • NO nitric oxide
  • vascular remodeling mainly on the normalization of endogenous antihypertensive mechanisms, rather than on their exacerbation or blockage of prohypertensors. In this way it aims to prevent or treat episodes of hypertension and, as we will discuss below, of endothelial dysfunction.
  • Endothelial dysfunction characterized by a reduced ability to synthesize the vasodilator NO
  • Vascular NO levels are strongly determined by the availability of the L-arginine substrate and oxidative stress, which produces peroxynitrite from NO and superoxide.
  • any pharmacological intervention aimed at maintaining a physiological availability of NO in situ represents a powerful approach to reverse vascular dysfunction.
  • Increasing the doses of antioxidants has not been shown to be effective, so it is necessary to combine the antioxidant with an arginase inhibitor, to intervene in the pro-oxidant signaling mechanisms, maintaining the bioavailability of NO in the vascular system.
  • Endothelial dysfunction plays a key role in the development of hypertension.
  • One of the factors that determine vascular dysfunction and development of vascular diseases is the reduction in the availability of the main vasodilator; he does not.
  • the factors that can decrease the availability of NO in the vessels is oxidative stress.
  • NO concentration can also be affected by increasing the expression and / or activity of arginase, an enzyme that competes for the substrate L-arginine, with eNOS, the enzyme that synthesizes NO.
  • the inventors developed a pharmacological combination to protect NO, and its function by combining an inhibitor of the enzyme arginase, 2 (S) -amino-6- boronohexanoic acid (ABH) and the antioxidant N-acetylcysteine (NAC ).
  • 2 (S) -amino-6-boronohexanoic acid (ABH) can be synthesized with the method protected in application CL 201402403 or its international presentation PCT / CI 2015/050035, by Dr. Marcelo Preite.
  • US6387890 Bl discloses the use of ABH as an inhibitor of arginase activity, and as a therapeutic agent for disorders related to the activity of this enzyme.
  • US6387890 Bl indicates that it is possible to combine the composition with antioxidant agents, however, there are no specific examples of these compounds, so this document does not anticipate the invention.
  • US7537785 B2 discloses compositions for the treatment of vascular diseases related to NO insufficiency, where the use of an antioxidant in combination with NO-releasing compounds is protected, where the antioxidant is a hydralazine compound, such as hydralazine hydrochloride.
  • the antioxidant is a hydralazine compound, such as hydralazine hydrochloride.
  • NAC is not mentioned as an antioxidant alternative.
  • arginase inhibitors such as BHA, could also be used. This document does not anticipate then the combination of the invention, of NAC and BHA.
  • the invention is distinguished from what has been known so far, and a synergistic effect is found between both components in the reduction of hypertension and arterial dysfunction, especially in the normalization of vascular reactivity, for example in the response to acetyl choline. (see figures 6 and 7), added to the normalization of blood pressure, to restore the structure and functionality of blood vessels and to the reduction of oxidative stress. All of which makes this new drug combination superior to usual treatments for hypertension, an effect that is also demonstrated in the comparative examples (see example 2).
  • Figure 2 Effect of ABH administration (400 ⁇ g / day per rat, white bar) on the rise in blood pressure produced by CIH.
  • Mean ⁇ SEM. (n 6).
  • FIG. 3 Hypotensive effect of the ABH and NAC combination in hypertensive rats.
  • ABH 400 ⁇ g / day per rat, white bar
  • osmotic and NAC pumps 100 mg / day per rat
  • FIG. 4 Left panel. Cross section of an external carotid artery obtained from a rat exposed to CIH and treated with ABH (400 mg / day per rat) at 10x. Right panel, increase to lOOx. In these arteries the internal thickness, the middle layer was measured and the contractile responses were recorded.
  • ABH 400 mg / day per rat
  • Figure 5 Records with a wire myograph of maximum contractile strength in response to 125 mM KCI in carotid arteries of control rats and hypertensive rats (CIH), and treated with ABH (400 ⁇ g / rat per day), NAC (100 mg / rat per day), or the combination of both in low doses (ABH 200 ⁇ g / rat per day + NAC 100 mg / rat per day) and high doses (ABH 400 ⁇ g / rat per day + NAC 100 mg / rat per day).
  • ABH 400 ⁇ g / rat per day
  • NAC 100 mg / rat per day
  • Figure 6 Relaxation curve in response to cumulative concentrations of acetylcholine (ACh) in pre-contracted carotid arteries with 125 mM KCL, from control rats, untreated hypertensive rats (CIH), or treated with ABH (400 ⁇ g / rat per day ), NAC (100 mg / rat per day), and the combination of both in low concentrations (ABH 200 ⁇ g / rat per day + NAC 100 mg / rat per day) and high concentrations (ABH 400 ⁇ g / rat per day + NAC 100 mg / rat per day).
  • ABH 400 ⁇ g / rat per day
  • NAC 100 mg / rat per day
  • Figure 8 Systemic stress measured with the plasma TBA S method of control rats, hypertensive rats (CIH) not treated or treated with ABH (400 ⁇ g / rat per day), NAC (100 mg / rat per day), and combination of both in low (ABH 200 ⁇ g / rat per day / NAC 100 mg / rat per day) and high (ABH 400 ⁇ g / rat per day / NAC 100 mg / rat per day).
  • * p ⁇ 0.05 vs control, ANOVA, followed by multiple Tukey comparisons. (n 6-8 per group).
  • ACh acetylcholine
  • FIG. 14 Systemic oxidative stress in control rats, CIH, and CIH rats treated with NAC (400 and 200 mg / rat per day), ABH (400 ⁇ g / rat per day), and with the combination of both in different doses (ABH 400 ⁇ g / rat per day + NAC 100 mg / rat per day; ABH 200 ⁇ g / rat per day + NAC 200 mg / rat per day; ABH 200 ⁇ g / rat per day + NAC 100 mg / rat per day) and Losartan ( THE 3 mg / rat per day).
  • * p ⁇ 0.05 vs control, ANOVA, followed by multiple Tukey comparisons. (n 6-8 per group).
  • the invention relates to a new pharmaceutical combination, which is useful in the treatment and prevention of arterial hypertension and vascular dysfunction.
  • This combination comprises as active components 2 (S) -amino-6-boronohexanoic acid (ABH) and N-acetylcysteine (NAC).
  • ABS -amino-6-boronohexanoic acid
  • NAC N-acetylcysteine
  • the invention also relates to the method of treatment for arterial hypertension and / or vascular dysfunction with said combination of active compounds.
  • the invention proposes that a new pharmacological therapy for the treatment and prevention of arterial hypertension and vascular dysfunction that includes as a strategy the inhibition of the enzyme arginase, combined with an antioxidant that decreases oxidative stress and prevents the deleterious effects of reactive species of oxygen, contribute to maintaining the bioavailability of NO.
  • the inventors have found that the ABH + NAC combination not only regularizes the increase in blood pressure in a model of rats subjected to intermittent hypoxia, but also regularizes endothelial function, reverses the remodeling of the arterial wall and reduces oxidative stress, being more effective than a usual medication for the treatment of hypertension, Losar ⁇ an, producing a comprehensive correction of the problem.
  • the invention points to a pharmaceutical combination of the active components 2 (S) -amino-6-boronohexanoic acid (ABH) or its pharmaceutically acceptable salts and N-acetylcysteine (NAC).
  • active components are provided together or separately in appropriate pharmaceutical forms, which obviously also comprise pharmaceutically acceptable carriers and / or adjuvants, so as to provide the combination in the form of a tablet, capsule, syrup, dragee, injectable solution, suppository , or any other pharmaceutical form existing in the art.
  • This pharmaceutical combination is used to prepare a medication useful for the treatment of high blood pressure and vascular dysfunction.
  • the invention also aims at a method of treatment or prevention of arterial hypertension and vascular dysfunction comprising administering to a person or animal simultaneously or separately both active compounds of the invention so that they act simultaneously in the body of the person or animal suffering from hypertension or vascular dysfunction.
  • the administration doses of each of these compounds vary between 1 ⁇ to 100 mg per kilo of weight of the animal or person in the case of ABH, and between 0.1 to 100 mg per kilo of weight of the animal or person in the NAC case.
  • the exact dose to be administered in each case depends on many factors, including among others the type of disorder to be treated, age, other pathologies present, the route of administration, etc.
  • the doses vary between 10 ⁇ g to 10 mg per kilo of weight of the animal or person in the case of ABH, and between 1 to 50 mg per kilo of weight of the animal or person in the case of NAC. And even more preferably, the doses vary between 100 ⁇ g to 5 mg per kilo of weight of the animal or person in the case of ABH, and between 5 to 30 mg per kilo of weight of the animal or person in the case of NAC.
  • the antihypertensive and anti-vascular remodeling effect of the ABH + NAC combination was determined in an animal model of chronic intermittent hypoxia hypertension (CIH).
  • CHI chronic intermittent hypoxia hypertension
  • awake rats blood pressure was measured using telemetric sensors.
  • This cutting-edge technology allowed to measure physiological variables in real time without disturbing the animals.
  • the effects of ABH + NAC on changes in vascular reactivity in arteries of hypertensive rats were measured by wire myography.
  • This technique is the "gold standard" to study the mechanisms that underlie to vascular function, as well as the pharmacology of the pathways that make up this function. This methodology allows to determine in detail the effects of ABH + NAC treatment on the reactivity and vascular remodeling.
  • ABH + NAC against Losartan, an ATI receptor blocker for angiotensin II, which is the standard treatment against hypertension
  • Losartan produces a decrease in high blood pressure due to CIH, but does not reduce the increase in contractile tension in arteries of rats-CIH, while ABH + NAC does.
  • ABH + NAC proved to be more effective in reducing the vasoconstrictor effect and in regulating ACh-induced vasodilation at values similar to those found in control rats.
  • Losartan does not normalize the response to exogenous NO and does not reduce the systemic oxidative stress produced by CIH, while the ABH + NAC combination does. Therefore, although Losartan is effective in reducing high blood pressure, it does not normalize endothelial dysfunction or systemic oxidative stress, and maintains exaggerated vasodilatory responses to ACh.
  • the ABH + NAC combination was studied, determining its effects on 1) Hypertension 2) Oxidative stress 3) Vascular remodeling and 4) Vascular reactivity in the hypertension model due to intermittent hypoxia in rats.
