RU2838908C2 - Inositol phosphate compounds for use in increasing tissue perfusion - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: presented is the use of a hexanodium salt of myo-inositol hexaphosphate or a pharmaceutical composition containing said salt together with pharmaceutically acceptable excipients and carriers for treating or preventing peripheral arterial disease.

EFFECT: invention provides improved blood perfusion in extremities and mobility (ability to walk) of patients suffering peripheral artery diseases and critical extremity ischemia.

12 cl, 17 dwg, 6 ex

Description

[0001] This application claims the benefit of priority from applications EP 19382061.0, filed January 30, 2019, and US 62/913259, filed October 10, 2019.

Field of technology

[0002] The present invention relates to the use of inositol phosphates (IP), their analogs and derivatives for increasing tissue perfusion and/or oxygenation. The present invention also relates to pharmaceutical compositions containing said IP, their analogs and derivatives, and their use in animal and human health.

State of the art

[0003] Peripheral arterial disease (PAD) is a common disorder characterized by stenosis and/or obstruction of the arteries of the lower extremities, resulting in decreased muscle perfusion and oxygenation. PAD is a major public health problem and poses a high risk of long-term suffering. PAD increases the risk of tissue necrosis (gangrene), amputation, and premature death.

[0004] PAD is the result of lower limb ischemia. The underlying cause is atherosclerosis. In mild forms, PAD may be limited to intermittent claudication and pain in the lower limbs. Lower limb PAD is the leading cause of disability and loss of mobility in older men and women and has a major impact on quality of life.

[0005] The prevalence of PAD is ~12% in the adult population. This prevalence increases to more than 20% in the population over 70 years of age. Currently, PAD affects more than eight million men and women in the United States alone. It is estimated that more than 200 million people worldwide suffer from PAD. The prevalence of PAD is likely to increase in the near future as the general population ages and the incidence of obesity-related type 2 diabetes increases. Cigarette smoking is also another important risk factor. Patients with PAD have increased rates of cardiovascular (CV) morbidity and mortality, faster rates of functional decline, and increased rates of mobility loss compared to the general population.

[0006] The goals of the protocol for treating patients with PAD include reducing the incidence of cardiovascular events, improving functional performance, and preventing functional decline and loss of mobility. Restoring or improving blood flow to the extremities may help achieve these goals.

[0007] Although endovascular and lower extremity revascularization procedures significantly improve walking in patients with PAD, revascularization procedures are not therapeutic options for many of them, either because of the presence of comorbidities or because the location and type of atherosclerotic lower extremity disease are not amenable to revascularization. Revascularization is invasive, expensive, and associated with risks, especially in older patients. For these reasons, clinicians need effective, affordable, and well-tolerated treatments to improve lower extremity function in patients with PAD.

[0008] Currently, only two drugs have been approved by the Federal Drug Administration (FDA) to improve walking in people with PAD: pentoxifylline (1984) and cilostazol (1999). Since then, no new drugs have been approved for the treatment of intermittent claudication. Furthermore, in recent studies of patients with PAD, pentoxifylline did not significantly improve claudication symptoms or maximum pain-free walking distance compared with placebo. Recent clinical practice guidelines recommend against prescribing pentoxifylline for intermittent claudication symptoms due to lack of therapeutic effect.

[0009] Cilostazol is a phosphodiesterase inhibitor that provides approximately 25% to 40% improvement in treadmill walking performance in individuals with symptomatic PAD. Cilostazol is a phosphodiesterase type 3 inhibitor that acts by increasing intracellular cyclic adenosine monophosphate concentrations; in doing so, the drug inhibits platelet aggregation and serves as a direct arterial vasodilator, improving blood perfusion. However, the mechanism by which cilostazol improves walking performance in patients with PAD remains unclear.

[0010] Side effects of cilostazol include headache, diarrhea, palpitations, and dizziness. There is a black box warning against prescribing cilostazol to patients with a history of cardiovascular disease. Cilostazol should not be given to patients with PAD who also have heart failure. Cilostazol interacts with drugs regularly prescribed to patients with renal impairment or cardiovascular disease, such as cinacalcet, clopidogrel, and ibandronate, increasing the risk of adverse reactions in these patients when cilostazol is co-administered with other drugs.

[0011] In conclusion, medical treatments for symptomatic relief are limited, surgical or endovascular interventions are feasible for some people, but long-term results are often disappointing. As a result, there is a need to develop more effective and safe new treatments for PAD.

The essence of the invention

[0012] According to a first aspect, the present invention relates to a compound of general formula I or a pharmaceutically acceptable salt thereof for use in increasing tissue perfusion and/or oxygenation in a subject in need thereof.

[0013] where R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are independently selected from OH, a radical of formula II, III, IV and a heterologous moiety:

[0014] provided that:

[0015] at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ is selected from a radical of formula II, III and IV, and

[0016] zero, one, two, or three of R _1, R ₃ , R ₅ , R ₇ , R ₉ , and R ₁₁ are

heter o logical fragment.

[0017] According to another aspect, the present invention relates to a compound of general formula I as defined above for use in the treatment or prevention of ischemia in a subject in need thereof. According to a version of this aspect, the present invention relates to a compound of general formula I as described above for use in the treatment or prevention of an ischemia-related disease or condition in a subject in need thereof.

[0018] According to some aspects, the present invention relates to a compound of general formula I, as defined above, wherein the heterologous moiety is selected from a radical of formula V, a radical of formula VI and a radical of formula VH:

[0019] and where n is an integer in the range from 2 to 200, and R ₁₃ is selected from H, methyl or ethyl.

[0020] According to a further aspect, the present invention also relates to a method for increasing tissue perfusion and/or oxygenation, which comprises administering a therapeutically effective amount of a compound of formula I, as defined above, together with pharmaceutically acceptable excipients or carriers, to a subject in need thereof. This aspect can also be formulated as the use of a compound of formula I, as defined above, for the manufacture of a medicament for increasing tissue perfusion and/or oxygenation in a subject in need thereof.

[0021] According to another aspect, the present invention also relates to a method for the treatment or prevention of ischemia and/or an ischemia-related disease or condition, which comprises administering a therapeutically effective amount of a compound of formula I, as defined above, together with pharmaceutically acceptable excipients or carriers, to a subject in need thereof. This aspect can also be formulated as the use of a compound of formula I, as defined above, for the manufacture of a medicament for the treatment or prevention of ischemia and/or an ischemia-related disease or condition in a subject in need thereof.

[0022] According to a further aspect, the present invention relates to a method for the treatment or prevention of peripheral arterial disease, which comprises administering a therapeutically effective amount of a compound of formula I, as defined above, together with pharmaceutically acceptable excipients or carriers, to a subject in need thereof. This aspect can also be formulated as the use of a compound of formula I, as defined above, for the manufacture of a medicament for the treatment or prevention of peripheral arterial disease in a subject in need thereof.

[0023] The compounds of the present invention are particularly useful for increasing tissue perfusion and/or oxygenation in the lower extremities and, in particular, for the treatment or prevention of peripheral arterial disease (PAD) and closely related conditions such as critical limb ischemia (CLI). These compounds also exhibit many beneficial properties (e.g., a better safety profile) compared to cilostazol, a reference drug currently indicated for the treatment of PAD.

[0024] The present invention also relates to a pharmaceutical composition comprising at least one compound of formula I as defined above for use in (i) increasing tissue perfusion and/or oxygenation, (ii) treating or preventing ischemia and/or an ischemia-related disease, and/or (iii) treating or preventing PAD in a subject in need thereof. This aspect can also be formulated as the use of a pharmaceutical composition comprising at least one compound of formula I as defined above for the manufacture of a medicament for (i) increasing tissue perfusion and/or oxygenation, (ii) treating or preventing ischemia and/or an ischemia-related disease, and/or (iii) treating or preventing PAD in a subject in need thereof.

