WO2013030564A1 - Vascular treatments - Google Patents

Vascular treatments Download PDF

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Publication number
WO2013030564A1
WO2013030564A1 PCT/GB2012/052109 GB2012052109W WO2013030564A1 WO 2013030564 A1 WO2013030564 A1 WO 2013030564A1 GB 2012052109 W GB2012052109 W GB 2012052109W WO 2013030564 A1 WO2013030564 A1 WO 2013030564A1
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Prior art keywords
nitrite
blood pressure
accordance
central
pressure
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PCT/GB2012/052109
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French (fr)
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Andrew Webb
Phil Chowienczyk
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King's College London
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00Medicinal preparations containing inorganic active ingredients
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00Drugs for disorders of the cardiovascular system
    • A61P9/08Vasodilators for multiple indications
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00Drugs for disorders of the cardiovascular system
    • A61P9/12Antihypertensives

Definitions

  • the present invention relates to the use of nitrites as vasodilators, and to their use in the treatment and prevention of cardiovascular conditions.
  • CVD cardiovascular disease
  • vasodilator drugs in general, has focused almost exclusively on their action on the small blood vessels in the periphery, known as the resistance vasculature,and their efficacy in reducing peripheral blood flow resistance.
  • Organic nitrates such as glyceryl trinitrate (GTN) are among the most commonly used drugs for the treatment of cardiovascular disease. They have wide therapeutic application, and are effective venodi!ators and also dilators of coronary arteries in the treatment of acute coronary syndromes. They are, however, used less frequently to control BP and, when they are, this is usually in the acute setting, as their long term effects are rapidly attenuated by the induction of tolerance.
  • GTN glyceryl trinitrate
  • ROS reactive oxygen species
  • nitrate tolerance manifests as reduced treadmill walking time and reduced time to onset of angina.
  • congestive heart failure it has been described as the loss of haemodynamic effect of the administered nitrate, and in hypertension, it is evident as the rapid loss of the hypotensive effects.
  • Elkayamei a/. [Circulation 1987; 31 (9): 1727-35]
  • the initial significant reduction in pulmonary artery wedge pressure caused by a continuous GTN infusion was no longer present at 12 hours, and by 20 hours there was a complete loss off effect, even to being numerically worse than placebo.
  • patients are often denied the beneficial haemodynamic effects of nitrates for several days, with the associated risk arising from failure to address the elevated SBP, while the skilled physician seeks to establish the optimal window in which to administer GTN.
  • Aortic dissection is a condition characterised by bleeding into and along the aorta wall. This results from a tear or damage to the inner wall of the aorta.
  • Such patients need very strict blood pressure control over several days to reduce the risk of further dissection.
  • GTN becomes ineffective after less than 24 hours, continuous good blood pressure control is frequently not possible and further dissection is common.
  • a further issue is that, whilst it is imperative to lower SBP, it is important not to excessively lower mean arterial pressure (MAP), as this is crucial for maintaining adequate perfusion to vital organs such as the kidneys.
  • MAP mean arterial pressure
  • the kidneys are at particular risk as the dissection often involves the renal arteries, and so their blood flow may already be compromised.
  • GTN is therefore potentially useful, in the short term, through its selective large, artery effect, but once tolerance has occurred, it becomes necessary to rely on alternative antihypertensive agents.
  • Most other antihypertensive medications have greater effects on resistance vessels and will cause a more marked reduction in MAP.
  • heart rate control agents such as beta-blockers (e.g. intravenous !abeta!ol)
  • beta-blockers e.g. intravenous !abeta!ol
  • a further problem with organic nitrates is that they frequently cause headaches, a very common side effect that can be very unpleasant for patients and sometimes limits the use of GTN.
  • vascular nitrite can serve as the largest source of nitric oxide (NO), provided that physiological mechanisms exist to reduce nitrite to NO.
  • Reduction of nitrite to NO is generally considered to be increased under conditions of hypoxia and ischaemia, by processes beyond simple chemical acidification/disproportionation. It is generally accepted that one of the most important mechanisms is reduction by deoxyhaemoglobin. This reaction has been well-characterised: the maximal rate of nitrite reduction and NO generation occurs at the P 50 of haemoglobin, the oxygen concentration at which half the haem is saturated with oxygen (Cosby et al., supra).
  • nitrite Whilst physiologically acceptable levels of nitrite have been shown to have a substantial venodilatory effect, and less but significant resistance arteriole dilatory affect, this has been shown to be dependent on hypoxia, and increases forearm blood flow while having no effect on pBP or MAP. As such, nitrite has not been popularly pursued in relation to general treatment of hypertension, and known uses for nitrite are dependent on, or are targeted at, conditions associated with hypoxia or ischemia. For example, one formulation currently under development is aerosolised sustained release nitrite for the treatment of pulmonary arterial hypertension that will work through inducing smail vessel dilation in the deoxygenated conditions of the pulmonary arterial circulation. Another product under development provides nitrite as an oral preparation for the selective increase of blood flow to ischaemic tissues in peripheral artery disease.
  • Ingram T er al. (Oxygen Transport to Tissue XXX, 21-25, 2009), used rabbit aorta, inferior vena cava and pulmonary artery to show the relaxing effects of 10 ⁇ sodium nitrite.
  • Such aorta preparations are frequently used to test dilators and constrictors but are not representative of their likely effect on large arteries in humans. Indeed, the test is more representative of their likely action in small arteries and resistance vessels.
  • Dilators that are poorly selective for large arteries, and which are effective in resistance vessels in humans, such as phentolamine and verapamil, are very effective in rabbit aorta preparations.
  • Such aorta preparations are also very artificial, as they lack red blood cells with the concomitant nitrite-scavenging oxyhaemoglobin, and flow resulting in shear stress and the resulting generation of endogenous NO production, but also because the rings are put under tension and then phenylephrine is applied to pre- constrict the vessel. Accordingly, even if such tests couid be considered predictive of aortic performance in humans, the indications are that a considerably higher concentration than 10 ⁇ of nitrite would be required in order to demonstrate any effect.
  • nitrite (10 ⁇ ) used in Ingram T et a!., (supra) is too high a systemic concentration to be sustained for any period of time, such as would be required in the chronic treatment of hypertension.
  • Pluta R et al., (Plos One 2011) infused different doses of sodium nitrite for 48 h in healthy volunteers and established that the maximum tolerated dose (MTD) was 266.9 pg/kg/h, equivalent to approximately 4.5 pmol/min and resulted in a systemic nitrite concentration of ⁇ 0.56 ⁇ and no change in MAP.
  • US 2006/182815 discloses treatment of cerebral artery spasm in the context of a stroke with hypoxia/ischaemia.
  • WO 2005/004884 discloses the prevention of vasospasm by nitrite due to a reaction with perivascular haemoglobin, i.e. in the cerebrospinal fluid around the vessels, where the nitrite level was found to be increased.
  • WO 2011/095389 discloses the use of theobromine for lowering centra! blood pressure.
  • inorganic nitrite at physiologically acceptable levels, is capable of reducing central systolic blood pressure and dilating the major arteries, without reducing central diastolic blood pressure, and without affecting peripheral blood pressure.
  • nitrite for use in the treatment of patients having one of elevated central systolic blood pressure and elevated central pulse pressure, or both, wherein the patient has normal peripheral blood pressure.
  • the term 'elevated' is used to mean a blood pressure reading that is determined by the skilled person, preferably a physician, to be higher than is desired, whether by comparison with the norm, taking into account such factors as the patient's age, sex, and health, or for other reasons, such as acute aortic dissection, for example, or for any other reason, including published guidelines from learned societies or published reference ranges.
  • nitrite for use in the reduction of, preferably elevated, central systolic blood pressure.
  • the level of nitrite does not exceed a systemic level of 10 ⁇ for up to 20 minutes, and for periods longer than 2 hours, preferably about 1 hour, and more preferably not for more than about 20 minutes. It is preferred that the level of nitrite not exceed 5 ⁇ systemically, with levels of between 0.3 ⁇ and 3 ⁇ , inclusive, being most preferred.
  • Chronic levels of 5 ⁇ nitrite and below are referred to herein as physiologically acceptable levels, or chronic physiologically acceptable levels. Acute physiologically acceptable levels can range as high as 100 pM, but the higher the concentration, the shorter the acceptable duration, which will be readily apparent to those skilled in the art.
  • a preferred position for a patient to adopt is the supine position.
  • the supine position is preferred for patients receiving i.v. infusion.
  • patients may receive an oral administration form prior to a period of sleep, for example, especially as nocturnal blood pressures are more strongly associated with cardiovascular events than daytime blood pressures.
  • Nitrite has been found to selectively reduce central systolic blood pressure in preference to peripheral blood pressure. It is an advantage that nitrite selectively targets large arteries over resistance arterioles, it is a further advantage that nitrite does not induce tolerance/resistance with long term use. It is a yet further advantage that nitrite treatment is not generally associated with any adverse reactions, such as headaches. It is a further advantage that the risk of suffering a cardiovascular event is not known to be increased by treatment with nitrite.
  • Nitrite as used in the present invention serves to reduce elevated central systolic blood pressure, or elevated central pulse pressure, or both, and appears to have little or no effect on peripheral systolic blood pressure at physiologically acceptable concentrations, in one embodiment, there is provided nitrite for use as described, wherein peripheral systolic blood pressure is reduced by no more than 7mmHg, most preferably by no more than 5mmHg.
  • the present invention further provides nitrite for use in the dilation of arteries, especially large arteries, preferably without causing substantial dilation of resistance arterioles.
  • Arteries having an internal diameter of 2 mm or more are preferred, although conduit-compliance small muscular arteries having an interna! diameter of 150 pm-2mm may be targeted.
  • the local average oxygen saturation of haemoglobin is preferably greater than 50%, more preferably greater than 90%, and most preferably between 95% and 100% inclusive. Nitrite is able to cause marked dilatation of the arteries under fully oxygenated conditions, including haemoglobin oxygen saturations of ⁇ 99%, without having any significant effect on pBP.
  • nitrite for use in reducing the central augmentation index, preferably by at least 3%, more preferably at least 5%.
  • augmentation index or "Alx” is used herein to refer to the ratio of the arterial augmentation pressure to the arterial pulse pressure.
  • the augmentation pressure is the difference between the pu!se pressure and the maximum pressure, and is caused by wave reflection and constructive wave overlap.
  • Alx can be expressed as a ratio or a percentage.
  • a large Alx results from arterial wall stiffness and inelasticity.
  • the "central augmentation index” or “cAlx” is the augmentation index as measured in the aorta.
  • the "peripheral augmentation index” or “pAlx” is the augmentation index derived from the waveform measured ata peripheral location. When it is not possible to measure cA!x directly, it can be estimated from pAlx by one skilled in the art.
  • Nitrite especially inorganic nitrite, as used in accordance with the present invention, may be used to alleviate pressure arising from elevated central pressure in a number of conditions. Such treatment may be sufficient, in itself, while a patient recovers from a condition, such as aortic dissection, or may be used in conjunction with known treatments for such conditions.
  • Suitable conditions are exemplified hereinbelow, and include conditions selected from: acute and chronic aortic dissection, impaired ventricular-vascular coupling, acute hypertensive heart failure, chronic hypertensive heart failure, diastolic heart failure due to hypertension, acute coronary syndromes, stroke due to occlusion of middle cerebral artery in children/adolescents, and adults with sickle cell disease.
  • Nitrite is also useful in the chronic management of hypertension, whereby the reduction of centra! systolic blood pressure or the dilation of arteries is sufficient to cause selective reduction of central haemodynamic indexes in order to reduce cardiovascular risk.
  • Nitrite can also be used in the chronic management of isolated systolic hypertension.
  • Nitrite may also be used during percutaneous coronary intervention (PCI), whereby the reduction of central systolic blood pressure or the dilation of arteries is of assistance in reducing the risk of peri-procedural infarction and/or to allow better implantation of one or more stents.
  • PCI percutaneous coronary intervention
  • Nitrite may be used for the dilation of large arteries supplying, or in, organs or organ systems to improve blood flow and function of that organ. Nitrite may be administered at such a dose as to produce a therapeutic decrease in centra! haemodynamic indices, which may include centra! systolic blood pressure, central pulse pressure or the central augmentation index, preferably in physiologically acceptable amounts.
  • Typical routes of administration of nitrite include intravenous, intracoronary, oral, buccal, and/or inhaled.
  • Nitrite may be provided as a pharmaceutically acceptable salt. More preferably, the cation is inorganic and selected from aluminium, caicium, potassium, sodium, magnesium, and zinc ions. The alkali metal salts are generally preferred. The nitrite is preferably soluble in water to at least a level of 10 ⁇ . Most preferably, nitrite for use according to the present invention is sodium nitrite.
  • the nitrite may be provided as a medicament comprising one or more pharmaceutically acceptably excipients.
  • Suitable administration forms may include: liquids, pastes, tablets, capsules, gums, lozenges, sprays, solutions, powders, suspensions, suppositories, pessaries, ointments, nebu!iser preparations, delayed release formulations, transdermal patches, and injectables.