  • Hypertension was induced in rats by chronic intermittent hypoxia, where animals were progressively subjected to hypoxia of 7% of 0 2 on the first day, until reaching 5% of 0 2 on the third day, which was maintained during the study, maintaining Rats in hypoxic chambers.
  • the blood pressure of the animals rose approximately 10 mm Hg on the third or fourth day.
  • FIG 1 illustrates the hypotensive effect of NAC in hypertensive rats.
  • NAC 100 mg / day per rat, SIGMA, USA
  • This result constitutes the first evidence that an antioxidant is able to reduce the pressure rise in this model.
  • Figure 2 illustrates the hypotensive effect of ABH, supplied by 2-ml osmotic pumps (Alzet, USA) implanted in the back under aseptic surgical conditions in hypoxic rats. It is observed that the administration of ABH (400 ⁇ g / day per rat), in osmotic pumps, effectively reduced blood pressure.
  • Figure 3 shows the hypotensive effect of the ABH and NAC combination in hypertensive rats.
  • ABH was administered in osmotic pumps, (400 ⁇ g / day per rat) and NAC (100 mg / day per rat) was administered in drinking water. It is seen that the combination of the invention reduces blood pressure to values similar to those obtained by each of the components themselves.
  • vascular reactivity was studied in segments of external 2 mm carotid arteries. Similar to the morphometric analysis results, the functional internal diameter of the external carotid arteries, determined by wire myography, was significantly reduced in rats exposed to CIH (775 ⁇ 22 ⁇ ) compared to control rats (883 ⁇ 38 ⁇ ).
  • Figure 5 shows the effect of treatments on the maximum contractile response induced by KCI (125 mM).
  • KCI 125 mM
  • ABH treatment 400 ⁇ g / rat per day
  • the ABH + NAC combinations in low doses (ABH 200 ⁇ / ⁇ 3 ⁇ 8 per day + NAC 100 mg / rat per day) and high (ABH 400 ⁇ / ⁇ 8 ⁇ 8 per day) + NAC 100 mg / rat per day), reverses the increase in the maximum tension produced by CIH.
  • NAC 100 mg / rat per day
  • NAC 100 mg / rat per day
  • Figure 6 shows the relaxation curve induced by acetylcholine (ACh) in external carotid arteries of normal rats, CIH and CIH treated with ABH, NAC and ABH + NAC. Relaxation in response to ACh in external carotid arteries was lower in CIH rats, and CIH rats treated with NAC compared to control responses. On the other hand, treatment with ABH (400 ⁇ g / rat per day) exaggeratedly increased vasodilator responses to acetylcholine above the control level.
  • ABH 400 ⁇ g / rat per day
  • CIH increased systemic oxidative stress measured by plasma lipid per oxidation (TBA S MDA).
  • TSA S MDA plasma lipid per oxidation
  • ABH treatment did not reduce oxidative stress.
  • the pharmacological therapeutic benefits of each of the ABH and NAC components are maintained by themselves, which are additive in the combination of the invention, that is, when using this combination the benefits of both components are obtained, without observing a decrease in them, as could occur in some cases of negative interaction, and additionally there is a synergistic result between both components, so that the combination of the invention attacks all the factors associated with the arterial hypertension and vascular dysfunction.
  • the combination of the invention reduces blood pressure, reverses the reduction of the internal diameter and regularizes the endothelial and contractile function in the arteries submitted to CIH.
  • Figure 9 shows the effect of Losar ⁇ an (3 mg / day, administered by means of subcutaneous osmotic pumps), on the increase in mean arterial pressure induced by chronic intermittent hypoxia in rats.
  • the mean arterial pressure measured in the 8 rats increased from 98.9 ⁇ 1.8 mmHg to 105.1 ⁇ 1.7 mmHg (p ⁇ 0.05).
  • the Losarian administration reduced the mean pressure below the previous baseline levels 88.2 ⁇ 2.4 mmHg (p ⁇ 0.05, measured on days 22-25).
  • Figure 10 shows the morphometric analyzes performed on external carotid arteries.
  • CIH produces a reduction in the internal diameter, but does not significantly increase the thickness of the intimate layer.
  • Treatments with ABH, NAC ABH + NAC and Losar ⁇ an in rats with CIH increased the inert diamere of the studied arteries.
  • the iremiamienio with Losar ⁇ an is more effective for lowering blood pressure in raises subjected to CIH, in relation to the ia ⁇ amien ⁇ os with ABH, NAC, and the ABH + NAC combination.
  • Losar ⁇ an does not reduce the increase in con ⁇ r ⁇ c ⁇ il dimension evoked by KCI in CIH raias, while the ABH + NAC combination does.
  • the iamiamienium with ABH + NAC is the most effective in reducing the KCI-induced vasoconstrictor effect in arterial vessels and in regularizing the acetylcholine-induced vasodilation at values similar to those found in the con ⁇ ral rails.
  • Losartan does not reduce systemic oxidative stress (TBARS) produced by CIH, while CIH rats treated with NAC or with the ABH + NAC combination have values similar to controls. Therefore, it is concluded that, although Losartan is the best agent to reduce high blood pressure, it does not normalize endothelial dysfunction or systemic oxidative stress, and presents exaggerated vasodilatory responses to ACh and more sensitive to SNP. These results suggest that NO is reduced in blood vessels during CIH, and that Losartan did not improve its bioavailability.

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Abstract

The invention relates to a pharmaceutical combination which can be used for the treatment and prevention of arterial hypertension and vascular dysfunction, comprising the active components 2(S)-amino-6-boronohexanoic acid (ABH) and N-acetylcysteine (NAC). The invention also relates to the method for the treatment or prevention of arterial hypertension and/or vascular dysfunction using said combination.

Description

COMBINACIÓN FARMACÉUTICA PARA EL TRATAMIENTO Y PREVENCIÓN DE LA HIPERTENSIÓN  PHARMACEUTICAL COMBINATION FOR THE TREATMENT AND PREVENTION OF HYPERTENSION
ARTERIAL Y DISFUNCIÓN VASCULAR ARTERIAL AND VASCULAR DYSFUNCTION
MEMORIA DESCRIPTIVA DESCRIPTIVE MEMORY
CAMPO TÉCNICO DE LA INVENCIÓN TECHNICAL FIELD OF THE INVENTION
La invención se refiere a una combinación farmacéutica, útil para el tratamiento y prevención de la hipertensión arterial y disfunción vascular que comprende los componentes activos ácido 2 (S)- amino-6-boronohexanoico (ABH) y N-acetilcisteína (NAC). Y al método de tratamiento o prevención para la hipertensión arterial y/o la disfunción vascular con dicha combinación. The invention relates to a pharmaceutical combination, useful for the treatment and prevention of arterial hypertension and vascular dysfunction comprising the active components 2 (S) -amino-6-boronohexanoic acid (ABH) and N-acetylcysteine (NAC). And to the method of treatment or prevention for arterial hypertension and / or vascular dysfunction with said combination.
ANTECEDENTES DE LA INVENCIÓN BACKGROUND OF THE INVENTION
La hipertensión arterial es el principal factor de riesgo cardiovascular en la población adulta mundial, con una alta prevalencia (40%). A pesar del desarrollo de fármacos, la efectividad de los tratamientos actuales es limitada. En un 50% de los pacientes, el tratamiento farmacológico no logra controlarla. Arterial hypertension is the main cardiovascular risk factor in the world adult population, with a high prevalence (40%). Despite the development of drugs, the effectiveness of current treatments is limited. In 50% of patients, drug treatment fails to control it.
Existen programas de prevención primaria de hipertensión arterial que tratan de reducir los factores de riesgo modificables, como el estilo de vida. Sin embargo, la gran mayoría de los pacientes requieren el uso de fármacos para regularizar la presión arterial. Una vez diagnostica la hipertensión, esta se clasifica de acuerdo a los valores de presión y se determinan factores de riesgo (edad, sexo, tabaquismo, obesidad, entre otros). Esto permite estimar el factor de riesgo cardiovascular (F CV) e iniciar el tratamiento antihipertensivo, que tiene como fin normalizar la presión arterial a niveles < 140/90 mm Hg. There are primary prevention programs for high blood pressure that try to reduce modifiable risk factors, such as lifestyle. However, the vast majority of patients require the use of drugs to regulate blood pressure. Once the hypertension is diagnosed, it is classified according to the pressure values and risk factors are determined (age, sex, smoking, obesity, among others). This allows estimating the cardiovascular risk factor (F CV) and initiating antihypertensive treatment, which aims to normalize blood pressure to levels <140/90 mm Hg.
El tratamiento farmacológico se aplica generalmente a pacientes que presentan un FRCV de moderado en adelante, optando en el caso de aquellos que presentan un FRCV bajo a intervenciones en el estilo de vida. Existen varias familias de fármacos hipotensores, las de uso más frecuente son los antagonistas del receptor de angiotensina II (ARAN), los inhibidores de la enzima convertidora de angiotensina (IECA), los bloqueadores de canales del calcio, los diuréticos, los β -bloqueadores y los bloqueadores a- adrenérgicos. Es pacientes hipertensos menores de 55 años, las familias de fármacos de primera línea corresponden principalmente a los ARAN e IECA, y en menor grado a β - bloqueadores. En pacientes mayores de 55 años se opta además por el uso de diuréticos y bloqueadores de los canales de calcio. Pharmacological treatment is generally applied to patients who have a moderate CVRF onwards, opting in the case of those who present a low CVRF to lifestyle interventions. There are several families of hypotensive drugs, the most commonly used are angiotensin II receptor antagonists (ARAN), angiotensin converting enzyme (ACEI) inhibitors, calcium channel blockers, diuretics, β-blockers and the adrenergic blockers. It is hypertensive patients under 55 years, the families of first-line drugs correspond mainly to the ARAN and IECA, and to a lesser extent to β - blockers In patients older than 55 years, the use of diuretics and calcium channel blockers is also chosen.