Brief description of the drawings

[0025] Fig. 1 shows representative examples of inositol phosphate analogs in which two of the six X's are OPSO ₂ ^2- and the remaining X's are OSO ³ . Two specific forms of 4,6-di-(O-thiophosphate)-inositol-1,2,3,5-tetra-O-sulfate are shown.

[0026] Fig. 2 shows inositol phosphate analogs and inositol phosphate derivatives that can be used to practice the methods of the present invention. The molecules shown are myo-inositol pentakisphosphate-2-PEG400, myo-inositol hexakisulfate (myo-inositol hexasulfate), and scyllo-myo-inositol hexakisulfate (scyllo-inositol hexasulfate).

[0027] Fig. 3 shows inositol phosphate analogs and inositol phosphate derivatives that can be used to practice the methods of the present invention. X independently represents phosphorus- and/or sulfur-containing groups (e.g., phosphate, sulfate, or thiophosphate). R ¹ represents a heterologous moiety (e.g., PEG or PG).

[0028] Fig. 4 shows exemplary inositol phosphate analogs and inositol phosphate derivatives that can be used to practice the methods of the present invention. R ¹ is a heterologous moiety (e.g., PEG or PG). n can be from 2 to 200.

[0029] Fig. 5 shows exemplary inositol phosphate analogs and inositol phosphate derivatives that can be used to practice the methods of the present invention. n can be from 2 to 200.

[0030] Fig. 6 shows exemplary inositol phosphate analogs and inositol phosphate derivatives that can be used to practice the methods of the present invention. n can be from 2 to 200.

[0031] Figure 7 shows the blood flow in the hind limbs of a rat model at D0, measured using laser Doppler imaging. The blood flow is shown in normalized perfusion units (PU). Normalization is achieved by comparing the raw data with the data of group 1 at D0.

[0032] Figure 8 shows blood flow in the hind limbs of a rat model at D6, measured using laser Doppler imaging. Blood flow is shown in normalized perfusion units (PU). Normalization is achieved by comparing the raw data to the data of group 1 at D6.

[0033] Figure 9 shows blood flow in the hind limbs of a rat model at D12, measured using laser Doppler imaging. Blood flow is shown in normalized perfusion units (PU). Normalization is achieved by comparing the raw data to the data of group 1 at D12.

[0034] Fig. 10 shows the percentage of inhibition of aortic calcification in a rat model using VitD at D12. Calcium levels at the time of sacrifice were measured by ICP-OES.

[0035] Figure 11 shows blood flow in the hind limbs of a rat model at D12 and D18 (6 days after treatment cessation) measured by laser Doppler imaging. Blood flow is shown in normalized perfusion units (PU). Normalization is achieved by comparing raw data to group 1 data at D12 and D18.

[0036] Fig. 12 shows (A) the maximum pain-free walking distance (MWD) and (B) the maximum pain-free walking time (MWT) in a rat model at D10, measured using a treadmill test. The maximum pain-free walking distance is displayed in meters (m), and the maximum pain-free walking time is displayed in minutes (min).

[0037] Fig. 13 shows the maximum pain-free walking distance (MWD) in the rat model at D17 (5 days after treatment cessation) measured using a treadmill test. The maximum pain-free walking distance is displayed in meters (m) up to 40 minutes of walking.

[0038] Figure 14 shows the percentage inhibition of aortic calcification in a rat model using VitD at D24 (12 days after treatment cessation). Calcium levels at the time of sacrifice were measured by ICP-OES.

[0039] Figure 15 shows the blood flow in the hind limbs of a rat model at D0, D5, and D13 (8 days after treatment initiation) measured using laser Doppler imaging. The blood flow is shown in normalized perfusion units (PU). Normalization is achieved by comparing the raw data to the data of Group 1 at D0, D5, and D13.

[0040] Fig. 16 shows (A) the maximum pain-free walking distance (MWD) and (B) the maximum pain-free walking time (MWT) in a rat model at D11 (7 days after the start of treatment) measured using a treadmill test. The maximum pain-free walking distance is shown in meters (m) and the maximum pain-free walking time is shown in minutes (min).

[0041] Fig. 17 shows the percentage inhibition of femoral artery calcification in a rat model using VitD at day 13 (9 days after the start of treatment). Calcium levels at the time of sacrifice were measured by ICP-OES.

Detailed description of the invention

[0042] The present invention relates to compounds, pharmaceutical compositions, methods and routes of administration for use in increasing tissue perfusion and/or oxygenation. The present invention also relates to compounds, pharmaceutical compositions, methods and routes of administration for use in the treatment or prevention of ischemia and ischemia-related diseases and conditions.

[0043] The compounds of the present invention are particularly useful for increasing tissue perfusion and/or oxygenation in the lower extremities and, in particular, for the treatment or prevention of peripheral arterial disease (PAD) and related conditions such as critical limb ischemia (CLI). These compounds also exhibit many beneficial properties compared to other approved drugs for the treatment of PAD and CLI.

1. Definitions of general terms and expressions

[0044] The present invention includes embodiments in which exactly one member of the group is present, used, or otherwise related to a given product or process. It also includes embodiments in which more than one or all members of the group are present, used, or otherwise related to a given product or process.

[0045] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. For example, Pei-Show J, Concise Dictionary of Biomedicine and Molecular Biology, 2nd ed. (CRC Press, Boca Raton, FL, USA 2002); Lackie J, The Dictionary of Cell and Molecular Biology, 5th ed. (Academic Press, Cambridge, MA, USA 2013) and Cammack R, et al, Oxford Dictionary of Biochemistry and Molecular Biology, 2nd ed., (Oxford University Press, Oxford, GB, 2006) provide the skilled artisan with a general dictionary of many of the terms used in this disclosure.

[0046] Units, prefixes and symbols are designated in their metric system (Systeme International de Unites, SI) form. Numerical ranges include the numbers defining the range. When a range of values is specified, it is understood that each intermediate integer value and each fraction thereof between the specified upper and lower limits of the range are also specifically disclosed, along with each subrange between such values. The upper and lower limits of any range may be independently included or excluded from the range, and each range that includes one, neither, or both limits is also encompassed by the present invention.

[0047] When a value is explicitly stated, it is to be understood that values that are approximately the same number or magnitude as the stated value are also within the scope of the present invention. When a combination is disclosed, each subcombination of the elements of that combination is also specifically disclosed and is within the scope of the present invention. Conversely, when different elements or groups of elements are separately disclosed, combinations thereof are also disclosed. Where any element of the present invention is disclosed as having multiple alternatives, examples of the present invention in which each alternative is excluded, either individually or in any combination with other alternatives, are also disclosed; more than one element of the present invention may have such exceptions, and all combinations of elements having such exceptions are disclosed.

[0048] As used herein, the term "and/or" is intended to mean a specific invention of each of the two stated features or components, with or without the other. Thus, the term "and/or" as used in a phrase such as "A and/or B" herein is intended to include "A and B," "A or B," "A" (alone) and "B" (alone). Likewise, the term "and/or" as used in a phrase such as "A, B and/or C" is intended to cover each of the following: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0049] The term "about" as used herein and applied to one or more values of interest refers to a value that is similar to a stated reference value. In some aspects, the term "about" refers to a range of values that are within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater or less) of the stated reference value, unless otherwise stated or otherwise apparent from context (except where such number is greater than 100% of the possible value).

[0050] As used herein, the terms "critical limb ischemia" or "CLI" refer to a severe obstruction of the arteries that markedly reduces blood flow to the extremities and progresses to severe pain and even skin ulcers, open skin lesions, or gangrene. Critical limb ischemia is a very severe peripheral arterial disease. In some aspects, administering an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) to a subject in need thereof improves the subject's ability to walk faster and for greater distances compared to no treatment.