  • the administration form may comprise one or more pharmaceutically acceptably excipients, including, as desired, one or more of, bulking agents, diluents, solvents, pH adjusting agents, buffers, antibiotics, sterilising agents, colourants, flavourings, and stabilising agents.
  • arteries is used to describe blood vessels that carry blood away from the heart, and are larger, and have a higher proportion of elastic fibres in their walls, than arterioles.
  • the oxygen saturation of haemoglobin is typically 97-100% in arteries.
  • “Arterioles” or “resistance arterioles” are the vessels that carry b!ood from the arteries to the capillaries and are part of the microcirculation, typically having a higher proportion of smooth muscle in their walls than arteries.
  • the oxygen saturation levels in haemoglobin in arterioles vary with exertion level and injury but, within the systemic circulation, are less than those of the arteries.
  • SBP cardiac blood pressure
  • systolic pressure is the maximum blood pressure measurable during the cardiac cycle.
  • Central systolic blood pressure or “cSBP” refers to the maximum aortic pressure measurable during the cardiac cycle.
  • Peripheral systolic blood pressure is the maximum blood pressure measurable by conventional blood pressure instrumentation in the arm.
  • Diastolic blood pressure (DBP) is the lowest blood pressure measured during the cardiac cycle.
  • Central diastolic blood pressure or “cDBP” refers to the lowest aortic pressure measurable during the cardiac cycle.
  • Peripheral diastolic blood pressure or “pDBP” is the lowest blood pressure measurable by conventional blood pressure instrumentation in the arm.
  • Pulse pressure or "PP” is the difference between systolic and diastolic pressure
  • MAP mean arterial pressure
  • central haemodynamic index refers to a measure of the circulatory system, and may include reference to any one the following: central systolic blood pressure, central pulse pressure, and central augmentation index.
  • “Hypertension” is a condition whereby an individual's blood pressure is persistently elevated above 139/89mmHg.
  • Ideal "ventricular/vascular coupling" is manifested as low pressure fluctuation in the ascending aorta.
  • Low pressure fluctuation results in pressure during systole being only slightly greater than pressure throughout the whole cardiac cycle, and pressure during diastole being only slightly less. This is desirable because pressure during systole determines ventricular output, in whichinotropic state and ventricular filling are constant, and ventricular metabolic requirement, while pressure during diastole in the ascending aorta is a major determinant of coronary blood flow.
  • CHD Coronary heart disease
  • Aortic dissection refers to a condition characterised by bleeding into and along the aorta wall.
  • Marfan syndrome is a genetic disorder of the connective tissue.
  • the main characterising feature in Marfan syndrome is aortic root dilation, which is aneurysm or vessel dilation at the base of the aorta, just above the aortic valve. This is associated with aortic regurgitation, dissection, and rupture, the major causes of morbidity and premature death. The exact mechanisms leading to dilatation are not fully understood, but steady and pulsatile stresses probably play an important role.
  • PCI Percutaneous coronary intervention
  • atherectomy whereby atherosclerotic plaques are excised from the artery wall
  • brachytherapy whereby an internally placed radioactive source is used to prevent restenosis
  • PCI is equivalent to coronary angioplasty or angioplasty.
  • Heart failure is a condition where the cardiac output of the heart is insufficient to meet the needs of the body.
  • Hapertensive heart failure is heart failure for which hypertension is a major causative factor.
  • Diastolic heart failure refers to decline in the ability of one or both of the ventricles to fill with blood during diastole.
  • ACS acute coronary syndrome
  • Myocardial infarction or "heart attack” refers to an event that leads to the interruption of the blood supply to the heart, resulting in myocyte death.
  • ST elevation myocardial infarction or "STEM!" refers to myocardial infarctions whereby the ST segment on the echocardiogram is elevated and non-ST elevation myocardial infarction or “non- STEMi” refers to myocardial infarction in the absence of an elevated ST segment.
  • Angina refers to severe chest pain due to ischaemia of the heart muscle.
  • “Dilator” refers to an agent or stimulus that can induce dilation, or dilatation, both of which terms and their related terms, are used interchangeably herein.
  • “Vasodilator” is an agent or stimulus that is able to induce the dilation of a target blood vessel, the term “vasodiiatory” to be construed accordingly.
  • Figure 1 shows the selectivity of increasing doses of the alpha-adrenergic antagonist phentolamine, the nitrovasodilators nitroglycerin (GTN) and sodium nitroprusside (SNP), and hydralazine on the diameter of the radial artery, a muscular conduit artery. Change in diameter of the radial artery is represented on the y axis;
  • Figure 2C shows peripheral Brachial Blood Pressure measurements during 60 min intrabrachial infusion of sodium nitrite (systolic, SBP, mean arterial, MAP, or diastolic, DBP);
  • FIG. 3C shows central Systolic Blood Pressure (cSBP) before and after 60 min infusion of sodium nitrite (8.7 ⁇ /min for 60 min), (*P ⁇ 0.05);
  • Figure 3D shows the peripheral augmentation index before and after 60 min infusion of sodium nitrite (8.7 ⁇ / ⁇ for 60 min) ( * P ⁇ 0.05);
  • Figure 4B shows peripheral Brachial Blood Pressure measurements during intra-arterial infusion of higher dose range sodium nitrite (8.7-87 ⁇ / min), (systolic, SBP, mean arterial, MAP, or diastolic, DBP);
  • Figure 5A shows the dose response for plasma nitrite concentration in infused arm during nitrite administration
  • Figure 5B shows the dose response for plasma nitrite concentration in infused arm during nitrite administration
  • Figure 6C is a comparison of radial artery dilatation during intra-arteria! infusion of GTN (1 pg/min) and nitrite (8.7 pmo!/min);
  • Figure 7 shows individuals' responses in terms of change in forearm blood flow (FBF) and radial artery diameter during the dose response studies, expressed independently of dose, with (A) nitrite and (B) GTN. Dashed lines represent 95% confidence intervals; and
  • Figure 8 shows the change in radial artery diameter versus change in FBF at each dose step of (A) nitrite and (B) GTN. Dashed lines represent 95% confidence intervals.
  • Nitrite is capable of causing marked dilatation of arteries under fully oxygenated conditions (haemoglobin oxygen saturations of ⁇ 99%) ( Figure2A and 3A). This is particularly surprising, and shows that the accepted model of action of nitrite, involving reduction by haemoglobin at its P 50( is either not correct, or that there is a further mode of action.
  • the discovery on which the present invention is based means that nitrite can now be used in therapy, to reduce cSBP, and there is no need to rely on, or to induce, reduced oxygen tension or ischaemic conditions.
  • nitrite is most effective at dilating conduit arteries under normal oxygen tension, with significant inhibition occurring under hypoxic and hyperoxic conditions, so that this action of nitrite appears to be optimised under normal physiological conditions.
  • This effect in conduit arteries in hypoxia is opposite to that seen in resistance arterioles (forearm blood flow) where dilatation to nitrite is enhanced.
  • nitrite This dilatory action of nitrite is highly selective for arteries, having little or no effect on the resistance arterioles. This is similar to GTN, before the induction of tolerance. In the case of nitrite, tolerance is not induced at physiological acceptable levels. While GTN is a selective large artery dilator, its drawbacks, as described above, and which include headaches and the rapid induction of tolerance, mean that it is only prescribed when the need is acute. Nitrite, on the other hand, does not induce headaches or tolerance, and can be used in chronic situations, and has greater large artery selectivity overall (Figure 8A and B), compared to GTN.
  • the dilatory effect of nitrite is not limited to the local site of administration, as nitrite is also effective systemically.
  • Figure2A we demonstrate dilation of the radial artery when nitrite was administered intra-arterial!y (Figure2A), and dilatation of the contralateral radial artery by 9% when nitrite was administered intravenously, at a surprisingly low dose rate of 8.7 mol/min for 60 minutes (Figure3A).
  • nitrite of 87 ⁇ / ⁇ resulted in a systemic concentration of -42 ⁇ , but some subjects had a reduction in MAP >15 mmHg and the procedure had to be stopped.
  • plasma concentrations of nitrite it is preferred to limit plasma concentrations of nitrite to ⁇ 5 ⁇ , preferably ⁇ 3 ⁇ , for chronic treatment or therapy, but plasma concentrations of up to 50 ⁇ , preferably no more than 40 ⁇ , may be employed for acute situations, such as where it is desired to have an immediate and substantial effect, although it is preferred to allow plasma concentrations to drop to those preferred for chronic administration in no more than an hour, and preferably inside 30 minutes, preferably inside 20 minutes. This is generally not a particular problem, as the half-life of nitrite is about 45 minutes.
  • the oral bioavailability of sodium nitrite is generally in excess of 95%. Accordingly, nitrite may be administered orally in amounts largely equivalent to the systemic quantity required.
  • the T m3X time to maximum concentration
  • Lower doses may be associated with lower bioavailability.
  • an oral dose of 40 mg is indicated, although it will be appreciated that uptake is dependent on many parameters including age, time of day, health, and whether the subject is eating or has recently eaten.
  • an oral dose of 72 mg an oral dose of about 80 mg is indicated.
  • nitrite concentrations are -0.3-3 ⁇ . These levels are preferably attained with doses of nitrite between 0.87 - 8.7 pmol/min, assuming a basal plasma nitrite concentration of -0.05 ⁇ . A more preferred range is 0.5-1.5 ⁇ . These ranges are considerably lower than the nitrite concentration of 10 ⁇ which would have non-selective effects and lower MAP.
  • Benefit is seen with doses as low as 0.087 pmol/min, achieving a concentration of -0.1 ⁇ , for example. Conversely, with a short infusion of 20 min, for example, higher dose rates may be administered with no changes in MAP, resulting in temporarily high systemic plasma concentrations of up to -100 ⁇ , although this is unlikely to be useful for prolonged use.
  • oral doses will be similar to intravenous doses.
  • a preferred range of nitrite is administered at between 0.087-8.7 ⁇ /min. Administration for 60 min at this rate equates to a dose range of 0.36-36 mg.
  • the administration may be as single doses of fast release nitrite, or in a slow release formulation, with nitrite being released at a rate of 0.36-36 mg/h, for example.
  • nitrite at 2.6 pmol/min for 20 min also reduced cPP by ⁇ 6.5 mmHg, and was associated with a similar increase in nitrite concentration to ⁇ 1 pmoi/L ( Figure 5B). Accordingly, it appears that administration of nitrate is not a preferred method of increasing plasma nitrite concentration for the purposes of the present invention.
  • nitrite has been found to be effective in lowering cSBP ( Figure 3C) at a dose not associated with any change in peripheral BP.
  • nitrite is capable of selectively lowering cSBP, in this case by approximately 12 mmHg, but with no effect on peripheral BP.
  • GTN is administered at a dose producing a similar local dilatation of the radial artery, there is no effect on cSBP.
  • nitrite has greater large artery selectivity than GTN, and this represents a substantial clinical advantage of nitrite over GTN. Whilst there may also be some effect on the myocardium, reducing cSBP while leaving DBP substantially unchanged reduces cPP and the pressure load on the left ventricle, thereby enhancing ventricular/vascular coupling. This is a particular advantage of nitrite over GTN.
  • Nitrite is further capable of reducing peripheral and central A!x.
  • nitrite significantly reduced the peripheral augmentation index (pAlx) by 11.9 ⁇ 4.6%, while GTN had no effect (Figure3D).
  • cAlx central augmentation index
  • levels of nitrite that are effective in the present invention have not been associated with headaches, which has been one significant factor mitigating against the use of GTN, and represents an advantage of the present invention.
  • ALDH-2 may also be involved in the bioactivation of nitrite, and increasing the activity of this receptor may partly explain the lack of tolerance induction with nitrite.
  • nitrite is associated with a reduction in reactive oxygen species in a model of renal ischaemia-reperfusion, a further possible factor in organic nitrate tolerance.
  • nitrite is useful to replace GTN in vascular therapy where arterial dilation is desirable or required, particularly where the selective dilation of arteries or a reduction in cSBP without lowering cDBP is desirable.
  • nitrates are not preferred for chronic indications
  • the present invention provides combinations of nitrite with an organic nitrate, with suitable formulations including GTN, isosorbidemononitrate, and/or isosorbidedintrate, such as a GTN patch, oral isosorbide mononitrate, or oral isosorbidedinitrate.
  • the present invention is particularly useful in the medical management of acute and chronic aortic dissection, where selective reduction in central aortic SBP/cPP/cAlx, is beneficial, !t is known in the art that the central indices cSBP, cPP and cAix are positively correlated with the occurrence and progression of aortic aneurysm or aortic dissection, particularly in patients with Marfan or Marfan-like syndromes. Because of the selectivity for arteries and, thereby, cSBP/cPP/cAix, a general reduction in blood pressure may substantially be avoided by the use of nitrite in accordance with the present invention. Substantia! maintenance of DBP and MAP helps to support perfusion of vital organs. For example, patients with aortic dissection benefit from strict blood pressure control over several days, and the administration of nitrite according the present invention is useful to help to reduce the risk of further dissection.
  • central haemodynamic indices including cSBP, cPP, and cAlx
  • cSBP central haemodynamic indices
  • cPP central haemodynamic indices
  • cAlx being capable of being used to predict clinical events independently of peripheral pressures.