Cabe destacar que la monoterapia es efectiva en sólo un 20-30% de los casos, y esto ocurre principalmente en individuos con un F CV bajo o moderado. Por lo tanto, la mayoría de los pacientes terminan siendo tratados con una combinación de al menos 2 fármacos, destacando el uso preferente de la combinación de un IECA (o ARAN) con algún otro fármaco. Actualmente un gran porcentaje del mercado de fármacos antihipertensivos a nivel mundial es dominado por los ARAN o IECA y por los β - bloqueadores, los que presentan un número considerable de efectos adversos. Por esto la búsqueda de alternativas terapéuticas para el tratamiento de la hipertensión es aún un desafío. It should be noted that monotherapy is effective in only 20-30% of cases, and this occurs mainly in individuals with a low or moderate F CV. Therefore, most patients end up being treated with a combination of at least 2 drugs, highlighting the preferred use of the combination of an ACEI (or ARAN) with some other drug. Currently, a large percentage of the global antihypertensive drug market is dominated by the ARAN or IECA and by β-blockers, which have a considerable number of adverse effects. This is why the search for therapeutic alternatives for the treatment of hypertension is still a challenge.
El principal aporte de esta invención es proponer un nuevo tratamiento antihipertensivo, el cual busca intervenir de manera simultánea, sobre los mecanismos preponderantes en la patología hipertensiva (déficit de vasodilatadores como el óxido nítrico (NO), aumento del estrés oxidativo y remodelamiento vascular). Basándose principalmente en la normalización de los mecanismos antihipertensivos endógenos, más que en la exacerbación de éstos o el bloqueo de los prohipertensores. De este modo se apunta a prevenir o tratar episodios de hipertensión y, como comentaremos a continuación, de disfunción endotelial.  The main contribution of this invention is to propose a new antihypertensive treatment, which seeks to intervene simultaneously, on the predominant mechanisms in hypertensive pathology (vasodilator deficit such as nitric oxide (NO), increased oxidative stress and vascular remodeling). Based mainly on the normalization of endogenous antihypertensive mechanisms, rather than on their exacerbation or blockage of prohypertensors. In this way it aims to prevent or treat episodes of hypertension and, as we will discuss below, of endothelial dysfunction.
La disfunción endotelial, caracterizada por una menor capacidad de síntesis del vasodilatador NO, representa un mecanismo común y temprano en el desarrollo de la hipertensión arterial. Los niveles de NO vasculares están fuertemente determinados por la disponibilidad del sustrato L-arginina y del estrés oxidativo, que produce peroxinitrito a partir del NO y superóxido. Dado la alta inestabilidad del NO y los efectos de resistencia que genera el tratamiento con fármacos donantes de NO, cualquier intervención farmacológica que apunte a mantener una disponibilidad fisiológica de la NO in situ, representa una potente aproximación para revertir la disfunción vascular. Aumentar las dosis de los antioxidantes no ha mostrado ser efectivo, por lo que es necesario combinar el antioxidante con un inhibidor de la arginasa, para intervenir en los mecanismos de señalización pro-oxidante, manteniendo la biodisponibilidad del NO en el sistema vascular. Endothelial dysfunction, characterized by a reduced ability to synthesize the vasodilator NO, represents a common and early mechanism in the development of arterial hypertension. Vascular NO levels are strongly determined by the availability of the L-arginine substrate and oxidative stress, which produces peroxynitrite from NO and superoxide. Given the high instability of NO and the resistance effects generated by treatment with NO donor drugs, any pharmacological intervention aimed at maintaining a physiological availability of NO in situ represents a powerful approach to reverse vascular dysfunction. Increasing the doses of antioxidants has not been shown to be effective, so it is necessary to combine the antioxidant with an arginase inhibitor, to intervene in the pro-oxidant signaling mechanisms, maintaining the bioavailability of NO in the vascular system.
La disfunción endotelial juega un papel primordial en el desarrollo de la hipertensión. Uno de los factores que determinan la disfunción vascular y desarrollo de enfermedades vasculares es la reducción de la disponibilidad del principal vasodilatador; el NO. Dentro de los factores que pueden disminuir la disponibilidad del NO en los vasos se encuentra el estrés oxidativo. Por otra parte, también puede afectarse la concentración de NO al aumentar la expresión y/o actividad de la arginasa, una enzima que compite por el sustrato L-arginina, con la eNOS, la enzima que sintetiza el NO. Endothelial dysfunction plays a key role in the development of hypertension. One of the factors that determine vascular dysfunction and development of vascular diseases is the reduction in the availability of the main vasodilator; he does not. Among the factors that can decrease the availability of NO in the vessels is oxidative stress. On the other hand, NO concentration can also be affected by increasing the expression and / or activity of arginase, an enzyme that competes for the substrate L-arginine, with eNOS, the enzyme that synthesizes NO.
Considerando estos dos factores, los inventores desarrollaron una combinación farmacológica para proteger el NO, y su función combinando un inhibidor de la enzima arginasa, el ácido 2(S)-amino-6- boronohexanoico (ABH) y el antioxidante N-acetilcisteína (NAC). Considering these two factors, the inventors developed a pharmacological combination to protect NO, and its function by combining an inhibitor of the enzyme arginase, 2 (S) -amino-6- boronohexanoic acid (ABH) and the antioxidant N-acetylcysteine (NAC ).
Preferentemente, el ácido 2(S)-amino-6-boronohexanoico (ABH) puede ser sintetizado con el método protegido en la solicitud CL 201402403 o su presentación internacional PCT/CI 2015/050035, del Dr. Marcelo Preite.  Preferably, 2 (S) -amino-6-boronohexanoic acid (ABH) can be synthesized with the method protected in application CL 201402403 or its international presentation PCT / CI 2015/050035, by Dr. Marcelo Preite.
En el estado de la técnica se ha sugerido estrategias similares, pero no se ha anticipado la combinación específica de la invención, cuyas propiedades muestran efectos sorprendentes respecto de otras terapias conocidas. Por ejemplo, la patente US6387890 Bl, divulga el uso de ABH como inhibidor de la actividad de arginasa, y como agente terapéutico para desórdenes relacionados con la actividad de esta enzima. El US6387890 Bl indica que es posible combinar la composición con agentes antioxidantes, no obstante, no hay ejemplos específicos de estos compuestos, por lo que este documento no anticipa la invención. Similar strategies have been suggested in the state of the art, but the specific combination of the invention has not been anticipated, whose properties show surprising effects with respect to other known therapies. For example, US6387890 Bl discloses the use of ABH as an inhibitor of arginase activity, and as a therapeutic agent for disorders related to the activity of this enzyme. US6387890 Bl indicates that it is possible to combine the composition with antioxidant agents, however, there are no specific examples of these compounds, so this document does not anticipate the invention.
Adicionalmente, la patente US7537785 B2 divulga composiciones para el tratamiento de enfermedades vasculares relacionadas con insuficiencia de NO, donde se protege el uso de un antioxidante en combinación con compuestos que liberan NO, donde el antioxidante es un compuesto de hidralazina, tal como clorhidrato de hidralazina, donde no se menciona NAC como alternativa de antioxidante. Adicionalmente, se indica como alternativa que se podrían utilizar además inhibidores de arginasa, tales como BHA. Este documento no anticipa entonces la combinación de la invención, de NAC y BHA. Additionally, US7537785 B2 discloses compositions for the treatment of vascular diseases related to NO insufficiency, where the use of an antioxidant in combination with NO-releasing compounds is protected, where the antioxidant is a hydralazine compound, such as hydralazine hydrochloride. , where NAC is not mentioned as an antioxidant alternative. Additionally, it is indicated as an alternative that arginase inhibitors, such as BHA, could also be used. This document does not anticipate then the combination of the invention, of NAC and BHA.
De este modo, la invención se distingue de lo conocido hasta ahora, encontrándose además un efecto sinérgico entre ambos componentes en la reducción de la hipertensión y la disfunción arterial, especialmente en la normalización de la reactividad vascular, por ejemplo en la respuesta a acetil colina (ver figuras 6 y 7) , sumado a la normalización de la presión arterial, a reestablecer la estructura y funcionalidad de los vasos sanguíneos y a la reducción del estrés oxidativo. Todo lo cual hace que esta nueva combinación farmacológica sea superior a tratamientos habituales para la hipertensión, efecto que también se demuestra en los ejemplos comparativos (ver ejemplo 2). BREVE DESCRIPCIÓN DE LAS FIGURAS In this way, the invention is distinguished from what has been known so far, and a synergistic effect is found between both components in the reduction of hypertension and arterial dysfunction, especially in the normalization of vascular reactivity, for example in the response to acetyl choline. (see figures 6 and 7), added to the normalization of blood pressure, to restore the structure and functionality of blood vessels and to the reduction of oxidative stress. All of which makes this new drug combination superior to usual treatments for hypertension, an effect that is also demonstrated in the comparative examples (see example 2). BRIEF DESCRIPTION OF THE FIGURES
Figura 1: Efecto hipotensor de la administración de NAC (barra blanca) sobre el alza de presión evocada por hipoxia intermitente crónica (CIH, barra llena). La hipoxia intermitente consiste en someter a las ratas a ciclos de 5-6% 02, 12 veces por hora al día, por 8 horas. Se administró NAC (100 mg/día por rata) en el agua de bebida a partir de los 14 días de exposición a CIH. La administración de NAC redujo la hipertensión en las ratas despiertas, (n = 6). * p < 0.05, ANOVA una vía. Media ± SEM. MABP, presión arterial media. Los registros de presión arterial se realizaron con un sistema telemétrico en ratas despiertas. Figure 1: Hypotensive effect of the administration of NAC (white bar) on the pressure rise evoked by chronic intermittent hypoxia (CIH, full bar). Intermittent hypoxia consists in subjecting the rats to cycles of 5-6% 02, 12 times per hour per day, for 8 hours. NAC (100 mg / day per rat) was administered in the drinking water after 14 days of exposure to CIH. The administration of NAC reduced hypertension in awake rats, (n = 6). * p <0.05, ANOVA one way. Mean ± SEM. MABP, mean blood pressure. Blood pressure records were performed with a telemetric system in awake rats.
Figura 2. Efecto de la administración de ABH (400 μg/día por rata, barra blanca) sobre el alza de presión arterial producido por CIH. La administración de ABH, mediante bombas osmóticas, efectivamente redujo la presión arterial. Media ± SEM. (n= 6). Figure 2. Effect of ABH administration (400 μg / day per rat, white bar) on the rise in blood pressure produced by CIH. The administration of ABH, through osmotic pumps, effectively reduced blood pressure. Mean ± SEM. (n = 6).