[0051] As used herein, the term "compound" includes all isomers and isotopes of the depicted structure. As used herein, the term "isomer" means any geometric isomer, tautomer, zwitterion, stereoisomer, enantiomer, or diastereomer of the compound. Compounds may include one or more chiral centers and/or double bonds and, thus, may exist as stereoisomers, such as double bond isomers (i.e., geometric E/Z isomers) or diastereomers (e.g., enantiomers (i.e., (+) or (-)) or cis/trans isomers). The present invention encompasses any and all isomers of the compounds described herein, including stereomerically pure forms (e.g., geometrically pure, enantiomerically pure, or diastereomerically pure) and enantiomeric and stereoisomeric mixtures, such as racemates. Enantiomeric and stereomeric mixtures of compounds and methods for separating them into their component enantiomers or stereoisomers are well known. A compound, salt, or complex of the present invention can be prepared in combination with solvent or water molecules to form solvates and hydrates by conventional methods.

[0052] As used herein, the term "cilostazol" refers to 6-[4-(1-cyclohexyl-1H-tetrazol-5-yl)butoxy]-3,4-dihydro-2(1H)-quinolinone [CAS-73963-72-1], a quinolinone derivative that inhibits cellular phosphodiesterase. The molecular formula and mass of cilostazol are C ₂₀ H ₂₇ N ₅ O ₂ and 369.46 g/mol, respectively. Its structural formula is:

[0053] As used herein, the term "effective amount" and the related terms "effective dose" and "effective dosage" when referring to (i) a compound of general formula I (e.g., inositol phosphate, an inositol phosphate analog, an inositol phosphate derivative, or a combination thereof), or (ii) a pharmaceutical composition comprising at least one of the compounds of (i), is such an amount sufficient to achieve beneficial or desired results. In some embodiments, beneficial or desired results are, for example, clinical results, and as such, "effective amount" depends on the context in which it is used. In the context of administering a therapeutic agent that increases tissue perfusion and/or oxygenation, an effective amount of the therapeutic agent is, for example, an amount sufficient to (a) increase tissue perfusion in a defined area, (b) stop, reduce, slow the progression of, or reverse ischemia in a defined area, or (c) improve mobility or walking ability (e.g., speed, distance) in a subject compared to the same parameters observed in the subject prior to administration of the therapeutic agent, or in a population of control subjects without administration of the therapeutic agent.

[0054] As used herein, the term "ischemia" refers to a limitation of blood supply to tissues that results in a lack of oxygen needed to support cellular metabolism. Ischemia includes not only a lack of oxygen, but also a decrease in nutrient availability and inadequate removal of metabolic waste. Ischemia may be partial (poor perfusion) or complete.

[0055] As used herein, the terms "maximum pain-free walking distance" or "MWD" refer to the distance at which a subject is unable to continue walking without assistance due to exhaustion or severe pain. In the context of assessing an increase in MWD, said increase is assessed by comparing the subject's MWD values before and after treatment with a therapeutic agent or by comparing the subject's MWD values after treatment with a population of control subjects who have not received treatment with a therapeutic agent.

[0056] As used herein, the terms "maximum pain-free walking time" or "MWT" refer to the time during which a subject is unable to continue walking without assistance due to exhaustion or severe pain. In the context of assessing an increase in MWT, said increase is assessed by comparing the subject's MWT values before and after treatment with a therapeutic agent or by comparing the subject's MWT values after treatment with a population of control subjects who have not received treatment with a therapeutic agent.

[0057] As used herein, the terms "parenteral administration" and "administered parenterally" mean routes of administration other than enteral and local administration, typically by injection, and include, without limitation, the following: intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion (e.g., renal dialysis infusion).

[0058] As used herein, the terms "peripheral arterial disease" or "PAD" refer to a narrowing of the peripheral arteries to the legs (most commonly), abdomen, arms, and head. Symptoms include intermittent claudication (e.g., leg pain with walking that goes away with rest), skin ulcers, bluish skin, cold skin, or poor nail and hair growth.

[0059] As used herein, the terms "prevent," "prevention," and "prophylaxis" refer to suppressing the onset or reducing the occurrence of a disease or condition in a subject (e.g., preventing the development of ischemic tissue in the limbs).

[0060] As used herein, the term "SNF472" refers to a formulation of myo-inositol hexaphosphate hexasodium for intravenous administration. SNF472 is manufactured by dissolving myo-inositol hexaphosphate hexasodium in saline, followed by pH adjustment and aseptic filtration. SNF472 is prepared in three different dosage strengths: (a) (i) 20 mg/mL and (ii) 90 mg/mL in single-use 5 mL vials formulated in saline, pH 5.8-6.2, and (b) 30 mg/mL in single-use 10 mL vials formulated in saline, pH 5.6-6.4.

[0061] As used herein, the terms "subject," "individual," "animal," or "mammal" mean any subject, particularly a mammalian subject, for which a diagnosis, prognosis, or treatment is desired. Mammalian subjects include, but are not limited to, humans, domesticated animals, farm animals, zoo animals, sport animals, pets such as dogs, cats, guinea pigs, rabbits, rats, mice, horses, cattle, cows; primates such as great apes, monkeys, orangutans, and chimpanzees; canines such as dogs and wolves; felines such as cats, lions, and tigers; odd-toed ungulates such as horses, donkeys, and zebras; bears, food animals such as cows, pigs, and sheep; ungulates such as deer and giraffes; rodents such as mice, rats, hamsters and guinea pigs; and so on. In certain aspects, the subject is a human. In some aspects, the subject is a human patient with reduced tissue perfusion and/or oxygenation in the muscles of the lower extremities, or at risk of developing said condition. In some further aspects, the subject is a human patient with ischemia and/or an ischemia-related disease or condition, or at risk of developing said ischemia, ischemia-related disease or condition.

[0062] As used herein, the term "substantially" refers to a qualitative condition of exhibiting a full or nearly full measure or degree of a characteristic or property of interest. One skilled in the art of biology will appreciate that biological and chemical phenomena rarely, if ever, reach completion and/or proceed to completion or achieve an absolute result or avoid achieving an absolute result. Thus, the term "substantially" is used herein to denote the potential lack of completion inherent in many biological and chemical phenomena.

[0063] As used herein, the term "tissue perfusion" refers to the flow of blood or other perfusate through the vessels of a particular tissue or organ. "Increase tissue perfusion" or "increase in tissue perfusion" as used herein refers to an increase in blood flow in a particular tissue region in a subject following administration of an inositol phosphate according to the present invention, compared to the same parameters observed in the subject prior to administration of said therapeutic agent or in a population of control subjects without administration of said therapeutic agent.

[0064] As used herein, the terms "treat" or "treatment" refer to the administration of a compound or pharmaceutical composition of the present invention to (i) slow, (ii) inhibit the progression of, (iii) stop, or (iv) reverse the progression of a disease or condition after clinical signs thereof have appeared. Control of disease progression refers to beneficial or desired clinical results that include, but are not limited to, reduction of symptoms, shortening of disease duration, stabilization of pathological conditions (especially to avoid further deterioration), delay of disease progression, improvement of pathological conditions, and remission (either partial or complete). Control of disease progression also includes an increase in survival compared to the expected survival if no treatment were administered. In the context of the present invention, the terms "treat" and "treatment" refer specifically to (a) increasing tissue perfusion and/or oxygenation or (b) stopping, reducing, slowing the progression of, or reversing the development of ischemic tissue, particularly in the lower extremities, or (c) improving mobility or walking ability (e.g., speed, distance, endurance) in a subject administered the compounds or pharmaceutical compositions of the present invention.

[0065] As used herein, the term "walking ability" refers to the ability of a subject to move autonomously without assistance. The MWD and MWT parameters indicate the ability of a subject to walk.