  • the present invention accordingly, further provides the use of nitrite in the modification of one or more of these indices to reduce the risk of adverse clinical events and/or in the prophylaxis or treatment of conditions associated with or aggravated by hypertension, such as aneurysms.
  • nitrite in accordance with the present invention is also useful when treatingvarious conditions that can be exacerbated by high central blood pressure, such heart failure and/or impaired ventricular-vascular coupling.
  • Suitable conditions particularly include acute hypertensive heart failure and chronic hypertensive heart failure, including diastolic heart failure due to hypertension.
  • Ventricular/vascular coupling becomes less than ideal in adult humans and is attributable to progressive arterial degeneration, which is known to commence in childhood and is apparent in the elderly as dilated tortuous arteries, high pulse pressure, and high likelihood of developing ventricular failure.
  • the present invention further provides nitrite for use during percutaneous coronary intervention (PCI), a therapeutic procedure known more commonly as coronary angioplasty or angioplasty, which is used to treat stenotic, or narrowed, arteries of the heart.
  • PCI percutaneous coronary intervention
  • PCI includes several related procedures including balloon angioplasty, atherectomy, wherein atherosclerotic plaques are excised from the artery wall, the implantation of stents, and brachytherapy, wherein an internally placed radioactive source is used to prevent restenosis.
  • Nitrite may be used in accordance with the present invention for the reduction of peri-procedural infarction in PCI. In particular, this allows better implantation of balloons, stents and radiotherapy sources.
  • nitrite in accordance with the present invention is also useful during the prophylaxis and/or treatment of various cardiovascular conditions, examples of which include, but are not limited to: the chronic management of hypertension, particularly the selective reduction of central aortic SBP/cPP/cAix to reduce cardiovascular risk; chronic management of isolated systolic hypertension, which has been associated with an increased risk of acute coronary syndromes, and for which it is desirable to selectively reduce systolic pressure in preference to diastolic BP; acute coronary syndromes, including both ST elevated myocardial infarction (STEM! and non-STEMI; and stroke due to occlusion of middle cerebral artery in children or adolescents and aduits with sickle cell disease, to assist in dilating the middle cerebral artery and restore blood flow.
  • various cardiovascular conditions examples of which include, but are not limited to: the chronic management of hypertension, particularly the selective reduction of central aortic SBP/cPP/cAix to reduce cardiovascular risk; chronic management of isolated sy
  • the present invention may also be used to induce dilatation of large arteries associated with a selected organ to improve blood flow and function of that organ.
  • nitrite may be administered to improve bloodflow to the kidneys or liver.
  • treatments of the conditions described herein are by reducing elevated central systolic blood pressure and/or elevated central pulse pressure, and will generally serve to reduce the stress on the condition in question that would otherwise be present through high central blood pressure. As such, it will generally also be entirely appropriate to administer any other medicaments normally administered for the condition concerned, unless the medicament in question is contra-indicated, or is likely to have an adverse reaction with nitrite.
  • nitrite surprisingly, has been found to possess potent effects under local fully oxygenated conditions. As such, - it is not necessary for medicaments comprising nitrite according to the invention to contain reducing agents, although these may be provided if desired.
  • reducing agents include hawthorn berry extract and vitamin C, for example.
  • nitrite has an effect on large arteries is surprising, as the art ascribes the action of nitrite to the reduction of nitrites to NO via deoxyhaemoglobin. As such, it has been assumed that the action of nitrite is inhibited by fully oxygenated haemoglobin, both because of a lack of deoxyhaem sites and also because of the oxidation of nitrite to nitrate by oxyhaemoglobin (Cosby et al. Nat Med, 2003; 9(12): 1498-505, Maher ef a/. Circulation, 2008;117:670-677).
  • Nitrite-induced dilation of the arteries under fully oxygenated conditions may possibly occur through a direct effect on the arterial wall, and seems unlikely to be dependent on an initial bioconversion to other NO species, such as nitrosothiols, as there appears to be no opportunity for the nitrite to be exposed to the necessary reduced oxygen conditions within the microcirculation or venous circulation in the timescale available.
  • the needle On successful cannu!ation the needle was secured in position and a saline infusion was commenced at a rate of 1 ml/min (InjectomatAgilia syringe driver, Fresenius Kabi, Homburg, GR).
  • injectomatAgilia syringe driver Fresenius Kabi, Homburg, GR.
  • the wrist cuffs were inflated to supra-systolic pressure (180 ⁇ 2 mmHg) to exclude the circulation in the hand. After a 30-60 s pause to allow the changes in the forearm circulation to reach equilibrium, the cuffs around the upper arm were inflated to 40 ⁇ 2 mmHg) and deflated a number of times in cycles consisting of ⁇ 0 s inflation and ⁇ 5 s deflation.
  • the degree of change in the circumferential size of the forearm measured by the mercury-in- silasticstrain gauge was acquired by channelling the signal through the EC-6 Pleihysmograph, and recorded and analysed using Chart 5 software to give a value of FBF in mi of blood / 100ml tissue / s.
  • the venous occlusion plethysmography technique was adapted to be able to measure venous capacitance.
  • the radial artery was imaged using an Acuson-Aspen advanced ultrasound (US) probe (Siemens GR) which was fixed in position using a magnetic flexible stand (Mitutoyo JP). The image was acquired for 120 s at a rate of 1 frame every 3 s giving a total of 40 frames per sequence of acquisition.
  • the radiai artery diameter US was analysed using dedicated vascular analysis software (Brachial analyzer, Medical Imaging Applications) which gave a measurement of the diameter of the artery for each of the 40 frames, and an average of the measurements obtained was used to calculate the vessel diameter at that time point.
  • nitrate-rich foods green leafy vegetables, beetroot and processed meats
  • caffeine for 24 hours prior to the study and to fast overnight (though were encouraged to take clear fluids to avoid the risk of dehydration). They were also advised to avoid smoking and heavy exercise in that time period.
  • the studies were conducted at a similar time of the day, around 11 :00 am under controlled temperatures 26.6 ⁇ 0.9°C and with minimal sensory stimulation to minimise confounding.
  • the samples were thawed and stored on ice.
  • the plasma levels of nitrite were analysed using the 280i Nitric Oxide Analyzer (NOA) (Sievers Instruments, GE analytic instruments).
  • NOA 280i Nitric Oxide Analyzer
  • the Purge vessel was filled with sodium iodide dissolved in 99.8% glacial acetic acid to which a few drops of antifoaming agent were added.
  • To generate a calibration curve a stock solution of 100 mM sodium nitrite was used to prepare the standards which were used in generating the calibration curves.
  • the series of standard dilutions was constructed by serial dilution of the stock solution giving concentrations of 100 nM, 0.5 ⁇ , 1 ⁇ , 5 ⁇ , 10 ⁇ , 50 ⁇ , 100 ⁇ .
  • HPLC nitrite-free water was used in the preparation of the stock and standard solutions.
  • 50 ⁇ _ samples of selected dilutions were injected, in duplicate, into the purge vessel directly onto the reducing agent and the NO generated was detected by the NOA and analysed by the software to generate the calibration curve. To maintain consistency, close attention was paid to the volume of reducing agent, the maintenance of the cell pressure and the maintenance of the gas pressure.
  • 50 ⁇ _ of the plasma samples were injected, in duplicate, into the purge vessel. The reducing agent in the purge vessel was changed after each duplicate. The concentrations of nitrite were derived from the calibration curve.
  • Intra-arterial infusion of sodium nitrite (8.7 ⁇ / min) into the brachial artery resulted in a rapid, marked (-30%) and highly significant dilatation of the radial artery within 5 min (P ⁇ 0.001 compared to baseline saline, Figure 2A). This degree of dilatation was almost maximal by 5 min, as no further significant dilatation compared to 5 min was demonstrated.
  • the average oxygen saturation of haemoglobin was 99% (range 97-100%), i.e.
  • FBF forearm blood flow
  • nitrite is capable of causing a significant arterial dilation of around 30%, without causing any significant drop in peripheral blood pressure. Without being bound by theory, it also appears that the effects of the present invention cannot be dependent on the conversion of nitrite to NO or another nitro-species in the micro-vascular beds.
  • Sodium nitrite was not only effective following direct intra-arteriai administration, but was a!so effective in dilating the radial artery following the intravenous infusion of the same dose of sodium nitrite into the contralateral forearm. Since the concentration in the contra-lateral artery depends on systemic intravenous administration rather than direct intra-arterial administration, the concentration of nitrite necessarily takes longer to build up, as it is determined by overall systemic concentration. Thus, the degree of dilatation was less and did not become significantiy different to baseline until 30 min, peaking at 9 ⁇ 3% at 45 min (P ⁇ 0.01). The results are shown in Figure 3A.
  • intravenous sodium nitrite did not result in any significant changes in peripheral brachial blood pressure (SBP, MAP, or DBP) during the 60 min infusion of sodium nitrite compared to baseline, (time 0 min) ( Figure 3B).
  • SBP peripheral brachial blood pressure
  • MAP MAP
  • DBP peripheral brachial blood pressure
  • peripheral augmentation index was also decreased by 11.9 ⁇ 4.6% (P ⁇ 0.05).
  • the results are shown in Figure 3D.
  • the profile of the increase in systemic plasma nitrite concentration with intravenous nitrite was similar to the intra-arterial study (see Figure 3E).
  • the baseline concentration appeared higher (379+65 nmol/L, compared to 64 ⁇ 19 nmol/L previously) owing to the use of a different method of calculation, requiring the deproteination of plasma using filters, which is typically associated with higher levels in this range.
  • the concentrations are higher than in the i.a. study shown in Fig 2D, and may be due to contamination through the iv canula used to infuse nitrite and collect samples, suggesting that the concentrations in the i.a. study (Fig 2D) are more representative.
  • nitrite is effective at inducing systemic arterial dilation, and that nitrite can significantiy reduce central systolic blood pressure (SPB) by at least 12 mmHg and the percent measure of the peripheral augmentation index (pA!x) by around 12%, without significantly reducing peripheral SBP.
  • SPB central systolic blood pressure
  • pA!x percent measure of the peripheral augmentation index
  • peripheral augmentation index and cSBP were slightly lower (5.8 ⁇ 7.6% and -1.5 mmHg respectively) with low dose nitrite, these differences were not significant. Following high dose nitrite, cSBP was reduced by ⁇ 18 mmHg.
  • Figures 5Aand 5B show the dose response for plasma nitrite concentration in infused and contralateral arm during nitrite administration respectively.
  • nitrite has a clear dose-response method of operation. No significant changes in peripheral blood pressure were seen up to a nitrite dose of to 26 pmol/min.
  • the dose response to GTN used six doses (0.003-1 pg/min), which resulted in a similar range of radial artery dilatations as sodium nitrite (-10-30%). The results are shown in Figure 6A.

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Abstract

Nitrites, such as sodium nitrite, are indicated as arterial dilators and act selectively to central systolic blood pressure without reducing central diastolic blood pressure.

Description

VASCULAR TREATMENTS
The present invention relates to the use of nitrites as vasodilators, and to their use in the treatment and prevention of cardiovascular conditions.
According to the World Heath Organisation, cardiovascular disease (CVD) represents the number one cause of global mortality. One of the most significant risk factors associated with the development of CVD, or in suffering reoccurring cardiovascular events and complications, is high blood pressure. Worldwide, around 40% of adults over the age of 25 have elevated blood pressure.
As such, there is a need for effective therapies both to mitigate chronic hypertension and to control blood pressure following acute cardiovascular events, in order to minimise further damage. In either case, it is often desirable to lower central aortic blood pressure, without lowering peripheral pressure. While reducing central pulse pressure (cPP) would help to protect against damage to the arteries, the vast majority of drugs that are currently available to reduce blood pressure have a limited effect on cPP,as they cause a fall in diastolic blood pressure (DBP) and mean arterial pressure (MAP, which are similar between centra! and peripheral locations), which can impair perfusion and aggravate any tissue hypoxia. Indeed, some drugs such as beta blockers having no/little effect on central aortic blood pressure and offer less protection than drugs which have more effect on central aortic blood pressures (Williams B, et al: principal results of the Conduit Artery Function Evaluation (CAFE) study. Circulation 2006 March 7;113(9):1213-25).Thus, therapies capable of selectively reducing cPP without lowering peripheral blood pressure are highly sought after.
Interest in vasodilator drugs, in general, has focused almost exclusively on their action on the small blood vessels in the periphery, known as the resistance vasculature,and their efficacy in reducing peripheral blood flow resistance.
Central aortic systolic blood pressure (cSBP) and pulse pressure (PP), the latter being the difference between systolic blood pressure (SBP) and diastolic blood pressure (DBP), are of importance in determining cardiovascular events, but less attention has been focussed on these parameters, as there is greater difficulty in establishing central readings, given that simple cuffs cannot measure these parameters, but also because it is generally assumed that there is a one ΐο one relationship between central and peripheral readings. However, evidence is emerging that central indices such as cPP are independent predictors of cardiovascular events (Vlachopoulos C, et al., Eur Heart J (2010) August;31 (15): 1865-71).