Figura 3. Efecto hipotensor de la combinación ABH y NAC en ratas hipertensas. ABH (400 μg/día por rata, barra blanca) administrada en bombas osmóticas y de NAC (100 mg/día por rata) administrada en el agua de bebida. Media ± SEM. (n= 6). Figure 3. Hypotensive effect of the ABH and NAC combination in hypertensive rats. ABH (400 μg / day per rat, white bar) administered in osmotic and NAC pumps (100 mg / day per rat) administered in drinking water. Mean ± SEM. (n = 6).
Figura 4. Panel izquierdo. Sección transversal de una arteria carótida externa obtenida de una rata expuestas a CIH y tratada con ABH (400 mg/día por rata) a lOx. Panel derecho, aumento a lOOx. En estas arterias se midió el grosor interno, la capa media y se registraron las respuestas contráctiles.  Figure 4. Left panel. Cross section of an external carotid artery obtained from a rat exposed to CIH and treated with ABH (400 mg / day per rat) at 10x. Right panel, increase to lOOx. In these arteries the internal thickness, the middle layer was measured and the contractile responses were recorded.
Figura 5. Registros con un miógrafo de alambre de máxima fuerza contráctil en respuesta a KCI 125 mM en arterias carótidas de ratas controles y ratas hipertensas (CIH), y tratadas con ABH (400 μg/rata por día), NAC (100 mg/rata por día), o la combinación de ambos en dosis bajas (ABH 200 μg/rata por día + NAC 100 mg/rata por día) y altas (ABH 400 μg/rata por día + NAC 100 mg/rata por día). *p < 0.05 vs control; * *p < 0.01 vs control, ANOVA, seguida por comparaciones múltiples Tukey. (n = 6-8 por grupo). Figure 5. Records with a wire myograph of maximum contractile strength in response to 125 mM KCI in carotid arteries of control rats and hypertensive rats (CIH), and treated with ABH (400 μg / rat per day), NAC (100 mg / rat per day), or the combination of both in low doses (ABH 200 μg / rat per day + NAC 100 mg / rat per day) and high doses (ABH 400 μg / rat per day + NAC 100 mg / rat per day). * p <0.05 vs control; * * p <0.01 vs control, ANOVA, followed by multiple Tukey comparisons. (n = 6-8 per group).
Figura 6. Curva de relajación en respuesta a concentraciones acumulativas de acetilcolina (ACh) en arterias carótidas pre-contraídas con KCL 125 mM, de ratas controles, ratas hipertensas (CIH) no tratadas, o tratadas con ABH (400 μg/rata por día), NAC (100 mg/rata por día), y la combinación de ambos en bajas concentraciones (ABH 200 μg/rata por día + NAC 100 mg/rata por día) y altas concentraciones (ABH 400 μg/rata por día + NAC 100 mg/rata por día). *p < 0.05 vs control; **p < 0.01 vs control, ANOVA, seguida por comparaciones múltiples Tukey. (n = 6-8 por grupo). Figura 7. Máxima respuesta de relajación (panel superior) y de sensibilidad (pD2, panel inferior) en respuesta a acetilcolina (ACh) en arterias carótidas de ratas controles, de ratas hipertensas CI H no tratadas, o tratadas con ABH (400 μg/rata por día), NAC (100 mg/rata por día), y la combinación de ambos en bajas (ABH 200 μg/rata por día + NAC 100 mg/rata por día) y altas (ABH 400 μg/rata por día + NAC 100 mg/rata por día). *p < 0.05 vs control, ANOVA, seguida por comparaciones múltiples Tukey. (n = 6-8 por grupo). Figure 6. Relaxation curve in response to cumulative concentrations of acetylcholine (ACh) in pre-contracted carotid arteries with 125 mM KCL, from control rats, untreated hypertensive rats (CIH), or treated with ABH (400 μg / rat per day ), NAC (100 mg / rat per day), and the combination of both in low concentrations (ABH 200 μg / rat per day + NAC 100 mg / rat per day) and high concentrations (ABH 400 μg / rat per day + NAC 100 mg / rat per day). * p <0.05 vs control; ** p <0.01 vs control, ANOVA, followed by multiple Tukey comparisons. (n = 6-8 per group). Figure 7. Maximum relaxation response (upper panel) and sensitivity (pD2, lower panel) in response to acetylcholine (ACh) in carotid arteries of control rats, of untreated CI H hypertensive rats, or treated with ABH (400 μg / rat per day), NAC (100 mg / rat per day), and the combination of both in low (ABH 200 μg / rat per day + NAC 100 mg / rat per day) and high (ABH 400 μg / rat per day + NAC 100 mg / rat per day). * p <0.05 vs control, ANOVA, followed by multiple Tukey comparisons. (n = 6-8 per group).
Figura 8. Estrés sistémico medido con el método de TBA S en plasma de ratas controles, ratas hipertensas (CIH) no tratadas o tratadas con ABH (400 μg/rata por día), NAC (100 mg/rata por día), y la combinación de ambos en bajas (ABH 200 μg/rata por día/NAC 100 mg/rata por día) y altas (ABH 400 μg/rata por día/NAC 100 mg/rata por día). *p < 0.05 vs control, ANOVA, seguida por comparaciones múltiples Tukey. (n = 6-8 por grupo). Figure 8. Systemic stress measured with the plasma TBA S method of control rats, hypertensive rats (CIH) not treated or treated with ABH (400 μg / rat per day), NAC (100 mg / rat per day), and combination of both in low (ABH 200 μg / rat per day / NAC 100 mg / rat per day) and high (ABH 400 μg / rat per day / NAC 100 mg / rat per day). * p <0.05 vs control, ANOVA, followed by multiple Tukey comparisons. (n = 6-8 per group).
Figura 9. La administración de Losarían redujo significativamente la hipertensión en ratas despiertas sometidas a CIH. * p < 0.05 vs control, ANOVA una vía, seguida por Tukey (Media ± SEM n = 8 ratas). Figure 9. Losarian administration significantly reduced hypertension in awake rats subjected to CIH. * p <0.05 vs control, ANOVA one way, followed by Tukey (Mean ± SEM n = 8 rats).
Figura 10. Diámetro interno medido en arterias carótidas externas de ratas controles, ratas sometidas a CIH, CIH +ABH (400
Figure imgf000006_0001
CIH ABH (400 μg/día) +NAC (100 mg/día), CIH+ NAC (100 mg/día) y CIH Losatran (3 mg/día). *p < 0.05 vs. CI H, ANOVA seguida por Tukey (n = 6-8 ratas por grupo). Figure 10. Internal diameter measured in external carotid arteries of control rats, rats subjected to CIH, CIH + ABH (400
Figure imgf000006_0001
CIH ABH (400 μg / day) + NAC (100 mg / day), CIH + NAC (100 mg / day) and CIH Losatran (3 mg / day). * p <0.05 vs. CI H, ANOVA followed by Tukey (n = 6-8 rats per group).
Figura 11. Máxima fuerza contráctil en respuesta a KCI 125 mM en arterias carótidas externas ratas controles y de ratas hipertensas CIH-no tratadas, CIH tratadas con la combinación ABH (400ug/rata por día) + NAC (100 mg/rata por día) y CIH tratadas con Losarían (LOS 3 mg/rata por día). *p < 0.05 vs control, ANOVA seguida por Tukey (n = 6-8 ratas por grupo). Figure 11. Maximum contractile force in response to 125 mM KCI in external carotid arteries control rats and CIH-untreated, CIH treated rats treated with the ABH combination (400ug / rat per day) + NAC (100 mg / rat per day) and CIH treated with Losarían (LOS 3 mg / rat per day). * p <0.05 vs control, ANOVA followed by Tukey (n = 6-8 rats per group).
Figura 12. Curva de relajación en respuesta a concentraciones acumulativas de acetilcolina (ACh) en arterias carótidas pre-contraídas con KCI, de ratas controles y ratas hipertensas (CIH) no tratadas, y ratas CIH tratadas con la combinación ABH (400 μg/día) +NAC (100 mg/día) y ratas-CIH tratadas con Losarían (3 mg/día). ***p < 0.001 vs control; **p < 0.01 vs control, *p < 0.05 vs control. ANOVA seguido por Tukey. (n = 6-8 ratas por grupo). Figure 12. Relaxation curve in response to cumulative concentrations of acetylcholine (ACh) in carotid arteries pre-contracted with KCI, from control rats and untreated hypertensive rats (CIH), and CIH rats treated with the ABH combination (400 μg / day ) + NAC (100 mg / day) and CIH rats treated with Losarían (3 mg / day). *** p <0.001 vs control; ** p <0.01 vs control, * p <0.05 vs control. ANOVA followed by Tukey. (n = 6-8 rats per group).
Figura 13. Máxima respuesta de relajación (panel superior) y sensibilidad (PD2, panel inferior) en respuesta a acetilcolina (ACh) en arterias carótidas pre-contraídas con KCI 125 mM, de ratas controles y de ratas hipertensas CIH-no tratadas, CIH tratadas con la combinación ABH (400ug/rata por día) + NAC (100 mg/rata por día) y CIH tratadas con Losartan (3 mg/rata por día). *p < 0.05 vs control, ANOVA seguida por Tukey (n = 6-8 ratas por grupo). Figure 13. Maximum relaxation response (upper panel) and sensitivity (PD2, lower panel) in response to acetylcholine (ACh) in pre-contracted carotid arteries with 125 mM KCI, control rats and CIH-untreated hypertensive rats, CIH treated with the ABH combination (400ug / rat per day) + NAC (100 mg / rat per day) and CIH treated with Losartan (3 mg / rat per day). * p <0.05 vs control, ANOVA followed by Tukey (n = 6-8 rats per group).