2. Connections

[0066] Compounds for use in the present invention are inositol phosphates as defined in the first aspect of the present invention, as well as analogs and derivatives thereof. The term "inositol phosphate" as used herein refers to a compound with an inositol ring and one, two, three, four, five or six phosphate groups, or a combination thereof. Myo-inositol hexaphosphate (IP6) is an exemplary inositol phosphate according to the present invention. In some aspects, the inositol phosphate is pure (e.g., greater than 99% of the inositol phosphate species are the same species, e.g., IP6) or substantially pure (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% of the inositol phosphate species are the same species, e.g., IP6). In some aspects, the inositol phosphate is a mixture, such as containing varying amounts of IP1, IP2, IP3, IP4, IP5, and IP6. In some aspects, the inositol phosphate is a racemic mixture.

[0067] The present invention also provides inositol phosphate analogs. As used herein, "inositol phosphate analog" refers to a compound that contains a ring with a different number of carbon atoms relative to the inositol ring (i.e., 5 or 7 carbon atoms) and/or contains at least one sulfate or thiophosphate group. For example, a compound containing a ring with 5, 6, or 7 carbon atoms and at least one phosphate, sulfate, or thiophosphate group would be considered an inositol phosphate analog.

[0068] As used herein, the term "inositol phosphate derivative" refers to inositol phosphate or an inositol phosphate analog that contains a heterologous moiety (i.e., a group that is not a phosphate, sulfate, or thiophosphate). For example, inositol pentasulfate, which contains a heterologous polyethylene glycol moiety, or myo-inositol hexaphosphate, which contains a heterologous polyglycerol moiety, should be considered as inositol phosphate derivatives.

[0069] As used herein, the term "heterologous moiety" refers to a moiety in a compound of Formula I that is not a phosphate, sulfate, or thiophosphate and imparts a desired property to such a compound. For example, a heterologous moiety (e.g., polyglycerol or polyethylene glycol) can increase the solubility of the compound. In some aspects, a heterologous moiety can impart multiple desired properties (e.g., polyglycerol and polyethylene glycol can both increase the solubility of the compound and decrease the rate of elimination of the compound).

[0070] As used herein, the terms "inositol phosphate of the invention" and "inositol phosphate of the present invention" are a generic term that encompasses "inositol phosphate," "inositol phosphate analog," "inositol phosphate derivative," and combinations thereof. In some aspects, the term "inositol phosphate of the present invention" encompasses compositions comprising "inositol phosphate," "inositol phosphate analog," "inositol phosphate derivative," or a combination thereof, and at least one additional therapeutic agent. In some aspects, the additional therapeutic agent comprises cilostazol, pentoxifylline, or a combination thereof.

[0071] Compounds of the present invention that contain a ring with 5, 6 or 7 carbon atoms and at least one sulfate or thiophosphate group, but without a phosphate group, will still be considered an "inositol phosphate analog" or an "inositol phosphate analog" in the context of the present invention. Thus, the term "inositol phosphate according to the present invention" covers not only phosphate-containing compounds, but also compounds without phosphate groups that contain a ring with 5, 6 or 7 carbon atoms and at least one sulfate or thiophosphate group.

[0072] Representative inositol phosphates of the present invention are shown in Figs. 1-6. Fig. 3 shows numerous examples of inositol phosphates, all of which are characterized by the myo conformation. In addition to myo-inositol, other naturally occurring stereoisomers of inositol include scyllo-, muco-, 1D-chiro-, 1L-chiro-, neo-inositol, allo-, epi-, and cis-inositol. As their names imply, 1L- and 1D-chiro-inositol are a single pair of inositol enantiomers, but they are enantiomers of each other and not of myo-inositol. It should be understood that any illustrative inositol phosphate shown in the present disclosure is not limited to the representative conformation shown. Thus, for example, the examples shown in Figs. 3, will also cover the corresponding equivalents in the scyllo-, muco-, 1D-chiro-, 1L-chiro-, neo-inositol, allo-, epi-, and cis-inositol conformations. In its most stable conformation, the myo-inositol isomer adopts a chair conformation that moves the maximum number of hydroxyls to the equatorial position, where they are furthest apart. In this conformation, the natural myo isomer has a structure in which five of the six hydroxyls (the first, third, fourth, fifth, and sixth) are equatorial, while the second hydroxyl group is axial.

2.1. Inositol phosphates, analogues and derivatives

[0073] According to some aspects, at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ of the compound of general Formula I is independently H, -X, -OX, -NHX, -NX ₂ , -SX, -OSO ₃ HX, -OSO ₃ X ₂ or a compound of Formula II, Formula III or Formula IV, wherein each X is independently H, C _1-30 alkyl, C _2-30 alkynyl or Cy ₁ wherein the C _1-30 alkyl, C _2-30 alkenyl and C _2-30 alkynyl are independently optionally substituted with one or more R ₁₄ and wherein Cy ₁ is optionally substituted with one or more R ₁₅ ; Cy ₁ is a carbocyclic or heterocyclic 3-10 membered ring which may be saturated, partially unsaturated or aromatic, wherein said heterocycle contains one to four heteroatoms selected from O, S and N, wherein said ring may be bonded to the rest of the molecule through any available C atom, and wherein Cy ₁ is optionally fused to one to four five- or six-membered rings each of which is saturated, partially unsaturated or aromatic, carbocyclic or heterocyclic, and wherein said fused heterocycle may contain one or two heteroatoms selected from O, N and S; each R ₁₃ is independently H, C _1-30 alkyl, -NH ₂ , -NHC _1-30 alkyl or N(C _1-30 alkyl) ₂ , wherein each C _1-30 alkyl is independently optionally substituted with one or more halogen, -OH, -CN and -NO ₂ groups; and each R ₁₄ and R ₁₅ independently represents -OH, C _1-30 alkoxy, C _1-30 alkylthionyl, C _1-30 acyloxy, phosphate, halogen, trihalogen C _1-30 alkyl, nitrile, azide.

[0074] In certain non-explicit aspects, each X is independently H, C _1-30 alkyl, or Cy ₁ , wherein the C _1-30 alkyl is optionally substituted with one or more R ₁₄ and wherein Cy ₁ is optionally substituted with one or more R ₁₅ ; and each R ₁₄ and R ₁₅ is independently -OH, C _1-30 alkoxy, C _1-30 alkylthionyl, C _1-30 acyloxy, phosphate, halogen, trihalogen C _1-30 alkyl, nitrile, or azide. In some aspects, each X is H, C _1-30 alkyl, or Cy ₁ . In some aspects, each X is H.

[0075] According to some further aspects, at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ is independently a compound of formula II, formula III or formula IV, each R ₁₃ is independently H, C _1-30 alkyl, -NH ₂ , -NHC _1-30 alkyl or -N(C _1-30 alkyl) ₂ , wherein each C _1-30 alkyl is independently optionally substituted with one or more halonene, -OH, -CN and -NO ₂ groups; and R ₂ , R ₄ , R ₆ , R ₈ , R ₁₀ and R ₁₂ are independently H.

[0076] According to a further aspect, R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ independently represent a compound of formula II, formula III or formula IV, each R ₁₃ independently represents H or C _1-30 alkyl, wherein each C _1-30 alkyl is independently optionally substituted with one or more halonene, -OH, -CN and -NO ₂ groups; and R ₂ , R ₄ , R ₆ , R ₈ , R ₁₀ and R ₁₂ independently represent H.

[0077] According to a further aspect, at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ is a compound of formula II, formula III or formula IV, and each R ₁₃ is independently H or C _1-30 alkyl. According to another aspect, at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ is a compound of formula II, formula III or formula IV, and each R ₁₃ is H.

[0078] In a particular aspect, the compound is inositol hexaphosphate (IP6). In other aspects, the compound is inositol monophosphate (IP1), inositol diphosphate (IP2), inositol triphosphate (IP3), inositol tetraphosphate (IP4), or inositol pentaphosphate (IP5). In some aspects, the compound comprises a combination of IP1, IP2, IP3, IP4, IP6, and IP6. In some aspects, IP6 can form other inositol phosphates (IP5, IP4, IP3, IP2, IP1) by dephosphorylation in vivo. Inositol is intended to mean any isomeric form of the molecule, such as myo-inositol.