In a population of 2,405 nativeAmericans, hypertension was present in 52% of the population, of whom 68% were taking antihypertensive medications.A central PP >50 mm Hg, including some 23% with normal blood pressure, was associated with double the event rate, compared to a central PP <50 mm Hg, (the Strong Heart Study, Roman et al. Hypertension 2007
July; 50(1): 197-203).
Furthermore, about 70% of individuals with high normal blood pressure have similarly elevated central aortic blood pressure as individuals with hypertension (McEniery CM, et al: the Anglo- Cardiff Collaborative Trial El. Hypertension 2008 June;51 (6):1476-82).
Organic nitrates, such as glyceryl trinitrate (GTN), are among the most commonly used drugs for the treatment of cardiovascular disease. They have wide therapeutic application, and are effective venodi!ators and also dilators of coronary arteries in the treatment of acute coronary syndromes. They are, however, used less frequently to control BP and, when they are, this is usually in the acute setting, as their long term effects are rapidly attenuated by the induction of tolerance.
Organic nitrates selectively lower centra! BP (cBP) over peripheral BP (pBP), an effect that may be due to their high efficacy in dilating large arteries. We have shown that the dilatory effect can occur separately from the effect on BP, and that at elevated levels, central SBP and central DBP are both reduced to a similar extent, reducing mean arterial pressure (MAP), but not necessarily central pulse pressure (cPP).
A recent, unpublished study on potential large, artery dilators (P. Chowienczyk) compared the effects of intrabrachial arterial infusions of increasing doses of the alpha-adrenergic antagonist phentolamine, the nitrovasodilators nitroglycerin (GTN) and sodium nitroprusside (SNP), and hydralazine, on the diameter of the radial artery, a muscular conduit artery. The results demonstrated that the nitrovasodilators, and in particular, GTN, had the greatest dilatory effect on radial artery diameter for a given change in forearm blood flow, /.e.GTN demonstrated the greatest selectivity for large arteries (see Figure 1). The usefulness of the organic nitrates is markedly limited by the rapid induction of tolerance. Mechanisms of nitrate tolerance development include increased production of reactive oxygen species (ROS), possibly through premature release of partially reduced oxygen from mitochondrial complex I or III, initiation of lipid peroxidation, depolarisation of mitochondrial membrane potential, mitochondrial swelling, resulting in the oxidation of thiol groups which may cause inhibition of several enzymes including (i) aldehyde dehydrogenase 2 (ALDH-2), which is important in the bioactivation of GTN, and (ii) soluble guany!ylcyclase (sGC), thus impairing NO signal transduction.
In respect of coronary artery disease, nitrate tolerance manifests as reduced treadmill walking time and reduced time to onset of angina. In congestive heart failure, it has been described as the loss of haemodynamic effect of the administered nitrate, and in hypertension, it is evident as the rapid loss of the hypotensive effects. In a study by Elkayamei a/. [Circulation 1987; 31 (9): 1727-35], the initial significant reduction in pulmonary artery wedge pressure caused by a continuous GTN infusion was no longer present at 12 hours, and by 20 hours there was a complete loss off effect, even to being numerically worse than placebo. As a result of this problem, patients are often denied the beneficial haemodynamic effects of nitrates for several days, with the associated risk arising from failure to address the elevated SBP, while the skilled physician seeks to establish the optimal window in which to administer GTN.
Patients with acute aortic dissection have been successfully managed in the short term with intravenous GTN. This approach is supported by a review (Vaderaef al. Ann Emerg Med 2011 ; 57(1}:64-5) which states that, "Pending further trials, use of [organic] nitrates to acutely decrease blood pressure in patients presenting with a hypertensive emergency is reasonable".
Aortic dissection is a condition characterised by bleeding into and along the aorta wall. This results from a tear or damage to the inner wall of the aorta. Such patients need very strict blood pressure control over several days to reduce the risk of further dissection. However, because GTN becomes ineffective after less than 24 hours, continuous good blood pressure control is frequently not possible and further dissection is common. A further issue is that, whilst it is imperative to lower SBP, it is important not to excessively lower mean arterial pressure (MAP), as this is crucial for maintaining adequate perfusion to vital organs such as the kidneys. The kidneys are at particular risk as the dissection often involves the renal arteries, and so their blood flow may already be compromised. GTN is therefore potentially useful, in the short term, through its selective large, artery effect, but once tolerance has occurred, it becomes necessary to rely on alternative antihypertensive agents. Most other antihypertensive medications have greater effects on resistance vessels and will cause a more marked reduction in MAP. Furthermore, whilst heart rate control agents, such as beta-blockers (e.g. intravenous !abeta!ol), have been demonstrated to improve outcomes, they actually increase cPP, while lowering MAP and peripheral biood pressure. Treatment that could reduce cSBP without reducing cDBP, would be particularly useful, especially if there were little or no effect on the pBP.
In patients with aortic dissection or aortic aneurysm, ~50% have an elevated central aortic systolic blood pressure despite optimally controlled/normalised (peripheral) systolic blood pressure.lndeed, even when excellent control of peripheral systolic BP is achieved (-118 mmHg), only patients with good control of central SBP (<106 mmHg) have been found to avoid further complications over 5 years, which occurred in all patients with elevated cSBP (>106 mmHg) (Moon Jet al, Acta Cardiol 2010 June;65(3):303-8).
A further problem with organic nitrates is that they frequently cause headaches, a very common side effect that can be very unpleasant for patients and sometimes limits the use of GTN.
There is also increasing evidence that long term use of nitrates increases the risk of suffering a cardiovascular event. Kosugi M et al. Circ J. 2011 Jun 14, describe patients who received organic nitrate therapy in addition to calcium channel blockers, were found to suffer a higher rate of cardiac events than those patients who received calcium channel blockers alone. Yiu KH er a/. (CardiovascDiabetol. 2011 Jun 13;10(1):52) followed the impact of long term use of nitrate (isosorbide mononitrate [ISMNj) in patients with type II diabetes following percutaneous coronary intervention (PCI), and found that nitrate use was an independent predictor for the development of major cardiac adverse events.
Until recently, the nitrite anion (N02 ~) was considered to be inactive in the circulation, and was present merely as an oxidation product of nitric oxide (NO) under normal physiological conditions. However, in 2003, Cosby et al. (Nat Med 2003 December;9(12): 1498-505) demonstrated that infusion of nitrite into the forearm brachial artery at two different doses, 36 μηιοΙ/min, and a 100-fold lower dose (0.36 μιηοΙ/min), increased forearm blood flow (FBF), in both cases, and also resulted in supra- and near-physiologic intravascular nitrite concentrations, respectively. These effects were demonstrated to be as a result of the generation of NO in the resistance vasculature. At the physiologically toxic levels of 1000 pmoi sodium nitrite and at 95% 02 saturation, rabbit arterial slices were shown to dilate, although no effect was noted at a lower concentration of nitrite.
It is now generally recognised that vascular nitrite can serve as the largest source of nitric oxide (NO), provided that physiological mechanisms exist to reduce nitrite to NO. Reduction of nitrite to NO is generally considered to be increased under conditions of hypoxia and ischaemia, by processes beyond simple chemical acidification/disproportionation. It is generally accepted that one of the most important mechanisms is reduction by deoxyhaemoglobin. This reaction has been well-characterised: the maximal rate of nitrite reduction and NO generation occurs at the P50 of haemoglobin, the oxygen concentration at which half the haem is saturated with oxygen (Cosby et al., supra). This thesis is supported by in vitro studies of the effect on nitrite on rat aortic rings, which found that in the presence of red blood cells, the aorta began to dilate when the partial pressure of oxygen (p02) dropped to a level corresponding to oxyhaemoglobin saturation of 80%. In the absence of red blood cells, the aorta did not dilate until the p02 dropped to a level corresponding to an oxyhaemoglobin saturation of around 0%. Furthermore, in the presence of free oxyhaemoglobin, the dilatory activity of nitrite on the aorta was shown to be reduced even below that of nitrite alone (i.e. in the absence of red blood cells or haemoglobin). Other studies suggest that, in fully oxygenated conditions, some oxyhaemoglobin-dependent oxidation of nitrite to nitrate (N03 ") takes place. aher et al., (2008) demonstrated that, under normoxic conditions, exogenous nitrite-induced vasodilatation in humans occurred predominantly in capacitance vessels (veins, associated with the lowest oxygen tension), with only a modest effect on small arteries (measured as changes to forearm blood flow (FBF)). Also, under hypoxic conditions (12% oxygen), rather than room air (21 % oxygen), FBF responses to nitrite were enhanced, but there was little additional effect in terms of venodilatation, which suggests that optimum conditions for nitrite reduction (around the P60) are already present under normal physiological conditions in the veins. In addition to haemoglobin, the other globins (myoglobin, cytoglobin and neuroglobin) have also been shown to possess nitrite reductase activity, which increases as the oxygen saturation falls.
Whilst physiologically acceptable levels of nitrite have been shown to have a substantial venodilatory effect, and less but significant resistance arteriole dilatory affect, this has been shown to be dependent on hypoxia, and increases forearm blood flow while having no effect on pBP or MAP. As such, nitrite has not been popularly pursued in relation to general treatment of hypertension, and known uses for nitrite are dependent on, or are targeted at, conditions associated with hypoxia or ischemia. For example, one formulation currently under development is aerosolised sustained release nitrite for the treatment of pulmonary arterial hypertension that will work through inducing smail vessel dilation in the deoxygenated conditions of the pulmonary arterial circulation. Another product under development provides nitrite as an oral preparation for the selective increase of blood flow to ischaemic tissues in peripheral artery disease.
Ingram T er al. (Oxygen Transport to Tissue XXX, 21-25, 2009), used rabbit aorta, inferior vena cava and pulmonary artery to show the relaxing effects of 10μΜ sodium nitrite. Such aorta preparations are frequently used to test dilators and constrictors but are not representative of their likely effect on large arteries in humans. Indeed, the test is more representative of their likely action in small arteries and resistance vessels. Dilators that are poorly selective for large arteries, and which are effective in resistance vessels in humans, such as phentolamine and verapamil, are very effective in rabbit aorta preparations. Such aorta preparations are also very artificial, as they lack red blood cells with the concomitant nitrite-scavenging oxyhaemoglobin, and flow resulting in shear stress and the resulting generation of endogenous NO production, but also because the rings are put under tension and then phenylephrine is applied to pre- constrict the vessel. Accordingly, even if such tests couid be considered predictive of aortic performance in humans, the indications are that a considerably higher concentration than 10μΜ of nitrite would be required in order to demonstrate any effect.
The concentration of nitrite (10 μΜ) used in Ingram T et a!., (supra) is too high a systemic concentration to be sustained for any period of time, such as would be required in the chronic treatment of hypertension. Pluta R et al., (Plos One 2011) infused different doses of sodium nitrite for 48 h in healthy volunteers and established that the maximum tolerated dose (MTD) was 266.9 pg/kg/h, equivalent to approximately 4.5 pmol/min and resulted in a systemic nitrite concentration of ~0.56 μΜ and no change in MAP. Dose-limiting toxicity, in terms of a drop in MAP greater than 15 mmHg, occurred with increases in plasma nitrite to ~4.8 μΜ, less than 50% of the levels used in vitro by Ingram. Given that higher levels would be expected in order to yield similar results in vivo, this would place requisite levels for physiological activity in the toxic range. Jung et al., Stroke 2006;37:2744-2750 "Early Intravenous Infusion of Sodium Nitrite Protects Brain Against In Vivo Ischemia-Reperfusion Injury", showed that sodium nitrite enhanced local cerebral blood flow in ischaemic rats. No effect on blood pressure was demonstrated.
Mack et al., Br. J. Haematol. 2008;142{6):971-978 "Sodium nitrite promotes regional blood flowin patients with sickle cell disease: a phase l/ll study", studied the effect of nitrite on small resistance arterioles, where forearm blood flow was measured using venous occlusion plethysmography. No effect onblood pressure was demonstrated.
Montenegro et al., Free Radic. Biol. Med. 2011 ;51(1):144-152 "Sodium nitrite..downregu!ates vascular NADPH oxidase and exerts antihypertensive effects in hypertension", showed a reduction in peripheral BP of rats using a tail cuff, due to dietary sodium nitrite.
Haas M et al., Arzneimiitelforschung. 1999 ;49(4):318-323 "Persistent antihypertensive effect of oral nitrite supplied up to one year via drinking water in spontaneously hypertensive rats", showed a persistent antihypertensive effect of oral nitrite supplied up to one year via the drinking water in spontaneously hypertensive rats.The effect of nitrite on large/condutt arteries or central pressures is not assessed.
US 2006/182815 discloses treatment of cerebral artery spasm in the context of a stroke with hypoxia/ischaemia.
WO 2005/004884 discloses the prevention of vasospasm by nitrite due to a reaction with perivascular haemoglobin, i.e. in the cerebrospinal fluid around the vessels, where the nitrite level was found to be increased.
Schnaar R. et al., ., "Response of large and small coronary arteries to nitroglycerin, NaN02, and adenosine." Am J Physiol. 1972; 223: 223-228, showed that helical strips cut from coronary arteries taken from dogs relaxed when exposed to sodium nitrite (0.145-450 pmol/L). The conditions were not physiological, involving salt solution in a muscle bath, with a 10-fold supraphysiological concentration of potassium of 35 mM to pre-constrict the tissue in order to be able to observe a dilatory effect of nitrite.