Figura 14. Estrés oxidativo sistémico en ratas controles, CIH, y ratas CIH tratadas con NAC (400 y 200 mg/rata por día), ABH (400 μg/rata por día), y con la combinación de ambos en diferentes dosis (ABH 400 μg/rata por día + NAC 100 mg/rata por día; ABH 200 μg/rata por día + NAC 200 mg/rata por día; ABH 200 μg/rata por día + NAC 100 mg/rata por día) y Losartan (LOS 3 mg/rata por día). *p < 0.05 vs control, ANOVA, seguida por comparaciones múltiples Tukey. (n = 6-8 por grupo). Figure 14. Systemic oxidative stress in control rats, CIH, and CIH rats treated with NAC (400 and 200 mg / rat per day), ABH (400 μg / rat per day), and with the combination of both in different doses (ABH 400 μg / rat per day + NAC 100 mg / rat per day; ABH 200 μg / rat per day + NAC 200 mg / rat per day; ABH 200 μg / rat per day + NAC 100 mg / rat per day) and Losartan ( THE 3 mg / rat per day). * p <0.05 vs control, ANOVA, followed by multiple Tukey comparisons. (n = 6-8 per group).
DESCRIPCIÓN DETALLADA DE LA INVENCIÓN DETAILED DESCRIPTION OF THE INVENTION
La invención se refiere a una nueva combinación farmacéutica, que es útil en el tratamiento y prevención de la hipertensión arterial y la disfunción vascular. Esta combinación comprende como componentes activos el ácido 2 (S)-amino-6-boronohexanoico (ABH) y la N-acetilcisteína (NAC). La invención también se refiere al método de tratamiento para la hipertensión arterial y/o la disfunción vascular con dicha combinación de compuestos activos. The invention relates to a new pharmaceutical combination, which is useful in the treatment and prevention of arterial hypertension and vascular dysfunction. This combination comprises as active components 2 (S) -amino-6-boronohexanoic acid (ABH) and N-acetylcysteine (NAC). The invention also relates to the method of treatment for arterial hypertension and / or vascular dysfunction with said combination of active compounds.
La invención propone que una nueva terapia farmacológica para el tratamiento y prevención de la hipertensión arterial y disfunción vascular que incluye como estrategia la inhibición de la enzima arginasa, combinado con un antioxidante que disminuye el estrés oxidativo y que previene los efectos deletéreos de las especies reactivas de oxígeno, contribuyen a mantener la biodisponibilidad del NO. The invention proposes that a new pharmacological therapy for the treatment and prevention of arterial hypertension and vascular dysfunction that includes as a strategy the inhibition of the enzyme arginase, combined with an antioxidant that decreases oxidative stress and prevents the deleterious effects of reactive species of oxygen, contribute to maintaining the bioavailability of NO.
Como se verá en los ejemplos, al atacar el problema hipertensivo conjuntamente desde los frentes propuestos, síntesis y estabilidad del NO, y estrés oxidativo, se obtiene un efecto importante sobre el control de la presión arterial y la disfunción endotelial en un modelo experimental de hipertensión, pues la combinación farmacológica de la invención permite intervenir en los mecanismos claves implicados en la disfunción vascular. As will be seen in the examples, when attacking the hypertensive problem together from the proposed fronts, synthesis and stability of NO, and oxidative stress, an important effect on the control of blood pressure and endothelial dysfunction is obtained in an experimental model of hypertension , since the pharmacological combination of the invention allows to intervene in the key mechanisms involved in vascular dysfunction.
Los inventores han encontrado que la combinación ABH+NAC no sólo regulariza el aumento de presión arterial en un modelo de ratas sometidas a hipoxia intermitente, sino que regulariza la función endotelia, revierte el remodelamiento de la pared arterial y reduce el estrés oxidativo, siendo más efectiva que un medicamento habitual para el tratamiento de hipertensión arterial, Losarían, produciendo una corrección integral del problema. The inventors have found that the ABH + NAC combination not only regularizes the increase in blood pressure in a model of rats subjected to intermittent hypoxia, but also regularizes endothelial function, reverses the remodeling of the arterial wall and reduces oxidative stress, being more effective than a usual medication for the treatment of hypertension, Losarían, producing a comprehensive correction of the problem.
Como indicamos, la invención apunta a una combinación farmacéutica de los componentes activos ácido 2 (S)-amino-6-boronohexanoico (ABH) o sus sales farmacéuticamente aceptables y N- acetilcisteína (NAC). Donde los componentes activos se proveen juntos o por separado en formas farmacéuticas apropiadas, las que obviamente también comprenden portadores y/o adyuvantes farmacéuticamente aceptables, de modo de proveer la combinación en forma de una tableta, cápsula, jarabe, gragea, solución inyectable, supositorio, o cualquier otra forma farmacéutica existente en la técnica. Esta combinación farmacéutica sirve para preparar un medicamento útil para el tratamiento de la hipertensión arterial y disfunción vascular. As indicated, the invention points to a pharmaceutical combination of the active components 2 (S) -amino-6-boronohexanoic acid (ABH) or its pharmaceutically acceptable salts and N-acetylcysteine (NAC). Where the active components are provided together or separately in appropriate pharmaceutical forms, which obviously also comprise pharmaceutically acceptable carriers and / or adjuvants, so as to provide the combination in the form of a tablet, capsule, syrup, dragee, injectable solution, suppository , or any other pharmaceutical form existing in the art. This pharmaceutical combination is used to prepare a medication useful for the treatment of high blood pressure and vascular dysfunction.
La invención también apunta a un método de tratamiento o prevención de la hipertensión arterial y disfunción vascular que comprende administrar a una persona o animal simultáneamente o por separado ambos compuestos activos de la invención de modo de que actúen simultáneamente en el cuerpo de la persona o animal que sufre hipertensión o disfunción vascular. Las dosis de administración de cada uno de estos compuestos varían entre 1 μ§ a 100 mg por kilo de peso del animal o persona en el caso de ABH, y entre 0,1 a 100 mg por kilo de peso del animal o persona en el caso de NAC. La dosis exacta a administrar en cada caso depende de muchos factores, incluyendo entre otros del tipo de desorden a tratar, la edad, otras patologías presentes, la vía de administración, etc. Preferentemente, las dosis varían entre 10 μg a 10 mg por kilo de peso del animal o persona en el caso de ABH, y entre 1 a 50 mg por kilo de peso del animal o persona en el caso de NAC. Y aún más preferentemente, las dosis varían entre 100 μg a 5 mg por kilo de peso del animal o persona en el caso de ABH, y entre 5 a 30 mg por kilo de peso del animal o persona en el caso de NAC. The invention also aims at a method of treatment or prevention of arterial hypertension and vascular dysfunction comprising administering to a person or animal simultaneously or separately both active compounds of the invention so that they act simultaneously in the body of the person or animal suffering from hypertension or vascular dysfunction. The administration doses of each of these compounds vary between 1 μ§ to 100 mg per kilo of weight of the animal or person in the case of ABH, and between 0.1 to 100 mg per kilo of weight of the animal or person in the NAC case. The exact dose to be administered in each case depends on many factors, including among others the type of disorder to be treated, age, other pathologies present, the route of administration, etc. Preferably, the doses vary between 10 μg to 10 mg per kilo of weight of the animal or person in the case of ABH, and between 1 to 50 mg per kilo of weight of the animal or person in the case of NAC. And even more preferably, the doses vary between 100 μg to 5 mg per kilo of weight of the animal or person in the case of ABH, and between 5 to 30 mg per kilo of weight of the animal or person in the case of NAC.
Sólo un porcentaje muy reducido de pacientes con hipertensión tienen una causa conocida para su enfermedad. En efecto, en la gran mayoría de los pacientes (85-90%) no se conoce el origen de la enfermedad. Existe limitada información respecto de los mecanismos patogénicos que llevan a la hipertensión arterial esencial en humanos, debido en parte a las consideraciones éticas que limitan los estudios invasivos y a la alta recurrencia de co-morbilidades que presentan los pacientes. En respuesta a este escenario, se han desarrollado modelos experimentales en roedores que desarrollan hipertensión debido a intervenciones como: selección de genes implicados en la fisiopatología hipertensiva, limitaciones del flujo renal, y la hipoxia intermitente que es la característica principal de la apnea del sueño. En este estudio, el efecto antihipertensivo y anti remodelamiento vascular de la combinación ABH+NAC se determinó en un modelo animal de hipertensión por hipoxia intermitente crónica (CIH). En ratas despiertas se midió la presión arterial usando sensores telemétricos. Esta tecnología de punta permitió medir variables fisiológicas en tiempo real sin perturbar a los animales. En el ámbito de la fisiología vascular, se midió los efectos de ABH+NAC sobre los cambios de reactividad vascular en arterias de ratas hipertensas mediante miografía de alambre. Esta técnica es el "gold standard" para estudiar los mecanismos que subyacen a la función vascular, así como la farmacología de las vías que componen esta función. Esta metodología permite determinar con detalle los efectos del tratamiento ABH+NAC sobre la reactividad y el remodelamiento vascular. Only a very small percentage of patients with hypertension have a known cause for their disease. In fact, in the vast majority of patients (85-90%) the origin of the disease is unknown. There is limited information regarding the pathogenic mechanisms that lead to essential arterial hypertension in humans, due in part to ethical considerations that limit invasive studies and the high recurrence of co-morbidities presented by patients. In response to this scenario, experimental models have been developed in rodents that develop hypertension due to interventions such as: selection of genes involved in hypertensive pathophysiology, renal flow limitations, and intermittent hypoxia that is the main characteristic of sleep apnea. In this study, the antihypertensive and anti-vascular remodeling effect of the ABH + NAC combination was determined in an animal model of chronic intermittent hypoxia hypertension (CIH). In awake rats, blood pressure was measured using telemetric sensors. This cutting-edge technology allowed to measure physiological variables in real time without disturbing the animals. In the field of vascular physiology, the effects of ABH + NAC on changes in vascular reactivity in arteries of hypertensive rats were measured by wire myography. This technique is the "gold standard" to study the mechanisms that underlie to vascular function, as well as the pharmacology of the pathways that make up this function. This methodology allows to determine in detail the effects of ABH + NAC treatment on the reactivity and vascular remodeling.