[0079] According to some aspects, the compounds for use in the present invention are compounds of formula I, wherein:

[0080] R ₇ is OSO ₃ ^- , and R ₉ , R ₅ , R ₃ , R ₁ and R ₁₁ are independently selected from OPO ₃ ^2- , OPSO ₂ ^2- or OSO ₃ ^- ;

[0081] R ₉ , R ₅ and R ₁ are OSO ₃ ^2- , and R ₇ , R ₃ and R ₁₁ are OSO ₃ ^- ;

[0082] R ₉ , R ₅ and R ₁ are OSO ₃ ^- , and R ₇ , R ₃ and R ₁₁ are OPO ₃ ^2- ;

[0083] R ₃ , R ₁ and R ₁₁ are OSO ₃ ^- , and R ₉ , R ₇ and R ₅ are OPO ₃ ^2- ;

[0084] R ₃ , R ₁ and R ₁₁ are OPO ₃ ^2- , and R ₉ , R ₇ and R ₅ are OSO ₃ ^- ;

[0085] R ₇ and R ₁ are OPO ₃ ^2- , and R ₉ , R ₅ , R ₃ and R ₁₁ are OPO ₃ ^2- ;

[0086] R ₇ and R ₁ are OSO ₃ ^- , and R ₉ , R ₅ , R ₃ , and R ₁₁ are OPO3 ^2- ;

[0087] R ₇ and R ₅ are OSO ₃ ^2- , and R ₉ , R ₃ , R ₁ and R ₁₁ are OSO ₃ ^- ; or,

[0088] R ₇ and R ₅ are OSO ₃ ^- , and R ₉ , R ₃ , R ₁ and R ₁₁ are OPO ₃ ^2- .

[0089] The inositol phosphates of the present invention also include compounds that are formed as metabolites during physiological dephosphorylation (or desulfation or desulfation in the case of compounds containing sulfate or thiophosphate groups).

[0090] In some aspects, the compound administered in the dosage according to the methods disclosed herein is a prodrug that, upon hydrolysis or other intracellular or extracellular transformation, yields an inositol phosphate of the present invention.

[0091] The inositol phosphates of the present invention also include any combination of inositol phosphate, inositol phosphate analogs and derivatives thereof disclosed herein.

[0092] All compounds of formula I contain radicals with C-O-P or C-O-S bonds, which give the compounds an affinity for calcium-containing crystals and a sufficiently labile bond to be hydrolyzed in vivo, thereby preventing irreversible binding to calcium-containing crystals such as hydroxyapatite (HAP) in bone, which can have a negative effect on bone remodeling, as is the case with bisphosphonates when administered long-term, since these compounds contain P-C-P bonds that cannot be hydrolyzed by the body. On the other hand, phosphorylated compounds that do not contain these C-O-P bonds, such as pyrophosphates, whose P-O-P bonds mean that they are too easily hydrolyzed in the intestine, meaning that only parenteral administration is advisable. The compounds of the present invention with C-O-P bonds, C-O-S bonds and combinations thereof represent an adequate middle ground due to their potency and the fact that the body has mechanisms to eliminate said compounds, thereby reducing the risk of side effects (e.g., compounds with P-C-P bonds can have half-lives of several months in vivo, thereby affecting, for example, bone remodeling).

[0093] The term "alkyl" or "alkyl group" in the context of the present invention refers to a saturated hydrocarbon moiety, which may be straight, branched, cyclic, or cyclic with straight or branched side chains. The term "alkyl" includes partially unsaturated hydrocarbons such as propenyl. Examples are methyl, ethyl, n- or isobutyl, n- or cyclohexyl. The term "alkyl" may extend to alkyl groups linked or bridged by heteroatoms. Heteroatoms in the context of the present invention are nitrogen (N), sulfur (S), and oxygen (O).

[0094] An "amine functional group" or "amino group" is an NR'R'' functional group, wherein R' and R'' are independently selected from hydrogen and _C1 - _C5 alkyl. In some aspects, R' and R'' are selected from hydrogen and _C1 - _C3 alkyl. A "hydroxy functional group" or "hydroxy group" is OH.

[0095] A "thiol functional group" or "thiol group" is SH. A "carboxylic acid functional group" or "carboxylic acid group" is COOH or its anion, COO ^- . A "carboxylic acid amide" is CONR'R'' with R' and R'' independently having the meanings given above. A "sulfonic acid" is SO ₃ H. A "sulfonic acid amide" is SO ₂ NR'R'' with R' and R'' independently having the meanings given above.

[0096] "C ₁ -C ₃ alkyl" in the context of the present invention refers to a saturated straight-chain or branched hydrocarbon containing 1, 2 or 3 carbon atoms, wherein one carbon-carbon bond may be unsaturated and one CH ₂ moiety may be replaced by oxygen (ether bridge). Non-limiting examples of C ₁ -C ₃ alkyl include methyl, ethyl, propyl, prop-2-enyl and prop-2-ynyl.

[0097] “C ₁ -C ₅ alkyl” in the context of the present invention refers to a saturated straight or branched hydrocarbon containing 1, 2, 3, 4 or 5 carbon atoms, wherein one or two carbon-carbon bonds may be unsaturated and one CH ₂ moiety may be replaced by oxygen (ether bridge). Non-limiting examples of _C1 - _C5 alkyl include the examples given above for _C1 - _C3 alkyl, and additionally n-butyl, 2-methylpropyl, tert-butyl, 3-methylbut-2-enyl, 2-methylbut-3-enyl, 3-methylbut-3-enyl, n-pentyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 1,2-dimethylpropyl, but-3-enyl, but-3-ynyl, and pent-4-ynyl. “ _C3 - _C10 alkyl” as used herein refers to a saturated straight-chain hydrocarbon containing 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms, wherein 1, 2 or 3 carbon-carbon bonds may be unsaturated and one _CH2 moiety may be replaced by oxygen (ether bridge).

[0098] The term "C _1-30 alkyl" as a group or part of a group refers to a straight or branched chain alkyl group containing from 1 to 30 carbon atoms, including, among others, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, decyl and dodecyl groups.

[0099] The term "C _2-30 alkenyl" refers to a straight or branched alkyl chain containing from 2 to 30 carbon atoms and also containing one or more double bonds. Examples include, but are not limited to, ethenyl, 1-propenyl, 2-propenyl, isopropenyl, 1-butenyl, 2-butenyl, 3-butenyl, and 1,3-butadienyl.

[0100] The term "C _2-30 alkynyl" refers to a straight or branched alkyl chain containing from 2 to 30 carbon atoms and also containing one or more triple bonds. Examples include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3-butynyl, and 1,3-butadiynyl.

[0101] "Cy ₁ group" refers to a 3- to 10-membered carbocyclic or heterocyclic ring, which may be saturated, partially unsaturated, or aromatic, and which is bonded to the rest of the molecule through any available carbon atom. In the heterocyclic state, Cy ₁ contains from one to four heteroatoms selected from N, O, and S. Moreover, Cy ₁ may optionally be fused with up to four five- or six-membered carbocyclic or heterocyclic rings, which may be saturated, partially unsaturated, or aromatic. If the fused ring is a heterocycle, said ring contains one or two heteroatoms selected from N, O and S. Examples of Cy ₁ include, but are not limited to, phenyl, naphthyl, thienyl, furyl, pyrrolyl, thiazolyl, isothiazolyl, imidazolyl, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, tetrazolyl, 1,3,4-thiadiazolyl, 1,2,4-thiadiazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, benzimidazolyl, benzofuranyl, isobenzofuranyl, indolyl, isoindolyl, benzothiophenyl, benzothiazolyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, azetidinyl and aziridinyl.