WO 2011/095389 discloses the use of theobromine for lowering centra! blood pressure. Thus, there remains a need to identify vasodilators that overcome one or more of the above- identified problems.
Surprisingly it has now been found thatinorganic nitrite, at physiologically acceptable levels, is capable of reducing central systolic blood pressure and dilating the major arteries, without reducing central diastolic blood pressure, and without affecting peripheral blood pressure.
Thus, in a first aspect, there is provided nitrite for use in the treatment of patients having one of elevated central systolic blood pressure and elevated central pulse pressure, or both, wherein the patient has normal peripheral blood pressure.
The term 'elevated' is used to mean a blood pressure reading that is determined by the skilled person, preferably a physician, to be higher than is desired, whether by comparison with the norm, taking into account such factors as the patient's age, sex, and health, or for other reasons, such as acute aortic dissection, for example, or for any other reason, including published guidelines from learned societies or published reference ranges.
In an alternative aspect, there is provided nitrite for use in the reduction of, preferably elevated, central systolic blood pressure.
It is preferred that the level of nitrite does not exceed a systemic level of 10 μΜ for up to 20 minutes, and for periods longer than 2 hours, preferably about 1 hour, and more preferably not for more than about 20 minutes. It is preferred that the level of nitrite not exceed 5 μ systemically, with levels of between 0.3 μ and 3 μΜ, inclusive, being most preferred. Chronic levels of 5 μΜ nitrite and below are referred to herein as physiologically acceptable levels, or chronic physiologically acceptable levels. Acute physiologically acceptable levels can range as high as 100 pM, but the higher the concentration, the shorter the acceptable duration, which will be readily apparent to those skilled in the art.
A preferred position for a patient to adopt is the supine position. The supine position is preferred for patients receiving i.v. infusion. For treatment regimens involving the supine position, then patients may receive an oral administration form prior to a period of sleep, for example, especially as nocturnal blood pressures are more strongly associated with cardiovascular events than daytime blood pressures.
Nitrite has been found to selectively reduce central systolic blood pressure in preference to peripheral blood pressure. It is an advantage that nitrite selectively targets large arteries over resistance arterioles, it is a further advantage that nitrite does not induce tolerance/resistance with long term use. It is a yet further advantage that nitrite treatment is not generally associated with any adverse reactions, such as headaches. It is a further advantage that the risk of suffering a cardiovascular event is not known to be increased by treatment with nitrite.
Nitrite as used in the present invention serves to reduce elevated central systolic blood pressure, or elevated central pulse pressure, or both, and appears to have little or no effect on peripheral systolic blood pressure at physiologically acceptable concentrations, in one embodiment, there is provided nitrite for use as described, wherein peripheral systolic blood pressure is reduced by no more than 7mmHg, most preferably by no more than 5mmHg.
The present invention further provides nitrite for use in the dilation of arteries, especially large arteries, preferably without causing substantial dilation of resistance arterioles. Arteries having an internal diameter of 2 mm or more are preferred, although conduit-compliance small muscular arteries having an interna! diameter of 150 pm-2mm may be targeted.
The established favourable effects of GTN on lowering central aortic blood pressures have been attributed to effects on dilating muscular conduit arteries, causing reduction in wave reflection (Pauca AL, et a\,, Heart 2005 November;91 (11):1428-32). Dilatation of conduit arteries is therefore a key mechanism of lowering central aortic blood pressure, in contrast to peripheral blood pressure, which is mainly determined by peripheral resistance as described above.
The local average oxygen saturation of haemoglobin is preferably greater than 50%, more preferably greater than 90%, and most preferably between 95% and 100% inclusive. Nitrite is able to cause marked dilatation of the arteries under fully oxygenated conditions, including haemoglobin oxygen saturations of ~99%, without having any significant effect on pBP.
There is further provided nitrite for use in reducing the central augmentation index, preferably by at least 3%, more preferably at least 5%. The term "augmentation index" or "Alx" is used herein to refer to the ratio of the arterial augmentation pressure to the arterial pulse pressure. The augmentation pressure is the difference between the pu!se pressure and the maximum pressure, and is caused by wave reflection and constructive wave overlap. Alx can be expressed as a ratio or a percentage. A large Alx results from arterial wall stiffness and inelasticity. The "central augmentation index" or "cAlx" is the augmentation index as measured in the aorta. The "peripheral augmentation index" or "pAlx" is the augmentation index derived from the waveform measured ata peripheral location. When it is not possible to measure cA!x directly, it can be estimated from pAlx by one skilled in the art.
Nitrite, especially inorganic nitrite, as used in accordance with the present invention, may be used to alleviate pressure arising from elevated central pressure in a number of conditions. Such treatment may be sufficient, in itself, while a patient recovers from a condition, such as aortic dissection, or may be used in conjunction with known treatments for such conditions.
Suitable conditions are exemplified hereinbelow, and include conditions selected from: acute and chronic aortic dissection, impaired ventricular-vascular coupling, acute hypertensive heart failure, chronic hypertensive heart failure, diastolic heart failure due to hypertension, acute coronary syndromes, stroke due to occlusion of middle cerebral artery in children/adolescents, and adults with sickle cell disease.
Nitrite is also useful in the chronic management of hypertension, whereby the reduction of centra! systolic blood pressure or the dilation of arteries is sufficient to cause selective reduction of central haemodynamic indexes in order to reduce cardiovascular risk.
Nitrite can also be used in the chronic management of isolated systolic hypertension.
Nitrite may also be used during percutaneous coronary intervention (PCI), whereby the reduction of central systolic blood pressure or the dilation of arteries is of assistance in reducing the risk of peri-procedural infarction and/or to allow better implantation of one or more stents.
Nitrite may be used for the dilation of large arteries supplying, or in, organs or organ systems to improve blood flow and function of that organ. Nitrite may be administered at such a dose as to produce a therapeutic decrease in centra! haemodynamic indices, which may include centra! systolic blood pressure, central pulse pressure or the central augmentation index, preferably in physiologically acceptable amounts.
Typical routes of administration of nitrite include intravenous, intracoronary, oral, buccal, and/or inhaled.
Nitrite may be provided as a pharmaceutically acceptable salt. More preferably, the cation is inorganic and selected from aluminium, caicium, potassium, sodium, magnesium, and zinc ions. The alkali metal salts are generally preferred. The nitrite is preferably soluble in water to at least a level of 10 μΜ. Most preferably, nitrite for use according to the present invention is sodium nitrite.
The nitrite may be provided as a medicament comprising one or more pharmaceutically acceptably excipients. Suitable administration forms may include: liquids, pastes, tablets, capsules, gums, lozenges, sprays, solutions, powders, suspensions, suppositories, pessaries, ointments, nebu!iser preparations, delayed release formulations, transdermal patches, and injectables.
The administration form may comprise one or more pharmaceutically acceptably excipients, including, as desired, one or more of, bulking agents, diluents, solvents, pH adjusting agents, buffers, antibiotics, sterilising agents, colourants, flavourings, and stabilising agents.
The term "arteries", as used herein, is used to describe blood vessels that carry blood away from the heart, and are larger, and have a higher proportion of elastic fibres in their walls, than arterioles. The oxygen saturation of haemoglobin is typically 97-100% in arteries. "Arterioles" or "resistance arterioles" are the vessels that carry b!ood from the arteries to the capillaries and are part of the microcirculation, typically having a higher proportion of smooth muscle in their walls than arteries. The oxygen saturation levels in haemoglobin in arterioles vary with exertion level and injury but, within the systemic circulation, are less than those of the arteries.
"Systolic blood pressure", "SBP" or "systolic pressure" is the maximum blood pressure measurable during the cardiac cycle. "Central systolic blood pressure" or "cSBP" refers to the maximum aortic pressure measurable during the cardiac cycle. "Peripheral systolic blood pressure" is the maximum blood pressure measurable by conventional blood pressure instrumentation in the arm.
"Diastolic blood pressure" (DBP) is the lowest blood pressure measured during the cardiac cycle. "Central diastolic blood pressure" or "cDBP" refers to the lowest aortic pressure measurable during the cardiac cycle. "Peripheral diastolic blood pressure" or "pDBP" is the lowest blood pressure measurable by conventional blood pressure instrumentation in the arm.
"Pulse pressure" or "PP" is the difference between systolic and diastolic pressure
measurements. "Centra! pulse pressure" or "cPP" is pulse pressure as measured in the aorta.
"Mean arterial pressure" or "MAP" is the average arterial blood pressure in an individual. It is conventionally estimated by summing diastolic pressure and a third of pulse pressure, which is standard practice in the art.
The term "central haemodynamic index", as used herein, refers to a measure of the circulatory system, and may include reference to any one the following: central systolic blood pressure, central pulse pressure, and central augmentation index.
"Hypertension" is a condition whereby an individual's blood pressure is persistently elevated above 139/89mmHg.
Ideal "ventricular/vascular coupling" is manifested as low pressure fluctuation in the ascending aorta. Low pressure fluctuation results in pressure during systole being only slightly greater than pressure throughout the whole cardiac cycle, and pressure during diastole being only slightly less. This is desirable because pressure during systole determines ventricular output, in whichinotropic state and ventricular filling are constant, and ventricular metabolic requirement, while pressure during diastole in the ascending aorta is a major determinant of coronary blood flow. Factors responsible for "ideal" coupling have been identified as high distensibiiity of proximal arteries, with corresponding decreasing distensibiiity in peripheral arteries, wave reflection at arterial terminations, and a "match" between heart rate on the one hand and arterial length and wave velocity on the other. "Coronary heart disease" or "CHD" refers to a condition characterised by narrowing of the coronary arteries.
"Aortic dissection" refers to a condition characterised by bleeding into and along the aorta wall.
"Marfan syndrome" is a genetic disorder of the connective tissue. The main characterising feature in Marfan syndrome is aortic root dilation, which is aneurysm or vessel dilation at the base of the aorta, just above the aortic valve. This is associated with aortic regurgitation, dissection, and rupture, the major causes of morbidity and premature death. The exact mechanisms leading to dilatation are not fully understood, but steady and pulsatile stresses probably play an important role.
"Percutaneous coronary intervention" or "PCI" is a therapeutic procedure which is used to treat stenotic (narrowed) arteries of the heart. PCI includes several related procedures including balloon angioplasty, atherectomy (whereby atherosclerotic plaques are excised from the artery wall) the implantation of stents and brachytherapy (whereby an internally placed radioactive source is used to prevent restenosis). PCI is equivalent to coronary angioplasty or angioplasty.
"Heart failure" is a condition where the cardiac output of the heart is insufficient to meet the needs of the body. "Hypertensive heart failure" is heart failure for which hypertension is a major causative factor. "Diastolic heart failure" refers to decline in the ability of one or both of the ventricles to fill with blood during diastole.
"Acute coronary syndrome" or "ACS" refers to diseases involving unstable coronary arteries, which have the common aetiology of the formation of a thrombus on an inflamed and complicated atheromatous plaque. Examples of ACS include myocardial infarction and angina.
"Myocardial infarction" or "heart attack" refers to an event that leads to the interruption of the blood supply to the heart, resulting in myocyte death.
"ST elevation myocardial infarction" or "STEM!" refers to myocardial infarctions whereby the ST segment on the echocardiogram is elevated and non-ST elevation myocardial infarction or "non- STEMi" refers to myocardial infarction in the absence of an elevated ST segment. "Angina" refers to severe chest pain due to ischaemia of the heart muscle.
"Dilator" refers to an agent or stimulus that can induce dilation, or dilatation, both of which terms and their related terms, are used interchangeably herein. "Vasodilator" is an agent or stimulus that is able to induce the dilation of a target blood vessel, the term "vasodiiatory" to be construed accordingly.