De este modo, en un modelo de hipertensión en ratas, inducido por hipoxia intermitente crónica, de aquí en adelante simplemente CIH, se evaluó el efecto de la combinación de la invención, ABH+NAC, sobre la presión arterial, el estrés oxidativo, la función renal y hepática, el remodelamiento vascular y las respuestas vasoactivas en arterias de resistencia. Se encontró que ABH y NAC administrados independientemente son igualmente efectivos que la administración de la combinación ABH+NAC para reducir la presión arterial, pero que sólo la combinación ABH+NAC revierte el aumento de tensión y la reducción diámetro en arterias de resistencia producido por CIH. Sorprendentemente, además la combinación ABH+NAC normalizó los valores máximos de relajación y de sensibilidad en respuesta a Acetilcolina (ACh) en el modelo estudiado. Se sabe que CIH también produce estrés oxidativo sistémico, que no fue reducido por ABH, pero si por ABH+NAC. Estos resultados demuestran que esta combinación específica de compuestos activos tiene resultados superiores a la suma de cada uno de ellos por separado. Thus, in a model of hypertension in rats, induced by chronic intermittent hypoxia, hereinafter simply CIH, the effect of the combination of the invention, ABH + NAC, on blood pressure, oxidative stress, the renal and hepatic function, vascular remodeling and vasoactive responses in resistance arteries. It was found that ABH and NAC administered independently are equally effective than the administration of the ABH + NAC combination to reduce blood pressure, but that only the ABH + NAC combination reverses the increased tension and reduced diameter in resistance arteries produced by CIH . Surprisingly, the ABH + NAC combination also normalized the maximum relaxation and sensitivity values in response to Acetylcholine (ACh) in the model studied. It is known that CIH also produces systemic oxidative stress, which was not reduced by ABH, but by ABH + NAC. These results demonstrate that this specific combination of active compounds has results greater than the sum of each of them separately.
Los resultados experimentales muestran que la combinación ABH+NAC reduce la presión arterial elevada, revierte la reducción del diámetro interno de los vasos y regulariza la función endotelial y contráctil en las arterias de ratas hipertensas. Experimental results show that the ABH + NAC combination reduces high blood pressure, reverses the reduction in the internal diameter of the vessels and regularizes endothelial and contractile function in the arteries of hypertensive rats.
También se comparó la efectividad de la combinación de la invención ABH+NAC contra Losartan, un bloqueador del receptor ATI a angiotensina II, que es el tratamiento estándar contra la hipertensión. Losartan produce una disminución de la presión arterial elevada por CIH, pero no reduce el aumento de tensión contráctil en arterias de ratas-CIH, mientras que ABH+NAC si lo hace. Además, ABH+NAC demostró ser más efectivo para reducir el efecto vasoconstrictor y para regularizar la vasodilatación inducida por ACh a valores similares a los encontrados en ratas controles. Por otra parte, Losartan no normaliza la respuesta a NO exógeno y no reduce el estrés oxidativo sistémico producido por CIH, mientras que la combinación ABH+NAC si lo hace. Por lo tanto, aunque Losartan es efectivo para reducir la presión arterial elevada, no normaliza la disfunción endotelial ni el estrés oxidativo sistémico, y mantiene respuestas vasodilatadoras exageradas a ACh. The effectiveness of the combination of the invention ABH + NAC against Losartan, an ATI receptor blocker for angiotensin II, which is the standard treatment against hypertension, was also compared. Losartan produces a decrease in high blood pressure due to CIH, but does not reduce the increase in contractile tension in arteries of rats-CIH, while ABH + NAC does. In addition, ABH + NAC proved to be more effective in reducing the vasoconstrictor effect and in regulating ACh-induced vasodilation at values similar to those found in control rats. On the other hand, Losartan does not normalize the response to exogenous NO and does not reduce the systemic oxidative stress produced by CIH, while the ABH + NAC combination does. Therefore, although Losartan is effective in reducing high blood pressure, it does not normalize endothelial dysfunction or systemic oxidative stress, and maintains exaggerated vasodilatory responses to ACh.
EJEMPLOS. Ejemplo 1. Estudio de efecto de ABH, NAC y de la combinación ABH+NAC EXAMPLES Example 1. Study of the effect of ABH, NAC and the combination ABH + NAC
Se estudió la combinación ABH+NAC, determinando sus efectos de sobre 1) Hipertensión 2) Estrés oxidativo 3) Remodelamiento vascular y 4) Reactividad vascular en el modelo de hipertensión por hipoxia intermitente en ratas. The ABH + NAC combination was studied, determining its effects on 1) Hypertension 2) Oxidative stress 3) Vascular remodeling and 4) Vascular reactivity in the hypertension model due to intermittent hypoxia in rats.
Se indujo hipertensión en ratas por hipoxia intermitente crónica, donde se somete a los animales progresivamente a hipoxia del 7% de 02 el primer día, hasta llegar a 5% de 02 el tercer día lo que se mantuvo durante el estudio, manteniendo a las ratas en cámaras hipóxicas. La presión arterial de los animales subió aproximadamente 10 mm Hg al tercer o cuarto día. Hypertension was induced in rats by chronic intermittent hypoxia, where animals were progressively subjected to hypoxia of 7% of 0 2 on the first day, until reaching 5% of 0 2 on the third day, which was maintained during the study, maintaining Rats in hypoxic chambers. The blood pressure of the animals rose approximately 10 mm Hg on the third or fourth day.
Se estudió el efecto sobre la presión en grupos de 6 ratas por serie de experimentos. A cada rata se le implantó sensores telemétricos TA11PA-C40 (Data Science International, USA) para registrar la presión arterial en la arteria femoral en animales despiertos. Se evaluaron distintas condiciones: The effect on pressure was studied in groups of 6 rats per series of experiments. Each rat was implanted with TA11PA-C40 telemetry sensors (Data Science International, USA) to record the blood pressure in the femoral artery in awake animals. Different conditions were evaluated:
• Serie Control • Control Series
• Serie CIH • CIH Series
• Serie CIH+ NAC (100 mg/día por rata, SIGMA, USA) agua bebida. • CIH + NAC Series (100 mg / day per rat, SIGMA, USA) drinking water.
• Serie CIH + ABH (400 μg/día rata) bomba osmótica • CIH + ABH series (400 μg / day rat) osmotic pump
• Serie CIH + ABH (200 μg/día rata) bomba osmótica + NAC (100 mg/día por rata) agua bebida) • CIH + ABH series (200 μg / day rat) osmotic pump + NAC (100 mg / day per rat) drinking water)
• Serie CIH +ABH (200 μg/día rata) bomba osmótica + NAC (200 mg/día por rata) agua bebida • CIH + ABH series (200 μg / day rat) osmotic pump + NAC (200 mg / day per rat) drinking water
• Serie CIH +ABH (200 μg/día rata bomba osmótica + NAC (100 mg/día por rata) agua bebida• CIH + ABH series (200 μg / day rat osmotic pump + NAC (100 mg / day per rat) drinking water
En paralelo se realizaron series de pruebas en animales sin telémetros (CONTROL, CIH, CIH+NAC, CIH + ABH bomba osmótica, CIH +ABH bomba osmótica + NAC) para obtener muestras de arterias, de sangre, para realizar los estudios de remodelamiento, de contracción vascular y estrés oxidativo. In parallel, series of tests were performed on animals without rangefinders (CONTROL, CIH, CIH + NAC, CIH + ABH osmotic pump, CIH + ABH osmotic pump + NAC) to obtain samples of arteries, blood, to carry out remodeling studies, of vascular contraction and oxidative stress.
1.1 Hipertensión. 1.1 Hypertension
La Figura 1 ilustra el efecto hipotensor de NAC en ratas hipertensas. En esta serie experimental se procedió a administrar NAC (100 mg/día por rata, SIGMA, USA) en el agua de bebida a partir de los 14 días de exposición a CIH. La administración de NAC redujo la hipertensión en las ratas despiertas (n = 6 ratas). Este resultado constituye la primera evidencia que un antioxidante es capaz de reducir el alza de presión en este modelo. La Figura 2 ilustra el efecto hipotensor de ABH, suministrado por bombas osmóticas de 2 mi (Alzet, USA) implantadas en la espalda bajo condiciones quirúrgicas asépticas en las ratas hipóxicas. Se observa que la administración de ABH (400 μg/día por rata), en las bombas osmóticas, efectivamente redujo la presión arterial. Figure 1 illustrates the hypotensive effect of NAC in hypertensive rats. In this experimental series, NAC (100 mg / day per rat, SIGMA, USA) was administered in the drinking water after 14 days of exposure to CIH. NAC administration reduced hypertension in awake rats (n = 6 rats). This result constitutes the first evidence that an antioxidant is able to reduce the pressure rise in this model. Figure 2 illustrates the hypotensive effect of ABH, supplied by 2-ml osmotic pumps (Alzet, USA) implanted in the back under aseptic surgical conditions in hypoxic rats. It is observed that the administration of ABH (400 μg / day per rat), in osmotic pumps, effectively reduced blood pressure.
Finalmente, la Figura 3 muestra el efecto hipotensor de la combinación ABH y NAC en ratas hipertensas. ABH se administró en bombas osmóticas, (400 μg/día por rata) y NAC (100 mg/día por rata) se administró en el agua de bebida. Se ve que la combinación de la invención reduce la presión arterial a valores similares a los obtenidos por cada uno de los componentes por sí mismos. Finally, Figure 3 shows the hypotensive effect of the ABH and NAC combination in hypertensive rats. ABH was administered in osmotic pumps, (400 μg / day per rat) and NAC (100 mg / day per rat) was administered in drinking water. It is seen that the combination of the invention reduces blood pressure to values similar to those obtained by each of the components themselves.
1.2 Remodelamiento vascular. 1.2 Vascular remodeling.
Medimos el diámetro interno y la capa media de arterias carotideas externas obtenidas de los animales sometidos a CIH y tratados con ABH, ABH +NAC, y NAC. La Figura 4 muestra ejemplos de cortes transversales de arterias carotideas externas. We measure the internal diameter and the middle layer of external carotid arteries obtained from animals subjected to CIH and treated with ABH, ABH + NAC, and NAC. Figure 4 shows examples of cross sections of external carotid arteries.