[0102] "C _1-30 alkoxy group" as a group or part of a group refers to a -OC _1-30 alkyl group, wherein the C _1-30 alkyl part has the same meaning as above. Examples include methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, sec-butoxy and tert-butoxy.

[0103] "C _1-30 alkylthionyl group" as a group or part of a group refers to a -SOC _1-30 alkyl group, wherein the C _1-30 alkyl part has the same meaning as above. Examples include methylthionyl, ethylthionyl, propithionyl, isopropylthionyl, butylthionyl, isobutylthionyl, sec-butylthionyl and tert-butylthionyl.

[0104] "C _1-30 acyloxy group" as a group or part of a group refers to a -COC _1-30 alkyl group, wherein the C _1-30 alkyl part has the same meaning as above. Examples include acetyl, ethanoyl, propanoyl and 2,2-diisopropylpentanoyl.

[0105] The "halogen radical" or its abbreviation "halogen" refers to fluorine, chlorine, bromine, and iodine.

[0106] "Trihalogen-C _1-30 alkyl group" refers to a group obtained by substituting three hydrogen atoms of a C _1-30 alkyl group with three halogen radicals as defined above. Examples include, but are not limited to, trifluoromethyl, tribromomethyl, trichloromethyl, triiodomethyl, trifluoroethyl, tribromoethyl, trichloroethyl, triiodoethyl, tribromopropyl, trichloropropyl, and triiodopropyl.

[0107] "-NHC _1-30 alkyl group" refers to a group obtained by replacing one hydrogen atom of the -NH ₂ group with a C _1-30 alkyl group as defined above. Examples include, but are not limited to, methylamine, ethylamine, propylamine, butylamine, and pentylamine.

[0108] "The -N(C _1-30 alkyl) ₂ group" refers to a group obtained by replacing two hydrogen atoms of the -NH ₂ group with a C _1-30 alkyl group as defined above. Examples include, but are not limited to, dimethylamine, diethylamine, diisopropylamine, dibutylamine, and diisobutylamine.

[0109] The expression "optionally substituted with one or more" means the possibility that the group can be substituted with one or more (e.g., 1, 2, 3, or 4) substituents. In some aspects, the group can be substituted with 1, 2, or 3 substituents, and even 1 or 2 substituents, provided that the group has a sufficient number of positions that can be accessed for substitution. If substituents are present, they can be the same or different and can be in any accessible position.

[0110] In some aspects, the inositol phosphates of the present invention include the compounds disclosed in WO2017098033 and WO2017098047 and US 9358243. In some aspects, the inositol phosphates useful in the present invention include the compounds shown in Figs. 1-6.

[0111] According to some aspects, inositol phosphates, inositol phosphate analogs and derivatives thereof comprise compounds of formula (VIII), formula (GC) or formula (X):

^[ 0112] wherein each X is independently selected from _OPO32- , _OPSO22- ^, or ^OSO3- ; Z is an alkyl chain containing 1-3 carbon atoms and/or heteroatoms, optionally containing a group X, wherein X is also selected from _OPO32- , ^OPSO22- , or _OSO3- ^; and _R1 is an optional heterologous moiety (see Section 2.2 below). In some aspects ^, the molecule comprises more than one heterologous moiety, in which case the heterologous moieties may be _the same or different.

[0113] According to some aspects, Z used in formula (VIII) is CH ₂ , CHX, CHR ¹ , CXR ¹ , CH ₂ -CH ₂ , CH ₂ -CHX, CHX-CHX, CHR ¹ -CHX, CXR ¹ -CHX, CHR ¹ -CH ₂ , CXR ¹ -CH ₂ , CHR ¹ -CHOH, CH ₂ -CH ₂ -CH ₂ , CH ₂ -O-CH ₂ , CHOH-CH ₂ -CH ₂ , CHOH-CHOH-CHR ¹ , CHOH-CHR ¹ -ChOH, CHX-CH ₂ -CH ₂ , CH ₂ -CHX-CH ₂ , CHX-CHX-CH ₂ , CHX-CH ₂ -CHX or CHX-CHR ¹ -CHX, wherein X is independently selected from OPO ₃ ^2- , OPSO ₂ ^{2- ,} and OSO ₃ ^- .

[0114] In some aspects, Z as used in formula (VIII) is (CHX) _p CHX(CHX) _q ; wherein each p and q is independently from 0 to 2, provided that (p+q) is 0, 1, or 2; one or two or three X may be a heterologous moiety (e.g., PEG), and the remaining X are independently selected from OPO ₃ ^2- , OPSO ₂ ^2- , and OSO ₃ ^- . In some aspects, not all of the X in Z are OPO ₃ ^2- . In some aspects, not all of the X in Z are OSO ₃ ^- .

[0115] In some aspects, one, two, or three of the X in the compounds of formula (VIII), formula (IX), or formula (X) can be a heterologous moiety, and the remaining X can be independently selected from OPO ₃ ^2- , OPSO ₂ ^{2- ,} or OSO ₃ ^- .

[0116] Formula (VII) above describes a five-membered, six-membered or seven-membered alkyl ring, and an optional heterologous moiety or moieties are attached to one of the carbon atoms forming the ring.

[0117] In some aspects, inositol phosphates, inositol phosphate analogs, and derivatives thereof used, for example, in the methods and compositions disclosed herein include compounds of formula (XI) or formula (XII):

[0118] where:

[0119] X ² is OSO ₃ ^- , and X ¹ , X ³ , X ⁴ , X ⁵ and X ⁶ are independently selected from OPO ₃ ^2- , OPSO ₂ ^2- or OSO ₃ ^- ;

[0120] X ¹ , X ³ and X ⁵ are OSO ₃ ^2- , and X ² , X ⁴ and X ⁶ are OSO ₃ ^- ;

[0121] X ¹ , X ³ and X ⁵ are OSO ₃ ^- , and X ² , X ⁴ and X ⁶ are OPO ₃ ^2- ;

[0122] X ⁴ , X ⁵ and X ⁶ are OSO ₃ ^- , and X ¹ , X ² and X ³ are OPO ₃ ^2- ;

[0123] X ⁴ , X ⁵ and X ⁶ are OSO ₃ ^2- , and X ¹ , X ² and X ³ are OSO ₃ ^- ;

[0124] X ² and X ⁵ are OPO ₃ ^2- and X ¹ , X ³ , X ⁴ , and X ⁶ are OPO ₃ ^2- ;

[0125] X ² and X ⁵ are OSO ₃ ^- , and X ¹ , X ³ , X ⁴ , and X ⁶ are OPO ₃ ^2- ;

[0126] X ² and X ³ are OSO ₃ ^2- , and X ¹ , X ⁴ , X ⁵ , and X ⁶ are OSO ₃ ^- ; or,

[0127] X ² and X ³ are OSO ₃ ^- , and X ¹ , X ⁴ , X ⁵ , and X ⁶ are OPO ₃ ^2- .

[0128] In some aspects, the inositol phosphates of the present invention or their metabolites can be detected and/or quantified using the methods disclosed in US 9,612,250. See also US 8,377,909, US 8,778,912 and US 20070066574.

[0129] The compounds disclosed herein may be present in any form commonly used in pharmaceutical technology. Particular aspects include, but are not limited to, a sodium salt, a magnesium salt, a potassium salt, an ammonium salt, a free acid, or a mixture of the preceding forms. Other pharmaceutically acceptable salts are known to those skilled in the art and can be readily prepared. According to a particular aspect, the compound for use as defined in the first aspect of the present invention is a sodium salt, such as inositol hexaphosphate hexasodium.

[0130] The present invention also provides sodium salts of inositol monophosphate, inositol diphosphate, inositol triphosphate, inositol tetraphosphate and inositol pentaphosphate in any isomeric form of inositol, in particular myo-inositol. A specific example of compounds for use in the present invention is the hexasodium salt of myo-inositol hexaphosphate. Sodium salts provide several advantages in terms of production and impurity levels in the resulting IP6 formulations.