The invention will be further described in relation to the accompanying drawings, in which:
Figure 1 shows the selectivity of increasing doses of the alpha-adrenergic antagonist phentolamine, the nitrovasodilators nitroglycerin (GTN) and sodium nitroprusside (SNP), and hydralazine on the diameter of the radial artery, a muscular conduit artery. Change in diameter of the radial artery is represented on the y axis;
Figure 2A shows the increase in large artery (radial) diameter (%) during 60 min intrabrachial infusion of sodium nitrite (8.7 μηηοΙ/ min). Data shown as mean±SEM, n=8, ***P<0.001 compared to baseline;
Figure 2B shows theincrease in small artery/forearm blood flow (FBF) during 60 min intrabrachial infusion of sodium nitrite (8.7 pmol/ min). Data shown as mean±SEM, n=8, **P<0.01 , ***P<0.001, compared to baseline;
Figure 2C shows peripheral Brachial Blood Pressure measurements during 60 min intrabrachial infusion of sodium nitrite (systolic, SBP, mean arterial, MAP, or diastolic, DBP);
Figure 2D shows systemic plasma nitrite concentrations during a 60 min intrabrachial infusion of sodium nitrite (8.7 μηηοΙ/ min; blood sampled from veins in contralateral arm). Assay performed using whole plasma. Data shown as mean±SEM, n=8, **P<0.01 compared to baseline;
Figure 3A shows the increase in large artery (radial) diameter (%) during 60 min intravenous infusion of sodium nitrite (8.7 pmol/ min) in the contralateral arm. Data shown as mean±SE , n=8, *P<0.05, **P<0.01 compared to baseline; Figure 38 shows peripheral Brachial Blood Pressure measurements during 60 min intravenous infusion of sodium nitrite (systolic, SBP, mean arterial, MAP, or diastolic, DBP);
Figure 3C shows central Systolic Blood Pressure (cSBP) before and after 60 min infusion of sodium nitrite (8.7 μιηοΙ/min for 60 min), (*P<0.05);
Figure 3D shows the peripheral augmentation index before and after 60 min infusion of sodium nitrite (8.7 μηιοΙ/ηηϊη for 60 min) (*P<0.05);
Figure 3E shows the systemic plasma nitrite concentrations during 60 min intravenous infusion of sodium nitrite (8.7 mol/min; blood sampled from veins in contralateral arm). Assay performed using deproteinated plasma. Data shown as mean±SEM, n=8, **P<0.01 compared to baseline;
Figure 4A shows the increase in large artery (radial) diameter (%) during an intrabrachiai infusion of sodium nitrite (dose response 0.087-87 μηιοΙ/ min). Data shown as mean±SEM, n=6 up to 2.6 μηηοΙ/ min, n=3 from 8.7 pmol/ min, **P<0.01 , ***P<0.001 compared to baseline;
Figure 4B shows peripheral Brachial Blood Pressure measurements during intra-arterial infusion of higher dose range sodium nitrite (8.7-87 μηιοΙ/ min), (systolic, SBP, mean arterial, MAP, or diastolic, DBP);
Figure 4C shows forearm venous capacitance during intrabrachiai infusion of sodium nitrite (low dose response 0.087-2.6 mol/ min). Data shown as mean±SEM, n=3, **P<0.01 on 2-way ANOVA comparing nitrite curve with saiine control, *P<0.05 nitrite (2.6 pmol/ min) compared to saline control;
Figure 5A shows the dose response for plasma nitrite concentration in infused arm during nitrite administration;
Figure 5B shows the dose response for plasma nitrite concentration in infused arm during nitrite administration; Figure 6A shows the increase in large artery (radial) diameter (%) during an intrabrachial infusion of GTN (dose response 0.003-1 g min). Data shown as mean±SE , n=5, **P<0.01, compared to baseline;
Figure 6B shows the efect of GTN (1 pg/min) on central SBP (cSBP), n=5;
Figure 6C is a comparison of radial artery dilatation during intra-arteria! infusion of GTN (1 pg/min) and nitrite (8.7 pmo!/min);
Figure 6D shows forearm venous capacitance during intrabrachial infusion of GTN (dose response 0.003-1 g min). Data shown as meantSEM, n=3;
Figure 7 shows individuals' responses in terms of change in forearm blood flow (FBF) and radial artery diameter during the dose response studies, expressed independently of dose, with (A) nitrite and (B) GTN. Dashed lines represent 95% confidence intervals; and
Figure 8 shows the change in radial artery diameter versus change in FBF at each dose step of (A) nitrite and (B) GTN. Dashed lines represent 95% confidence intervals.
Nitrite is capable of causing marked dilatation of arteries under fully oxygenated conditions (haemoglobin oxygen saturations of ~99%) (Figure2A and 3A). This is particularly surprising, and shows that the accepted model of action of nitrite, involving reduction by haemoglobin at its P50( is either not correct, or that there is a further mode of action. However, the discovery on which the present invention is based means that nitrite can now be used in therapy, to reduce cSBP, and there is no need to rely on, or to induce, reduced oxygen tension or ischaemic conditions. Indeed, our recent studies also indicate that nitrite is most effective at dilating conduit arteries under normal oxygen tension, with significant inhibition occurring under hypoxic and hyperoxic conditions, so that this action of nitrite appears to be optimised under normal physiological conditions. This effect in conduit arteries in hypoxia is opposite to that seen in resistance arterioles (forearm blood flow) where dilatation to nitrite is enhanced.
This dilatory action of nitrite is highly selective for arteries, having little or no effect on the resistance arterioles. This is similar to GTN, before the induction of tolerance. In the case of nitrite, tolerance is not induced at physiological acceptable levels. While GTN is a selective large artery dilator, its drawbacks, as described above, and which include headaches and the rapid induction of tolerance, mean that it is only prescribed when the need is acute. Nitrite, on the other hand, does not induce headaches or tolerance, and can be used in chronic situations, and has greater large artery selectivity overall (Figure 8A and B), compared to GTN.
It has also been found that the large artery and small vessel responses to nitrite are more predictable than the responses to GTN. In the accompanying Example 4, it is shown that nitrite results in far more predictable responses than GTN and that, at physiologically effective doses, nitrite was at least as selective as GTN, with nitrite having greater large artery selectivity overall. There was much less variability in responses for nitrite compared to GTN, indicating more predictable responses with nitrite.
Advantageously, the dilatory effect of nitrite is not limited to the local site of administration, as nitrite is also effective systemically. in the accompanying Examples, we demonstrate dilation of the radial artery when nitrite was administered intra-arterial!y (Figure2A), and dilatation of the contralateral radial artery by 9% when nitrite was administered intravenously, at a surprisingly low dose rate of 8.7 mol/min for 60 minutes (Figure3A).
Our data shows that infusion of nitrite at 8.7 pmo!/min increased plasma nitrite from 0.064±19 μΜ at baseline to 2.85±0.314 μΜ at 60 min, and resulted in a large drop in central SBP of ~12 mmHg, with no effect on peripheral BP. The larger dose of nitrite of 26 pmol/min resulted in a systemic nitrite concentration of ~13 μΜ, which was tolerated for the duration of the infusion of 20 min. Maintaining this concentration for chronic treatment would be unlikely to be tolerated in most subjects. The dose of nitrite of 87 μηιοΙ/ηιϊη resulted in a systemic concentration of -42 μΜ, but some subjects had a reduction in MAP >15 mmHg and the procedure had to be stopped. Thus, it is preferred to limit plasma concentrations of nitrite to <5 μΜ, preferably <3 μΜ, for chronic treatment or therapy, but plasma concentrations of up to 50 μΜ, preferably no more than 40 μΜ, may be employed for acute situations, such as where it is desired to have an immediate and substantial effect, although it is preferred to allow plasma concentrations to drop to those preferred for chronic administration in no more than an hour, and preferably inside 30 minutes, preferably inside 20 minutes. This is generally not a particular problem, as the half-life of nitrite is about 45 minutes. The oral bioavailability of sodium nitrite is generally in excess of 95%. Accordingly, nitrite may be administered orally in amounts largely equivalent to the systemic quantity required. A study by Hunault C. et a/., showed that the bioavailability of nitrite was 98% after oral administration of 0.12mmol NaN02/mmol Hb (mean dose of 310 mg), and 95% after oral administration of 0.06mmol NaN02/mmol Hb (mean dose of 160 mg). The Tm3X (time to maximum concentration) was 15 min. !n the accompanying Examples, sodium nitrite (8.7 μΐΓΐοΙ/ητπη) was infused over 60 min = 522 μιηοί - 36 mg, and resulting in a plasma concentration of ~3 μΜ. Similarly, sodium nitrite (26 μιηοΙ/min) was infused over 20 min = 520 μηηο! = 36 mg. This was followed by the maximum dose of sodium nitrite (87 μιηοΙ/ηΊΪη) over 20 min = 1.74 mmol = 120 mg, and resulting in a plasma concentration of ~42 μΜ, but this dose reduced peripheral pressure.
Lower doses may be associated with lower bioavailability. As a guide, for an intravenous dose of 36 mg, an oral dose of 40 mg is indicated, although it will be appreciated that uptake is dependent on many parameters including age, time of day, health, and whether the subject is eating or has recently eaten. For an intravenous dose of 72 mg, an oral dose of about 80 mg is indicated.
For intravenous/intra-arterial administration, a preferred range of plasma nitrite concentrations is -0.3-3 μΜ. These levels are preferably attained with doses of nitrite between 0.87 - 8.7 pmol/min, assuming a basal plasma nitrite concentration of -0.05 μ . A more preferred range is 0.5-1.5 μΜ. These ranges are considerably lower than the nitrite concentration of 10 μΜ which would have non-selective effects and lower MAP.
Benefit is seen with doses as low as 0.087 pmol/min, achieving a concentration of -0.1 μΜ, for example. Conversely, with a short infusion of 20 min, for example, higher dose rates may be administered with no changes in MAP, resulting in temporarily high systemic plasma concentrations of up to -100 μΜ, although this is unlikely to be useful for prolonged use.
For oral dosing, given that bioavailability of nitrite is -95-98%, as discussed above, oral doses will be similar to intravenous doses. A preferred range of nitrite is administered at between 0.087-8.7 μηηοΙ/min. Administration for 60 min at this rate equates to a dose range of 0.36-36 mg. The administration may be as single doses of fast release nitrite, or in a slow release formulation, with nitrite being released at a rate of 0.36-36 mg/h, for example. To correct for bioavailability, this may be amended to 0.4-4 mg/h: We have found that ingestion of a high dose of potassium nitrate (24 mmol) increased plasma nitrite from 0.05±0.01 to 1.0±0.13 pmol/L at 3 hours, but that this had no effect on cSBP or Alx (n=8). In contrast, systemic nitrite infusion (8.7 pmo!/min) increased plasma nitrite from 0.06±0.02 to 2.9±0.31 pmo!/L at 60 min, i.e. ~3-fold greater. In addition, nitrite at 2.6 pmol/min for 20 min also reduced cPP by ~6.5 mmHg, and was associated with a similar increase in nitrite concentration to ~1 pmoi/L (Figure 5B). Accordingly, it appears that administration of nitrate is not a preferred method of increasing plasma nitrite concentration for the purposes of the present invention.
It is particularly advantageous that nitrite has been found to be effective in lowering cSBP (Figure 3C) at a dose not associated with any change in peripheral BP.
Significantly, at a dose of 8.7 pmol/ min, for example, nitrite is capable of selectively lowering cSBP, in this case by approximately 12 mmHg, but with no effect on peripheral BP. In contrast, when GTN is administered at a dose producing a similar local dilatation of the radial artery, there is no effect on cSBP. Thus, at a systemic level, nitrite has greater large artery selectivity than GTN, and this represents a substantial clinical advantage of nitrite over GTN. Whilst there may also be some effect on the myocardium, reducing cSBP while leaving DBP substantially unchanged reduces cPP and the pressure load on the left ventricle, thereby enhancing ventricular/vascular coupling. This is a particular advantage of nitrite over GTN.
Nitrite is further capable of reducing peripheral and central A!x. For example, at a dose of 8.7 pmol/min, nitrite significantly reduced the peripheral augmentation index (pAlx) by 11.9±4.6%, while GTN had no effect (Figure3D). While the central augmentation index (cAlx) was not measured directly, it has previously been established that a change in pAix of ~12% equates to a change in cAlx of ~10% (Vadera, .,ef a/., Ann.Emerg. Med., 2011 , January; 57(1):64-5).
Advantageously, levels of nitrite that are effective in the present invention have not been associated with headaches, which has been one significant factor mitigating against the use of GTN, and represents an advantage of the present invention.
Dejamei a/., (Circulation 2007;116(16): 1821-31) demonstrated that nitrite was not subject to tolerance formation in nonhuman primates. Nitrite-induced hypotension was similar during the 14 days of continuous nitrite infusion, indicating the absence of tolerance induction typical for other exogenous NO donors such as inorganic nitrates. inhibition of ALDH2 by GTN is an important mechanism both for the bioactivation of GTN and the development of tolerance, but nitrite has been associated with the opposite effect, having been shown to increase ALDH-2 activity in the rat heart. Without being bound by theory, it is believed that ALDH-2 may also be involved in the bioactivation of nitrite, and increasing the activity of this receptor may partly explain the lack of tolerance induction with nitrite. In addition, it has been previously disclosed that nitrite is associated with a reduction in reactive oxygen species in a model of renal ischaemia-reperfusion, a further possible factor in organic nitrate tolerance.
Thus, nitrite is useful to replace GTN in vascular therapy where arterial dilation is desirable or required, particularly where the selective dilation of arteries or a reduction in cSBP without lowering cDBP is desirable.
While nitrates are not preferred for chronic indications, the present invention provides combinations of nitrite with an organic nitrate, with suitable formulations including GTN, isosorbidemononitrate, and/or isosorbidedintrate, such as a GTN patch, oral isosorbide mononitrate, or oral isosorbidedinitrate.
It will be appreciated that, while the main effect of treatment with nitrite is to reduce cSBP, proportionally lower effects may also be observed, such as a lowering of pBP or cDBP or increase in FBF, but these tend to be negligible to the point of statistical non-existence at physiologically acceptable levels of nitrite.