El análisis morfométrico realizado en arterias carótidas externas de ratas tratadas con CIH, mostró que la hipoxia intermitente causa una disminución significativa de 30% en el diámetro interno de las arterias carótidas, sin cambios en el grosor de la capa media. Estos cambios fueron acompañados por una mayor fuerza contráctil en respuesta a KCI 125 mM y un menor diámetro óptimo, determinados ex vivo por miografía de alambre, resultados que confirman la presencia de cambios en la estructura y funcionalidad de vasos sanguíneos en este modelo. Los tratamientos ABH (400 μg/día por rata), ABH (400 μg/día por rata) + NAC (100 mg/día por rata) y NAC (100 mg/día por rata) aumentaron el diámetro interno por encima del valor control. The morphometric analysis performed on external carotid arteries of rats treated with CIH showed that intermittent hypoxia causes a significant 30% decrease in the internal diameter of the carotid arteries, without changes in the thickness of the middle layer. These changes were accompanied by a greater contractile force in response to 125 mM KCI and a smaller optimal diameter, determined ex vivo by wire myography, results that confirm the presence of changes in the structure and functionality of blood vessels in this model. ABH (400 μg / day per rat), ABH (400 μg / day per rat) + NAC (100 mg / day per rat) and NAC (100 mg / day per rat) treatments increased the internal diameter above the control value .
1.3 Reactividad vascular. 1.3 Vascular reactivity.
Se estudió la reactividad vascular en segmentos de arterias carotideas externas de 2 mm. De manera similar a los resultados del análisis morfométrico, el diámetro interno funcional de las arterias carótidas externas, determinadas por la miografía de alambre, se redujo significativamente en ratas expuestas a CIH (775 ± 22 μιτι) en comparación con las ratas control (883 ± 38 μιτι).  Vascular reactivity was studied in segments of external 2 mm carotid arteries. Similar to the morphometric analysis results, the functional internal diameter of the external carotid arteries, determined by wire myography, was significantly reduced in rats exposed to CIH (775 ± 22 μιτι) compared to control rats (883 ± 38 μιτι).
La Figura 5 muestra el efecto de los tratamientos sobre la respuesta contráctil máxima inducida por KCI (125 mM). Claramente, la tensión máxima producida por KCI en arterias carótidas de ratas tratadas con CIH es significativamente mayor en comparación a la respuesta en animales control. El tratamiento con ABH (400 μg/rata por día) redujo el aumento de tensión generado por CIH, sugiriendo un efecto anti-remodelante del tratamiento con la combinación farmacológica. Las combinaciones ABH+NAC, en dosis bajas (ABH 200 μ§/Γ3ΐ8 por día + NAC 100 mg/rata por día) y altas (ABH 400 μ§/Γ8ΐ8 por día) + NAC 100 mg/rata por día), revierte el aumento de la tensión máxima producida por CIH. Sin embargo, NAC (100 mg/rata por día) por sí solo no es capaz de revertir el aumento de tensión. Figure 5 shows the effect of treatments on the maximum contractile response induced by KCI (125 mM). Clearly, the maximum tension produced by KCI in carotid arteries of rats treated with CIH is significantly higher compared to the response in control animals. ABH treatment (400 μg / rat per day) reduced the increase in tension generated by CIH, suggesting an anti-remodeling effect of the treatment with the pharmacological combination. The ABH + NAC combinations, in low doses (ABH 200 μ§ / Γ3ΐ8 per day + NAC 100 mg / rat per day) and high (ABH 400 μ§ / Γ8ΐ8 per day) + NAC 100 mg / rat per day), reverses the increase in the maximum tension produced by CIH. However, NAC (100 mg / rat per day) alone is not able to reverse the increase in tension.
La Figura 6 muestra la curva de relajación inducida por acetilcolina (ACh) en arterias carótidas externas de ratas normales, CIH y CIH tratadas con ABH, NAC y ABH+NAC. La relajación en respuesta a ACh en arterias carótidas externas fue menor en ratas CIH, y ratas CIH tratadas con NAC comparado con las respuestas controles. Por otra parte, el tratamiento con ABH (400 μg/rata por día), aumentó exageradamente las respuestas vasodilatadoras a acetilcolina por sobre el nivel control. Sólo la combinación ABH+NAC tanto a bajas concentraciones (ABH 200 μg/rata por día+ NAC 100 mg/rata por día) y como a altas concentraciones (ABH 400 μg/rata por día + NAC 100 mg/rata por día) normalizó el efecto vasodilatador de acetilcolina (Figura 7 panel superior) y potenció la sensibilidad a este agente (Figura 7 panel inferior). Por lo tanto, sólo la combinación ABH y NAC fue capaz de normalizar en términos de efecto vasodilatador la respuesta a acetilcolina. Este resultado es importante ya que demuestra que la combinación de la invención logra un resultado que no es logrado por ninguno de sus componentes por sí solos, y que la combinación de la invención lo logra incluso en bajas concentraciones de ABH (200 mg). Por lo tanto, se requiere de ambos componentes (ABH y NAC) para restablecer la respuesta normal vasodilatadora de ACh.Figure 6 shows the relaxation curve induced by acetylcholine (ACh) in external carotid arteries of normal rats, CIH and CIH treated with ABH, NAC and ABH + NAC. Relaxation in response to ACh in external carotid arteries was lower in CIH rats, and CIH rats treated with NAC compared to control responses. On the other hand, treatment with ABH (400 μg / rat per day) exaggeratedly increased vasodilator responses to acetylcholine above the control level. Only the ABH + NAC combination both at low concentrations (ABH 200 μg / rat per day + NAC 100 mg / rat per day) and at high concentrations (ABH 400 μg / rat per day + NAC 100 mg / rat per day) normalized the vasodilator effect of acetylcholine (Figure 7 upper panel) and enhanced sensitivity to this agent (Figure 7 lower panel). Therefore, only the ABH and NAC combination was able to normalize the response to acetylcholine in terms of vasodilator effect. This result is important as it demonstrates that the combination of the invention achieves a result that is not achieved by any of its components alone, and that the combination of the invention achieves it even at low concentrations of ABH (200 mg). Therefore, both components (ABH and NAC) are required to restore the normal ACh vasodilator response.
1.4 Estrés oxidativo. 1.4 Oxidative stress
CIH aumento el estrés oxidativo sistémico medido por la per oxidación de lípidos en el plasma (TBA S MDA). El tratamiento con ABH no redujo el estrés oxidativo. La combinación CIH y AC (200 y 400 μg/día por rata) y ABH + NAC (todas las dosis estudiadas) como se aprecia en la Figura 8. CIH increased systemic oxidative stress measured by plasma lipid per oxidation (TBA S MDA). ABH treatment did not reduce oxidative stress. The combination CIH and AC (200 and 400 μg / day per rat) and ABH + NAC (all doses studied) as seen in Figure 8.
De este modo se observa que en la combinación de la invención se mantienen los beneficios terapéuticos farmacológicos de cada uno de los componentes ABH y NAC por sí mismos, los que son aditivos en la combinación de la invención, es decir, que al usar esta combinación se obtienen los beneficios de ambos componentes, sin observarse una merma de ellos, como podría producirse en algunos casos de interacción negativa, y adicionalmente hay un resultado sinérgico entre ambos componentes, por lo que la combinación de la invención ataca todos los factores asociados a la hipertensión arterial y disfunción vascular. La combinación de la invención reduce la presión arterial, revierte la reducción del diámetro interno y regulariza la función endotelial y contráctil en las arterias sometidas a CIH. Thus, it is observed that in the combination of the invention the pharmacological therapeutic benefits of each of the ABH and NAC components are maintained by themselves, which are additive in the combination of the invention, that is, when using this combination the benefits of both components are obtained, without observing a decrease in them, as could occur in some cases of negative interaction, and additionally there is a synergistic result between both components, so that the combination of the invention attacks all the factors associated with the arterial hypertension and vascular dysfunction. The combination of the invention reduces blood pressure, reverses the reduction of the internal diameter and regularizes the endothelial and contractile function in the arteries submitted to CIH.
Ejemplo 2. Estudio comparativo de la combinación de la invención ABH+NAC contra Losarían Example 2. Comparative study of the combination of the invention ABH + NAC against Losarían
Se compararon los efectos de Losarían -tratamiento gold-standard para la hipertensión, con la combinación de la invención ABH+NAC sobre los distintos indicadores: 1) Hipertensión, 2) Reactividad vascular, 3) Remodelamiento vascular y 4) Estrés oxidativo; en el modelo de hipertensión por hipoxia intermitente en ratas. The effects of Losarían-gold-standard treatment for hypertension were compared with the combination of the ABH + NAC invention on the different indicators: 1) Hypertension, 2) Vascular reactivity, 3) Vascular remodeling and 4) Oxidative stress; in the model of hypertension due to intermittent hypoxia in rats.
Se realizaron dos series experimentales: Two experimental series were carried out:
Una serie experimental para medir el efecto de Losarían sobre la presión arterial elevada y sobre la contractilidad de arterias carótidas externas, en ratas sometidas a hipoxia intermitente crónica (n = 8 ratas) An experimental series to measure the effect of Losarían on high blood pressure and on the contractility of external carotid arteries, in rats subjected to chronic intermittent hypoxia (n = 8 rats)
Una serie para experimental para medir el efecto de Losarían sobre la contractilidad de arterias carótidas externas, morfometría de arterias carótidas externas y estrés oxidativo en ratas sometidas a hipoxia intermitente crónica (n = 8 ratas) An experimental series to measure the effect of Losarían on contractility of external carotid arteries, morphometry of external carotid arteries and oxidative stress in rats subjected to chronic intermittent hypoxia (n = 8 rats)
2.1 Hipertensión. 2.1 Hypertension
La Figura 9 muestra el efecto de Losarían (3 mg/día, administrado medíanle bombas osmóticas subcutáneas), sobre el aumento de presión arterial media inducido por la hipoxia intermitente crónica en ratas. La presión arterial media en medida en las 8 ratas subió de 98,9± 1,8 mmHg a 105,1±1,7 mmHg (p < 0,05). La administración de Losarían redujo la presión media bajo los niveles básales previos 88,2±2,4 mmHg (p< 0,05, medida los días 22-25). Figure 9 shows the effect of Losarían (3 mg / day, administered by means of subcutaneous osmotic pumps), on the increase in mean arterial pressure induced by chronic intermittent hypoxia in rats. The mean arterial pressure measured in the 8 rats increased from 98.9 ± 1.8 mmHg to 105.1 ± 1.7 mmHg (p <0.05). The Losarian administration reduced the mean pressure below the previous baseline levels 88.2 ± 2.4 mmHg (p <0.05, measured on days 22-25).