2.2 Heterologous fragment

[0131] According to some aspects, the present invention relates to a compound of general formula I as defined above, wherein the heterologous moiety is selected from a radical of formula V, a radical of formula VI and a radical of formula VH

[0132] and where n is an integer in the range from 2 to 200, and R ₁₃ is selected from H, methyl or ethyl.

[0133] According to some aspects, the compounds for use in the present invention, for example, the inositol phosphate derivatives of the present invention, may comprise one or two radicals selected from radicals of formulas V, VI and VII. These radicals are heterologous moieties that impart an advantageous property relative to a corresponding molecule lacking such heterologous moiety or moieties. Examples of such advantageous properties that can be imparted by a heterologous moiety or combination thereof to inositol phosphate or inositol phosphate analogs include, but are not limited to, (a) increased solubility, (b) decreased degradation or metabolism rate, (c) increased plasma half-life, (d) decreased hepatic metabolism rate, (e) decreased clearance rate, (f) decreased toxicity, (g) decreased irritability and (h) decreased side effects, among others. These advantageous properties can be assessed or quantified using methods known in the art without undue experimentation.

[0134] In some aspects, the heterologous moiety is, for example, polyethylene glycol (PEG) or polyglycerol (PG). Thus, in certain aspects, the compound for use in the present invention is any of the compounds as defined in the aspects disclosed above, comprising a heterologous moiety, that is, one of the radicals of formula I is selected from the radicals of formulas V, VI and VII. In some aspects, the heterologous moiety comprises polyethylene glycol (PEG). In certain aspects, the heterologous moiety consists of polyethylene glycol, that is, at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ of the compound of formula I according to the first aspect of the present invention is a radical of formula V. Alternatively, the heterologous moiety comprises polyglycerol. According to certain aspects, the heterologous moiety consists of a polyglycerol, i.e. at least one of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ of the compound of formula I according to the first aspect of the present invention is selected from a radical of formula VI or VII. According to other aspects, the compound of formula I according to the first aspect of the present invention comprises one, two or three radicals selected from a radical of formula VI or VII, such as two PEGs (a radical of formula V), three PEGs, two polyglycerols (a radical of formula VI), three PGs or any combinations thereof, such as one PEG and one PG, or two PEGs and one polyglycerol. According to certain aspects, all remaining radicals of formula I (i.e. those that are not radicals selected from V, VI and VII) are radicals selected from II, III and IV. According to some aspects, the compound of formula I according to the first aspect of the present invention comprises two radicals selected from a radical of formula VI or VII, for example two PEG (radical of formula V) or two polyglycerols (radical of formula VI) or one PEG, one polyglycerol and all remaining radicals are radicals of formula II. According to some aspects, R ₃ and R ₇ of the compound of formula I are selected from a radical of formula V, VI and VII. According to some aspects, R ₃ and R ₇ of the compound of formula I are radicals of formula V, and R ₁ , R ₅ , R ₉ and R ₁₁ of the compound of formula I are radicals of formula II.

[0135] The radicals of formulas V, VI and VII contain R ₁₃ =H, methyl or ethyl, and n is an integer from 2 to 200. In some aspects, R ₁₃ =H. According to specific aspects, n is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 189, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199 or 200.

[0136] In some aspects, n is 2-10, 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90, 90-100, 100-110, 110-120, 120-130, 130-140, 140-150, 150-160, 160-170, 170-180, 180-190, or 190-200.

[0137] According to some specific aspects, n is characterized by a value from 2 to 200, from 2 to 20, from 10 to 30, or from 9 to 45.

[0138] In some aspects, the PEG is a branched PEG. Branched PEGs contain three to ten PEG chains extending from a central backbone group.

[0139] According to certain aspects, the PEG moiety is a monodisperse polyethylene glycol. In the context of the present invention, monodisperse polyethylene glycol (mdPEG) is a PEG that has one specified chain length and molecular weight. mdPEG is typically obtained by isolation from a polymerization mixture by chromatography. In some formulas, the monodisperse PEG moiety is abbreviated as mdPEG. According to some aspects, the PEG is a star PEG. Star PEGs contain from 10 to 100 PEG chains originating from a central backbone group.

[0140] In some aspects, the PEG is a comb PEG. Comb PEG comprises multiple PEG chains, typically grafted onto a polymer backbone.

[0141] According to certain aspects, the PEG has a molar mass of from 100 g/mol to 3000 g/mol, in particular from 100 g/mol to 2500 g/mol, more in particular from about 100 g/mol to 2000 g/mol. According to certain aspects, the PEG has a molar mass of from 200 g/mol to 3000 g/mol, in particular from 300 g/mol to 2500 g/mol, more in particular from about 400 g/mol to 2000 g/mol.

[0142] In some aspects, the PEG is PEG ₁₀₀ , PEG ₂₀₀ , PEG ₃₀₀ , PEG ₄₀₀ , PEG ₅₀₀ , PEG ₆₀₀ , PEG ₇₀₀ , PEG ₈₀₀ , PEG ₉₀₀ , PEG ₁₀₀₀ , PEG ₁₁₀₀ , PEG ₁₂₀₀ , PEG ₁₃₀₀ , PEG ₁₄₀₀ , PEG ₁₅₀₀ , PEG ₁₆₀₀ , PEG ₁₇₀₀ , PEG ₁₈₀₀ , PEG ₁₉₀₀ , PEG ₂₀₀₀ , PEG ₂₁₀₀ , PEG _{2200 , PEG 2300, PEG 2400, PEG 2500 , PEG1600} , PEG ₁₇₀₀ , PEG ₁₈₀₀ , PEG ₁₉₀₀ , PEG ₂₀₀₀ , PEG ₂₁₀₀ , PEG ₂₂₀₀ , PEG ₂₃₀₀ , PEG ₂₄₀₀ , PEG ₂₅₀₀ , PEG ₂₆₀₀ , PEG ₂₇₀₀ , PEG ₂₈₀₀ , PEG ₂₉₀₀ , or PEG _3000. In one particular aspect, the PEG is PEG _400. In another particular aspect, the PEG is PEG ₂₀₀₀ .

[0143] According to other specific aspects, R ₃ and/or R ₇ of the compound of formula I is a radical of formula V wherein R ₁₃ is H and n is an integer from 9 to 45. According to other specific aspects, R ₃ and R ₇ of the compound of formula I is a radical of formula V wherein R ₁₃ is H and n is an integer from 9 to 45, and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II. According to other specific aspects, the compound of formula I is a sodium salt, wherein R ₃ and R ₇ = a radical of formula V wherein R ₁₃ is H and n is an integer from 9 to 45, and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II.

[0144] In certain other aspects, the heterologous moiety is a polyglycerol (PG) of the formula ((R ₃ -O-(CH ₂ -CHOH-CH ₂ O) _n -), wherein R ₃ is hydrogen, methyl, or ethyl, and n is from 3 to 200. In some aspects, n is from 3 to 20. In some aspects, n is from 10 to 30. In some alternatives of these aspects, n is from 9 to 45. In some aspects, the heterologous moiety is a branched polyglycerol of the formula (R ³ -O-(CH ₂ -CHOR ⁵ -CH ₂ -O) _n -), wherein R ⁵ is hydrogen, or a linear glycerol chain of the formula (R ³ -O-(CH ₂ -CHOH-CH ₂ -O) _n -), and R ³ is hydrogen, methyl or ethyl. In some aspects, the heterologous moiety is a hyperbranched polyglycerol described by the formula (R ³ -O-(CH ₂ -CHOR ⁵ -CH ₂ -O) _n -), wherein R ⁵ is hydrogen or a glycerol chain described by the formula (R ³ -O-(CH ₂ -CHOR ⁶ -CH ₂ -O) _n -), wherein R ⁶ is hydrogen or a glycerol chain described by the formula (R ³ -O-(CH ₂ -CHOR ⁷ -CH ₂ -O) _n -), wherein R ⁷ is hydrogen or a linear glycerol chain described by the formula (R ³ -O-(CH ₂ -CHOH-CH ₂ -O) _n -), and R ³ is hydrogen, methyl, or ethyl. Hyperbranched glycerol and methods for synthesizing the same are known in the art. See Oudshorn M, et al, Biomaterials 2006;27:5471–5479, Wilms D, et al, Acc Chem Res 2010;43:129–141 and references cited therein.