The present invention is particularly useful in the medical management of acute and chronic aortic dissection, where selective reduction in central aortic SBP/cPP/cAlx, is beneficial, !t is known in the art that the central indices cSBP, cPP and cAix are positively correlated with the occurrence and progression of aortic aneurysm or aortic dissection, particularly in patients with Marfan or Marfan-like syndromes. Because of the selectivity for arteries and, thereby, cSBP/cPP/cAix, a general reduction in blood pressure may substantially be avoided by the use of nitrite in accordance with the present invention. Substantia! maintenance of DBP and MAP helps to support perfusion of vital organs. For example, patients with aortic dissection benefit from strict blood pressure control over several days, and the administration of nitrite according the present invention is useful to help to reduce the risk of further dissection.
Recent studies have also suggested that central haemodynamic indices, including cSBP, cPP, and cAlx, are independently predictors of future cardiovascular events and all-cause mortality, with cAlx being capable of being used to predict clinical events independently of peripheral pressures. The present invention, accordingly, further provides the use of nitrite in the modification of one or more of these indices to reduce the risk of adverse clinical events and/or in the prophylaxis or treatment of conditions associated with or aggravated by hypertension, such as aneurysms. in accordance with the present invention, the use of nitrite in accordance with the present invention is also useful when treatingvarious conditions that can be exacerbated by high central blood pressure, such heart failure and/or impaired ventricular-vascular coupling. Suitable conditions particularly include acute hypertensive heart failure and chronic hypertensive heart failure, including diastolic heart failure due to hypertension. Ventricular/vascular coupling becomes less than ideal in adult humans and is attributable to progressive arterial degeneration, which is known to commence in childhood and is apparent in the elderly as dilated tortuous arteries, high pulse pressure, and high likelihood of developing ventricular failure. The present invention further provides nitrite for use during percutaneous coronary intervention (PCI), a therapeutic procedure known more commonly as coronary angioplasty or angioplasty, which is used to treat stenotic, or narrowed, arteries of the heart. PCI includes several related procedures including balloon angioplasty, atherectomy, wherein atherosclerotic plaques are excised from the artery wall, the implantation of stents, and brachytherapy, wherein an internally placed radioactive source is used to prevent restenosis. Nitrite may be used in accordance with the present invention for the reduction of peri-procedural infarction in PCI. In particular, this allows better implantation of balloons, stents and radiotherapy sources.
The use of nitrite in accordance with the present invention is also useful during the prophylaxis and/or treatment of various cardiovascular conditions, examples of which include, but are not limited to: the chronic management of hypertension, particularly the selective reduction of central aortic SBP/cPP/cAix to reduce cardiovascular risk; chronic management of isolated systolic hypertension, which has been associated with an increased risk of acute coronary syndromes, and for which it is desirable to selectively reduce systolic pressure in preference to diastolic BP; acute coronary syndromes, including both ST elevated myocardial infarction (STEM!) and non-STEMI; and stroke due to occlusion of middle cerebral artery in children or adolescents and aduits with sickle cell disease, to assist in dilating the middle cerebral artery and restore blood flow.
The present invention may also be used to induce dilatation of large arteries associated with a selected organ to improve blood flow and function of that organ. For example, nitrite may be administered to improve bloodflow to the kidneys or liver.
It will be appreciated that treatments of the conditions described herein are by reducing elevated central systolic blood pressure and/or elevated central pulse pressure, and will generally serve to reduce the stress on the condition in question that would otherwise be present through high central blood pressure. As such, it will generally also be entirely appropriate to administer any other medicaments normally administered for the condition concerned, unless the medicament in question is contra-indicated, or is likely to have an adverse reaction with nitrite.
As noted above, nitrite, surprisingly, has been found to possess potent effects under local fully oxygenated conditions. As such, - it is not necessary for medicaments comprising nitrite according to the invention to contain reducing agents, although these may be provided if desired. For oral administration, such agents include hawthorn berry extract and vitamin C, for example.
That nitrite has an effect on large arteries is surprising, as the art ascribes the action of nitrite to the reduction of nitrites to NO via deoxyhaemoglobin. As such, it has been assumed that the action of nitrite is inhibited by fully oxygenated haemoglobin, both because of a lack of deoxyhaem sites and also because of the oxidation of nitrite to nitrate by oxyhaemoglobin (Cosby et al. Nat Med, 2003; 9(12): 1498-505, Maher ef a/. Circulation, 2008;117:670-677).
Without being bound by theory, it is apparent that the accepted model of nitrite action cannot account for the effect on fully oxygenated large arteries, and that, while other known effects of nitrite may be accounted for by the mechanisms accepted in the art, the effect on large arteries is independent of the effects on the peripheral vasculature, and is apparent at generally lower concentrations. It is noteworthy that in vitro studies using excised rat aortic rings, in the presence of nitrite and red blood cells or free haemoglobin, showed that vasodilation was enhanced with falling oxygenation with noticeable effects when saturation fell to 80%, which levels will not be encountered in an artery under normal circumstances, and also when the concentration of nitrite was at physiologically impractical and unsustainable levels of up to 000 μ .
The accompanying Examples demonstrate that intrabrachiai infusion of nitrite immediately led to the vasodilation of the radial artery (Figure 2A), thereby excluding the possibility of conversion to NO within the microvasculature. In addition, the short half life of NO, measurable in seconds, precludes the possibility that NO generated elsewhere was responsible for the arterial dilation observed. Furthermore the numerically greater and instantaneous effect on the ipsiiateral radial artery with intra-arterial infusion of nitrite (Figure 2A) compared with the effect on the contralateral radial arterial with intravenous infusion (Figure 3A), indicates the effect is not due to gradual NO release from iron-nitrosylated haemoglobin.
Nitrite-induced dilation of the arteries under fully oxygenated conditions may possibly occur through a direct effect on the arterial wall, and seems unlikely to be dependent on an initial bioconversion to other NO species, such as nitrosothiols, as there appears to be no opportunity for the nitrite to be exposed to the necessary reduced oxygen conditions within the microcirculation or venous circulation in the timescale available.
The present invention will now be further illustrated by the following, non-limiting Examples.
EXAMPLES
Subjects
The subjects were healthy male volunteers. Systemic haemodynamic measurements
Following a minimum period of 5 min lying supine, baseline readings of non-invasive measurements of the following systemic haemodynamics were performed: carotid-femora! and brachiai-femorai pulse wave velocity (PWV) (Vicorder, Skidmore Medical ltd, Bristol UK), peripheral blood pressure and heart rate (IntelfiVue IV1P30, Phillips, NL), central blood pressure, augmentation index and heart rate (Finometer®Finapres Medical systems, Amsterdam NL); an average of three readings was taken for the final results. Measurements were repeated during and/or at the end of the study. In addition, Sp02, SpMet (Radicai-7, Rainbow, CA, US) were measured continuously.
Measurement of local Forearm Blood Flow (small vessels)
This was performed as described previously by Cheriyan J et al. Circulation 2011 February 8;123(5):515-23.
Patients remained supine and the arms were slightly elevated and supported using pre-shaped light foam blocs. Blood pressure cuffs were placed around the upper arms and the wrists. The cuffs themselves were connected to an E20 Rapid Cuff Inflators which in turn were supplied by an AG101 Cuff Inflator Air Source (Hokanson Inc. WA US). The circumference of the forearms was measured using a mercury-in-silastic strain gauge placed around each forearm connected to an EC6 Pleihysmograph (Hokanson Inc. WA US). The brachial artery was cannulaied using a 27 gauge needle (Cooper's Needle Works, Birmingham, UK). This part of the procedure was performed with the use of local anaesthesia and under strict aseptic techniques. On successful cannu!ation the needle was secured in position and a saline infusion was commenced at a rate of 1 ml/min (InjectomatAgilia syringe driver, Fresenius Kabi, Homburg, GR). To measure forearm blood flow (FBF) the wrist cuffs were inflated to supra-systolic pressure (180±2 mmHg) to exclude the circulation in the hand. After a 30-60 s pause to allow the changes in the forearm circulation to reach equilibrium, the cuffs around the upper arm were inflated to 40±2 mmHg) and deflated a number of times in cycles consisting of ~ 0 s inflation and ~5 s deflation. The degree of change in the circumferential size of the forearm measured by the mercury-in- silasticstrain gauge was acquired by channelling the signal through the EC-6 Pleihysmograph, and recorded and analysed using Chart 5 software to give a value of FBF in mi of blood / 100ml tissue / s. In addition to measuring forearm blood flow, the venous occlusion plethysmography technique was adapted to be able to measure venous capacitance.
Measurement of local Radial Artery Diameter (large/conduit vessel)
The radial artery was imaged using an Acuson-Aspen advanced ultrasound (US) probe (Siemens GR) which was fixed in position using a magnetic flexible stand (Mitutoyo JP). The image was acquired for 120 s at a rate of 1 frame every 3 s giving a total of 40 frames per sequence of acquisition. The radiai artery diameter US was analysed using dedicated vascular analysis software (Brachial analyzer, Medical Imaging Applications) which gave a measurement of the diameter of the artery for each of the 40 frames, and an average of the measurements obtained was used to calculate the vessel diameter at that time point.
Study Protocols
The participants were asked to avoid ingesting nitrate-rich foods (green leafy vegetables, beetroot and processed meats) and caffeine for 24 hours prior to the study and to fast overnight (though were encouraged to take clear fluids to avoid the risk of dehydration). They were also advised to avoid smoking and heavy exercise in that time period. The studies were conducted at a similar time of the day, around 11 :00 am under controlled temperatures 26.6 ± 0.9°C and with minimal sensory stimulation to minimise confounding. Following an initial equilibration period of 15 min and recording of baseline measurements with 0.9% saline, the effects of intra- arterial/intravenous sodium nitrite (Martindafe Pharmaceuticals, UK and Ipswich Hospital Pharmacy Manufacturing Unit, UK)/ GTN (Hospira Ltd, UK) on radial artery diameter, FBF and systemic haemodynamic parameters were assessed in the following 4 studies:
© Study 1: Fixed dose intra-arterial sodium nitrite (8.7 pmol/ min): effects on ipsilateral radial artery
® Study 2: Fixed dose intravenous sodium nitrite (8.7 pmol/ min): effects on contralateral radial artery
β Study 3: Dose response to intra-arterial sodium nitrite (0.087-87 pmol/ min): ipsilateral radial artery
• Study 4: Dose response to intra-arterial GTN (0.003-1 pg/min): ipsilateral radial artery Measurement of plasma nitrite
Blood was sampled serially through an 18 gauge Venflon® cannula placed in a forearm vein, in the relevant studies, blood was taken from the arm receiving the infusion (ipsilateral arm) in addition to the contralateral arm. Five ml samples of blood were aspirated and immediately transferred to pre-chilled Lithium Heparin tubes (Vacuette) and immediately spun at 4 °C for 5 min at 4700 RPM (Mikro 220R centrifuge, Hettich GR). The haemolysis-free supernatant plasma was removed and placed into two 1.5 ml pre-labelled Epindorff tubes. These samples were snap-frozen in liquid nitrogen and stored at -80°C for future analysis. On the day of the analysis, the samples were thawed and stored on ice. The plasma levels of nitrite were analysed using the 280i Nitric Oxide Analyzer (NOA) (Sievers Instruments, GE analytic instruments). The Purge vessel was filled with sodium iodide dissolved in 99.8% glacial acetic acid to which a few drops of antifoaming agent were added. To generate a calibration curve, a stock solution of 100 mM sodium nitrite was used to prepare the standards which were used in generating the calibration curves. The series of standard dilutions was constructed by serial dilution of the stock solution giving concentrations of 100 nM, 0.5 μΜ, 1 μΜ, 5 μΜ, 10 μ , 50 μΜ, 100 μΜ. HPLC nitrite-free water was used in the preparation of the stock and standard solutions. Using an analytical syringe, 50 μΙ_ samples of selected dilutions were injected, in duplicate, into the purge vessel directly onto the reducing agent and the NO generated was detected by the NOA and analysed by the software to generate the calibration curve. To maintain consistency, close attention was paid to the volume of reducing agent, the maintenance of the cell pressure and the maintenance of the gas pressure. Once a satisfactory calibration curve had been generated, 50 μΐ_ of the plasma samples were injected, in duplicate, into the purge vessel. The reducing agent in the purge vessel was changed after each duplicate. The concentrations of nitrite were derived from the calibration curve.
Data and Statistical Analysis
The data was analysed using the Graph Pad Prism Software. All data are expressed as mean±SE unless otherwise stated. Data were compared by repeated- measures ANOVA (2- tailed) with Dunnett's post test for comparison with baseline and Bonferroni post test for comparison with the control group. In all cases, P<0.05 was considered statistically significant.
Results
Subject characteristics
Subjects
A total of 17 healthy volunteers took part, some participating in more than one part of the study. They were all male with the following characteristics age of 29.9 ± 8.2, weight 76.5Kg ± 1.5, height 181.5cm + 3.9 giving a B I of 23.2±3.1.
EXAMPLE 1
Fixed dose intra-arterial sodium nitrite {8.7 umol/ min) over 60 min
Intra-arterial infusion of sodium nitrite (8.7 μηιοΐ/ min) into the brachial artery resulted in a rapid, marked (-30%) and highly significant dilatation of the radial artery within 5 min (P<0.001 compared to baseline saline, Figure 2A). This degree of dilatation was almost maximal by 5 min, as no further significant dilatation compared to 5 min was demonstrated. The average oxygen saturation of haemoglobin was 99% (range 97-100%), i.e. this dilatation did not occur under conditions of reduced oxygen tension found in small arterioles, or hypoxia (or ischaemia) which have been considered essential for the effects of nitrite via its reduction to NO, but rather occurred under fully oxygenated conditions that have previously been considered to inactivate nitrite/NO activity.