2.2 Remodelamiento vascular. 2.2 Vascular remodeling.
La Figura 10 muestran los análisis morfométricos realizados en arterias carótidas externas. CIH produce una reducción del diámetro interno, pero no aumenta significativamente el grosor de la capa íntima. Los tratamientos con ABH, NAC ABH+NAC y Losarían en ratas con CIH aumenían el diámeíro iníerno de las arterias esíudiadas. Figure 10 shows the morphometric analyzes performed on external carotid arteries. CIH produces a reduction in the internal diameter, but does not significantly increase the thickness of the intimate layer. Treatments with ABH, NAC ABH + NAC and Losarían in rats with CIH increased the inert diamere of the studied arteries.
2.3 Reactividad vascular.  2.3 Vascular reactivity.
Se comparó la respuesía vasoacíiva in vitro de arterias caróíidas exíernas de raías CIH íraíadas con Losarían con las respuesías arteriales de raías íraíadas con ABH (400 μg/día) + NAC (100 mg/día). Evaluamos la respuesta contráctil inducida por KCI como un índice del remodelamiento vascular, la respuesta a acetilcolina como indicador de la función endotelial y respuesta producida por nitroprusiato de sodio, un generador de NO. In vitro vasoactive response of exogenous carioid arteries of iariate CIH raias was compared with Losaria with arterial responses of iariate raias with ABH (400 μg / day) + CAP (100 mg / day). We evaluate the contractile response induced by KCI as an index of vascular remodeling, the response to acetylcholine as an indicator of endothelial function and response produced by sodium nitroprusside, a NO generator.
Al comprar los efectos de la combinación ABH (400 μg/día) +NAC (100 mg/día) sobre la función endotelial y vascular de carótidas en ratas sometidas a CIH, encontramos que Losarían no revierte el aumento en la fuerza contráctil inducido por CIH, ver Figura 11, presentando una mayor fuerza contráctil comparado con los controles y ratas-CIH tratadas con la combinación ABH+NAC. When buying the effects of the combination ABH (400 μg / day) + NAC (100 mg / day) on the endothelial and vascular function of carotids in rats subjected to CIH, we find that Losarían does not reverse the increase in contractile force induced by CIH , see Figure 11, presenting a greater contractile force compared to controls and CIH-rats treated with the ABH + NAC combination.
Por el contrario, la respuesta a acetilcolina fue mayor en ratas-CIH tratadas con Losarían en términos del efecto máximo y la sensibilidad, comparados con ratas controles y ratas-CIH tratadas con ABH+NAC, ver la Figura 12 y la Figura 13. Estos resultados sugieren que el efecto antihipertensivo de Losarían mejoran la función endotelial, pero no revierte el aumento de la fuerza contráctil inducido por la CIH, reforzando la idea que la combinación ABH+NAC es el normalizador de la función vascular. Este efecto antiremodelante exclusivo de la combinación frente a Losarían, estarían dados por los mecanismos a mediano y largo plazo acíivados por niveles de NO endógenos (derivados de la acíividad de eNOS), el cual acíúa como un regulador negaíivo de la proliferación del músculo liso vascular, así como la ausencia de esírés oxidaíivo. In contrast, the response to acetylcholine was higher in CI-rats treated with Losarían in terms of maximum effect and sensitivity, compared to control rats and CI-rats treated with ABH + NAC, see Figure 12 and Figure 13. These Results suggest that Losarían's antihypertensive effect improves endothelial function, but does not reverse the increase in contractile strength induced by CIH, reinforcing the idea that the ABH + NAC combination is the normalizer of vascular function. This exclusive antiremodelant effect of the combination against Losarían, would be given by the mechanisms in the medium and long term activated by endogenous levels of NO (derived from the eNOS activity), which acts as a negative regulator of vascular smooth muscle proliferation. , as well as the absence of oxidative esirés.
2.4 Estrés oxidativo sistémico. 2.4 Systemic oxidative stress.
Tal como esperábamos, la exposición a CIH produjo esírés oxidaíivo sisíémico medido por la peroxidación de lípidos en el plasma (MDA). El íraíamienío con Losarían o con ABH no redujo el esírés oxidaíivo, mieníras que la combinación CIH y NAC (200 y 400 μg/día por raía) y ABH + NAC (íodas las dosis esíudiadas) fueron efecíivas para disminuir el esírés oxidaíivo inducido por CIH, los resulíados se grafican en la Figura 14 As we expected, exposure to CIH produced seismic oxidative esyrés measured by plasma lipid peroxidation (MDA). The irony with Losarían or with ABH did not reduce oxidative esirés, while the combination CIH and NAC (200 and 400 μg / day per raia) and ABH + NAC (iodized the doses studied) were effective in reducing the oxidative esirés induced by CIH , the results are plotted in Figure 14
De esíos esíudios se puede concluir que, por una parte, el íraíamienío con Losarían es más efecíivo para bajar la presión arterial en raías someíidas a CIH, en relación a los íraíamieníos con ABH, NAC, y a la combinación ABH+NAC. Sin embargo, Losarían no reduce el aumenío de íensión conírácíil evocado por KCI en raías CIH, mieníras que la combinación ABH +NAC si lo hace. Adicionalmeníe el íraíamienío con ABH+NAC es el más efecíivo para reducir el efecío vasoconsíricíor inducido por KCI en vasos arteriales in viíro y para regularizar la vasodilaíación inducida por Aceíilcolina a valores similares a los enconírados en las raías coníroles. Losartan no reduce el estrés oxidativo sistémico (TBARS) producido por CIH, mientras que las ratas CIH tratadas con NAC o con la combinación ABH+NAC presentan valores similares a los controles. Por lo tanto, se concluye que, aunque Losartan es el mejor agente para reducir la presión arterial elevada, no normaliza la disfunción endotelial ni el estrés oxidativo sistémico, y presenta respuestas vasodilatadores exageradas a ACh y más sensibles a SNP. Estos resultados sugieren que el NO esta reducido en los vasos sanguíneos durante CIH, y que Losartan no mejoría su biodisponibilidad. From these studies it can be concluded that, on the one hand, the iremiamienio with Losarían is more effective for lowering blood pressure in raises subjected to CIH, in relation to the iaíamieníos with ABH, NAC, and the ABH + NAC combination. However, Losarían does not reduce the increase in conírícíil dimension evoked by KCI in CIH raias, while the ABH + NAC combination does. Additionally, the iamiamienium with ABH + NAC is the most effective in reducing the KCI-induced vasoconstrictor effect in arterial vessels and in regularizing the acetylcholine-induced vasodilation at values similar to those found in the coníral rails. Losartan does not reduce systemic oxidative stress (TBARS) produced by CIH, while CIH rats treated with NAC or with the ABH + NAC combination have values similar to controls. Therefore, it is concluded that, although Losartan is the best agent to reduce high blood pressure, it does not normalize endothelial dysfunction or systemic oxidative stress, and presents exaggerated vasodilatory responses to ACh and more sensitive to SNP. These results suggest that NO is reduced in blood vessels during CIH, and that Losartan did not improve its bioavailability.

Claims

REIVINDICACIONES
1. Combinación farmacéutica CARACTERIZADA porque comprende los componentes activos ácido 2 (S)-amino-6-boronohexanoico (ABH) o sus sales farmacéuticamente aceptables y N- acetilcisteína (NAC). 1. CHARACTERIZED pharmaceutical combination because it comprises the active components 2 (S) -amino-6-boronohexanoic acid (ABH) or its pharmaceutically acceptable salts and N-acetylcysteine (NAC).
2. La combinación farmacéutica de la reivindicación 1 CARACTERIZADA porque adicionalmente comprende portadores y adyuvantes farmacéuticamente aceptables. 2. The pharmaceutical combination of claim 1 CHARACTERIZED in that it additionally comprises pharmaceutically acceptable carriers and adjuvants.
3. La combinación farmacéutica de la reivindicación 2 CARACTERIZADA porque está en forma de una tableta, cápsula, jarabe, gragea, solución inyectable, supositorio, o cualquier otra forma farmacéutica.  3. The pharmaceutical combination of claim 2 CHARACTERIZED in that it is in the form of a tablet, capsule, syrup, dragee, injectable solution, suppository, or any other pharmaceutical form.
4. Uso de la combinación farmacéutica de la reivindicación 1 CARACTERIZADA porque sirve para preparar un medicamento útil para el tratamiento y prevención de la hipertensión arterial y disfunción vascular. 4. Use of the pharmaceutical combination of claim 1 CHARACTERIZED because it serves to prepare a medicament useful for the treatment and prevention of arterial hypertension and vascular dysfunction.
5. Método de tratamiento y prevención de la hipertensión arterial y/o la disfunción vascular CARACTERIZADO porque comprende administrar a un ser humano o animal una combinación de ácido 2 (S)-amino-6-boronohexanoico (ABH) o sus sales farmacéuticamente aceptables y N- acetilcisteína (NAC). 5. Method of treatment and prevention of arterial hypertension and / or CHARACTERIZED vascular dysfunction because it comprises administering to a human or animal a combination of 2 (S) -amino-6-boronohexanoic acid (ABH) or its pharmaceutically acceptable salts and N-acetylcysteine (NAC).
6. Método de acuerdo a la reivindicación 6 CARACTERIZADO porque la dosis de administración de ABH varía entre 1 μg a 100 mg por kilo de peso del ser humano o animal, y la dosis de administración de NAC varía entre 0,1 a 100 mg por kilo de peso del ser humano o animal. 6. Method according to claim 6 CHARACTERIZED in that the dose of ABH administration varies between 1 μg to 100 mg per kilo of human or animal weight, and the dose of NAC administration varies between 0.1 to 100 mg per kilo of weight of the human or animal.
7. Método de acuerdo a la reivindicación 7 CARACTERIZADO porque preferentemente la dosis de administración de ABH varía entre 10 μg a 10 mg por kilo de peso del ser humano o animal, y la dosis de administración de NAC varía entre 1 a 50 mg por kilo de peso del ser humano o animal. 7. Method according to claim 7 CHARACTERIZED because preferably the dose of ABH administration varies between 10 μg to 10 mg per kilo of human or animal weight, and the dose of NAC administration varies between 1 to 50 mg per kilo of human or animal weight.
PCT/CL2016/050062 2016-11-18 2016-11-18 Pharmaceutical combination for the treatment and prevention of arterial hypertension and vascular dysfunction WO2018090151A1 (en)

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