As expected and consistent with the art, sodium nitrite infusion resulted in a significant increase in forearm blood flow (FABF, also referred to as FBF herein) i.e. b!ood flow in the small arteries/arterioles, after 5 min (see Figure 2B). While sodium nitrite infusion into the brachial artery (8.7 μηηοΙ/ηηϊη) increased the FBF in the same (ipsifateral) forearm by 2.6 ± 1.5 ml/100 ml tissue/min (mean±SD) at 60 min (P<0.001), there was no effect in the contralateral forearm: - 0.067 ± 0.60 ml/100 ml tissue/min (data not shown).
No significant changes in peripheral blood pressure (SBP, MAP, or DBP) were detected during the 60 min infusion of sodium nitrite compared to baseline, i.e. time 0 min (Figure 2C). This was despite an eight-fold elevation in systemic plasma nitrite concentration from 64±19 nmo!/L at baseline to 528±128 nmol/L at 5 min and a 45-fold elevation to 2853±314 nmol/L at 60 min (Figure 2D).
This demonstrates that nitrite is capable of causing a significant arterial dilation of around 30%, without causing any significant drop in peripheral blood pressure. Without being bound by theory, it also appears that the effects of the present invention cannot be dependent on the conversion of nitrite to NO or another nitro-species in the micro-vascular beds.
EXAMPLE 2
Fixed dose intra-venous sodium nitrite f8.7 umol/ mini over 60 min.
Sodium nitrite was not only effective following direct intra-arteriai administration, but was a!so effective in dilating the radial artery following the intravenous infusion of the same dose of sodium nitrite into the contralateral forearm. Since the concentration in the contra-lateral artery depends on systemic intravenous administration rather than direct intra-arterial administration, the concentration of nitrite necessarily takes longer to build up, as it is determined by overall systemic concentration. Thus, the degree of dilatation was less and did not become significantiy different to baseline until 30 min, peaking at 9±3% at 45 min (P<0.01). The results are shown in Figure 3A.
As with the intra-arterial administration of sodium nitrite, intravenous sodium nitrite did not result in any significant changes in peripheral brachial blood pressure (SBP, MAP, or DBP) during the 60 min infusion of sodium nitrite compared to baseline, (time 0 min) (Figure 3B).
Despite the lack of effect on peripheral BP, sodium nitrite (8.7 pmol/min for 60 min) resulted in a large drop in centra! SBP of -12 mmHg, from 98.44±12.61 to 86.69+14.93 mmHg (P=0.02). The results are shown in Figure 3C.
The peripheral augmentation index was also decreased by 11.9±4.6% (P<0.05). The results are shown in Figure 3D.
The profile of the increase in systemic plasma nitrite concentration with intravenous nitrite was similar to the intra-arterial study (see Figure 3E). The baseline concentration appeared higher (379+65 nmol/L, compared to 64±19 nmol/L previously) owing to the use of a different method of calculation, requiring the deproteination of plasma using filters, which is typically associated with higher levels in this range. The concentrations are higher than in the i.a. study shown in Fig 2D, and may be due to contamination through the iv canula used to infuse nitrite and collect samples, suggesting that the concentrations in the i.a. study (Fig 2D) are more representative.
Thus, we show that nitrite is effective at inducing systemic arterial dilation, and that nitrite can significantiy reduce central systolic blood pressure (SPB) by at least 12 mmHg and the percent measure of the peripheral augmentation index (pA!x) by around 12%, without significantly reducing peripheral SBP.
EXAMPLE 3
Dose response to intra-arterial sodium nitrite
This study comprised a Sow dose regimen series (0.0, 0.087, 0.26, 0.87 and 2.6 μηιοΙ/ min) with subjects returning to complete the higher dose series (8.7, 26 and 87 pmol/ min): 8 subjects had completed both parts of the study. The mean base line arterial measurements were nearly identical: 2.32±±0.53cm and 2.50±0.28cm respectively (mean±SD). Figure 4A demonstrates a clear dose response to nitrite in terms of radial artery diameter, with significant dilatations compared to baseline seen with doses of 2.6 μιηοΙ/ min (resulting in a dilatation of 16.9±3.3%) and above. The highest dose of 87 μπιοί/ min resulted in a mean dilatation of 35.0+4.2%.
No changes in peripheral blood pressure were seen with doses up to 26 μηηοΙ/ min (see Figure 4B). However, at the highest dose (87 pmol/ min) BP decreased by -6/12 (9) mmHg (SBP/DBP (MAP) respectively) (see also Table 1). Heart rate (HR) did not change until the highest dose of nitrite (87 pmoS/ min) when it increased by 19 bpm (P<0.001). The highest methaemoglobin (metHb) levels reached were -4%, with a concomitant decrease in Hb02 saturations to -96%. Sodium nitrite at 2.6 μιηοΙ/ min increased systemic plasma nitrite concentration to 0.96±0.2 Μ (Figure 5B) and reduced cPP by 6.6 mmHg (95% CI, 0.3-13.0) P=0.04. cSBP decreased by -10 mmHg. Most radial artery measurements were made at 5 min following the initiation of a given dose. However, at 26 μηιοΙ/ min measurements were made at 5 min (26(E) - early) and 20 min (26(L) - late). Only minor changes were seen between these time points.
Table 1 Higher dose nitrite infusion and associated changes in radial artery (RA) diameter, forearm blood flow (FBF), BP, 02sats and metHb. 26(E) and 26(L) signify early and late changes at 5 and 20 min respectively.
Dose 0.0 8.7 26 (E) 26 (L) 87
pmol/min
RA diameter 2.52+0.28 2.95±0.32 3.16±0.28 3.21 +0.28 3.46±0.29
%Δ diameter 16.96±6.19 25.5±9.27 27.82±13 37.77±9.43
; FBF - Cont 1 95±0.62 2.00±0.84 2.28+0.77 2.36±0.64 2.77±1.52
FBF - Intv 2.03±0.96 4.59±2.68 6.46+3.18 7.15±2.41 8.30+2^5
AFBF- Cont 0.05+0.24 0.32+0.47 0.41±0.19 0.83+1.05
Δ FBF - Intv 2.55±1.72 4.42±2.23 5.12±1.45 6.27+1.58
BP mmHg
Systolic 113.5±9.5 113.5±7 111+9 109±9.5 107.5±3
Diastolic 58.5±5 67±11 59.5+4.5 61 ±5 46.5±6
MAP 71±1 78+9 72±5.5 72±5 62+5 MAP 1.67 -9 02 Sats% 98 98 98 97.67 96.33
et Hb% m 1.50 1.93 2.23 3.90
Whilst the peripheral augmentation index and cSBP were slightly lower (5.8±7.6% and -1.5 mmHg respectively) with low dose nitrite, these differences were not significant. Following high dose nitrite, cSBP was reduced by ~18 mmHg.
In addition to exploring the effects of nitrite on forearm blood flow, the effects of nitrite on forearm venous capacitance were also investigated: nitrite significantly increased forearm venous capacitance compared with saline control, and this was independently different at 2.6 pmol/ min (Figure 4C).
Figures 5Aand 5B show the dose response for plasma nitrite concentration in infused and contralateral arm during nitrite administration respectively.
This demonstrates that nitrite has a clear dose-response method of operation. No significant changes in peripheral blood pressure were seen up to a nitrite dose of to 26 pmol/min.
EXAMPLE 4
Dose response to intra -arterial GTN
The dose response to GTN used six doses (0.003-1 pg/min), which resulted in a similar range of radial artery dilatations as sodium nitrite (-10-30%). The results are shown in Figure 6A.
No changes in peripheral SBP, DBP, MAP, or HR were found following the highest GTN dose (SBP: ~+6 mmHg, DBP: ~0 mmHg, MAP: -0 mmHg, HR 0 bpm). However, in marked contrast to nitrite, GTN had no effect whatsoever on cSBP following the highest dose of GTN (Figure 6B). There was also no effect on pAlx, which was 37.0±3.4% at baseline, and 33.8+3.9 following GTN (P=0.55). This was despite the effect of this dose of GTN on large artery dilatation being at least as marked as that seen with sodium nitrite (8.7 pmol/min) which, as described above, was associated with a decrease in cSBP of -12 mmHg. The results are shown in Figure 6C. In contrast to low dose nitrite, GTN had no significant effect on venous capacitance, despite a broader dose range being investigated. Figure 6D shows forearm venous capacitance during intrabrachial infusion of GTN (dose response 0.003-1 g/ min). Data shown as mean+SEM, n=3.
The selectivity of nitrite and GTN for causing vasodilatation in large vs. small arteries was compared using the data obtained from Examples 3 and 4 respectively, and plotting all of the individuals' changes in FBF against the respective changes in radial artery diameter, independent of dose. The results are shown in Figure 7. Using this approach, changes in FBF due to nitrite were correlated with changes in radial artery diameter: ^=0.45, P<0.0001. However, there was no relationship between FBF and radial artery diameter when the changes were due to GTN r^O.00, P=0.8. Furthermore, the y-intercept for nitrite was 0.3±0.4, i.e. a 0% radial artery dilatation would result in no appreciable change in FBF. By contrast, the y- intercept for GTN was 2.3±0.6, i.e. a 0% radial artery dilatation due to GTN already results in a considerable increase in FBF. Presenting the data in this format, i.e. individual responses independent of dose, demonstrates that nitrite is more selective for large arteries than GTN and that nitrite results in a predictable response in an individual, while GTN results in an unpredictable response.
Plotting the data by dose-step revealed that nitrite resulted in a steeper gradient of 5.7±0.5, r2 - 0.78, PO.0001 compared to GTN where the gradient was 3.7±1.5, r2 = 0.18, P=0.019, showing that nitrite is more selective for the radial artery than GTN overall (i.e. a larger change for radial artery diameter for a given change in FBF) and that the responses to nitrite are more predictable than those to GTN (see Figure 8).

Claims

Claims:
1. Nitrite for use in the treatment of a patient having one of elevated central systolic b!ood pressure and elevated central pulse pressure, or both.
2. Nitrite for use according to claim 1 , wherein the nitrite is an inorganic nitrite, and preferably wherein the cation is inorganic and is selected from aluminium, calcium, potassium, sodium, magnesium, and zinc ions, particularly preferably wherein the nitrite is sodium nitrite.
3. Nitrite for use in accordance with claim 1 or 2, wherein said patient has a peripheral blood pressure of no higher than 139/89mmHg.
4. Nitrite for use in accordance with any preceding claim, wherein the treatment is selected such as to reduce peripheral systolic blood pressure by no more than 7mmHg, preferably by no more than 5mmHg, and more preferably to have substantially no effect on peripheral b!ood pressure.
5. Nitrite for use in accordance with any preceding claim, wherein the treatment takes the form of chronic administration.
6. Nitrite for use in accordance with any preceding claim, administered so as to attain plasma concentrations of up to 5 μ , and preferably levels of between 0.3 μΜ and 3 μ , inclusive.
7. Nitrite for use in accordance with any preceding claim, wherein the amount of nitrite used is selected such as to reduce the central augmentation index by at least 3%, and preferably by at least 5%.
8. Nitrite for use in accordance with any preceding claim, wherein the patient has a vascular condition selected from: acute and chronic aortic dissection, acute and chronic aortic aneurysm, impaired ventricular-vascular coupling, acute hypertensive heart failure, chronic hypertensive heart failure, diastolic heart failure due to hypertension, acute coronary syndromes, including both ST elevated myocardial infarction (STE I) and non-STEMI, stroke due to occlusion of middle cerebral artery in children/adolescents, and adults with sickle ceil disease.
9. Nitrite for use in accordance with any preceding claim, wherein the patient has hypertension or isolated systolic hypertension, and the treatment is chronic.
10. Nitrite for use in accordance with any of claims 1 to 7, during the peri-operational period for percutaneous coronary intervention (PCi).
11. Nitrite for use as defined in any of claims 1 to 7, to dilate large or conduit arteries.
12. Nitrite for use in accordance with any preceding claim, wherein the nitrite is sodium nitrite.
13. Nitrite for use in accordance with any of claims 1 to 4, wherein the treatment is acute, preferably for no longer than 20 minutes, and preferably wherein plasma nitrite does not exceed 80μΜ, and preferably does not exceed 40μ .
14. Nitrite for use as defined in any preceding claim, to reduce central puise pressure.
15. Nitrite for use as defined in any preceding claim, to reduce centra! blood pressure where glyceryl trinitrate is a treatment option, nitrite being used instead of GTN in an appropriate amount.
16. Nitrite for use in accordance with any preceding claim, wherein the central arteries relevant to the elevated central systolic blood pressure and elevated central pulse pressure, or both, are substantially normoxic.
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WO2020210707A1 (en) * 2019-04-10 2020-10-15 Mayo Foundation For Medical Education And Research Methods and materials for gender-dependent treatment of cardiovascular dysfunction

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