[0145] According to certain aspects, PG has a molar mass of from 100 g/mol to 3000 g/mol, in particular from 100 g/mol to 2500 g/mol, more in particular about 100 g/mol to 2000 g/mol. According to certain aspects, PG has a molar mass of from 200 g/mol to 3000 g/mol, in particular from 300 g/mol to 2500 g/mol, more in particular about 400 g/mol to 2000 g/mol.

[0146] In some aspects, PG is PG ₁₀₀ , PG ₂₀₀ , PG ₃₀₀ , PG ₄₀₀ , PG ₅₀₀ , PG ₆₀₀ , PG ₇₀₀ , PG ₈₀₀ , PG ₉₀₀ , PG ₁₀₀₀ , PG ₁₁₀₀ , PG ₁₂₀₀ , PG ₁₃₀₀ , PG ₁₄₀₀ , PG ₁₅₀₀ , PG ₁₆₀₀ , PG ₁₇₀₀ , PG ₁₈₀₀ , PG ₁₉₀₀ , PG ₂₀₀₀ , PG ₂₁₀₀ , PG ₂₂₀₀ , PG ₂₃₀₀ , PG ₂₄₀₀ , PG ₂₅₀₀ , PG ₁₆₀₀ , PG ₁₇₀₀ , PG ₁₈₀₀ , PG ₁₉₀₀ , PG ₂₀₀₀ , PG ₂₁₀₀ , PG ₂₂₀₀ , PG ₂₃₀₀ , PG ₂₄₀₀ , PG ₂₅₀₀ , PG ₂₆₀₀ , PG ₂₇₀₀ , PG ₂₈₀₀ , PG ₂₉₀₀ or PG _3000. According to one particular aspect, the PG is PG _400. According to another particular aspect, the PG is PG ₂₀₀₀ .

[0147] According to other specific aspects, R ₃ and/or R ₇ of the compound of formula I is a radical of formula VI wherein R ₁₃ is H and n is an integer from 9 to 45. According to other specific aspects, R ₃ and R ₇ of the compound of formula I is a radical of formula VI wherein R ₁₃ is H and n is an integer from 9 to 45 and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II. According to other specific aspects, the compound of formula I is a sodium salt wherein R ₃ and R ₇ = a radical of formula VI wherein R ₁₃ is H and n is an integer from 9 to 45 and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II.

3. Pharmaceutical compositions

[0148] According to another aspect, the present invention also relates to pharmaceutical compositions comprising a compound as defined in any of the aspects disclosed above. According to some aspects, a pharmaceutical composition comprises a compound as defined in any of the aspects disclosed above, together with one or more pharmaceutically acceptable excipients or carriers. These pharmaceutical compositions are intended for use in increasing tissue perfusion and/or oxygenation in a subject in need thereof. According to some aspects, these pharmaceutical compositions are intended for use in the treatment or prevention of ischemia and/or an ischemia-related disease or condition. According to some aspects, the pharmaceutical compositions of the present invention are used to treat or prevent PAD or CLI.

[0149] As used herein, the term "excipient" refers to a substance that facilitates the absorption of the elements of a pharmaceutical composition, stabilizes said elements, activates, or assists in the preparation of the composition. Thus, examples of excipients used in parenteral formulations include, but are not limited to, antimicrobial agents (e.g., benzalkonium chloride, metacresol, thiomersal), cosolvents (e.g., ethanol), buffers, and pH adjusting factors (e.g., carbonate, citrate, phosphate solutions).

[0150] As in the case of an excipient, a "pharmaceutically acceptable carrier" is a substance used in a composition to dilute any of the components contained therein to a certain volume or weight. A pharmaceutically acceptable carrier is an inert substance or a substance with an effect similar to any of the elements that make up the pharmaceutical composition according to the present invention. The role of said carrier is to ensure the inclusion of other elements, to improve dosing and administration, or to give the composition consistency and form.

[0151] The pharmaceutical compositions may comprise from about 1% to about 95% of a compound as defined in any of the aspects disclosed above. According to some aspects, the pharmaceutical compositions of the present invention may comprise, for example, from about 20% to about 90%, or from 20% to 80%, or from 20% to 70%, or from 20% to 60%, or from 20% to 50%, or from 30% to 90%, or from 40% to 90%, or from 50% to 90%, or from 60% to 90%, or from 30% to 70% of a compound as defined in any of the aspects disclosed above.

[0152] In some aspects, the concentration of the inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate or an analog or derivative thereof, or a combination thereof) in each dose of the pharmaceutical composition is from about 12.5 mM to about 135 mM. In some versions of this aspect, the concentration of the inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate or an analog or derivative thereof, or a combination thereof) in each dose of the pharmaceutical composition is about 25 mM, about 39 mM, or about 114 mM.

[0153] Formulations of a pharmaceutical composition suitable for parenteral administration comprise a compound as defined in any of the aspects disclosed above, mixed with a pharmaceutically acceptable carrier (e.g., in the form of sterile water or sterile isotonic saline). Such formulations may be prepared, packaged, or sold in a form suitable for bolus administration or for continuous administration. Formulations for injection may be prepared, packaged, or sold in unit dosage form, such as ampoules or multi-dose containers containing a preservative. Formulations for parenteral administration include, but are not limited to, suspensions, solutions, emulsions in oily or aqueous vehicles, pastes, and implantable sustained-release formulations or biodegradable formulations. Such formulations may further include one or more additional ingredients, including, but not limited to, suspending, stabilizing, or dispersing agents.

[0154] According to some aspects, in a parenteral formulation, the active agent (e.g., a compound as defined in any of the aspects disclosed above) is provided in dry (i.e., powder or granular) form for reconstitution with a suitable vehicle (e.g., sterile, pyrogen-free water) prior to parenteral administration of the reconstituted composition.

[0155] The pharmaceutical compositions can be prepared, packaged, or sold in the form of a sterile aqueous or oily injection suspension or solution. This suspension or solution can be formulated according to the known art and can contain, in addition to the active agent (e.g., a compound as defined in any of the aspects disclosed above), additional ingredients such as dispersing agents, wetting agents, or suspending agents described herein. Such sterile injection formulations can be prepared using a non-toxic parenterally acceptable diluent or solvent, such as, for example, water or 1,3-butanediol. Other acceptable diluents and solvents include, but are not limited to, Ringer's solution, isotonic sodium chloride solution, and fixed oils such as synthetic mono- or diglycerides.

[0156] Other formulations useful for parenteral administration include those comprising the active agent (e.g., a compound as defined in any of the aspects disclosed above) in microcrystalline form, in a liposomal preparation, or as a component of a biodegradable polymer system.

[0157] Compositions for sustained release or implantation may contain pharmaceutically acceptable polymeric or hydrophobic materials such as an emulsion, an ion exchange resin, a sparingly soluble polymer, or a sparingly soluble salt.

[0158] Controlled or sustained release formulations of the pharmaceutical composition of the present invention can be prepared using conventional technology. In some cases, the dosage forms used can be presented as slow or controlled release of one or more active agents therein using, for example, hydropropyl methylcellulose, other polymer matrices, gels, permeable membranes, osmotic systems, multilayer coatings, microparticles, liposomes or microspheres, or a combination thereof to provide the desired release profile in varying proportions. Suitable controlled release formulations known in the art, including those described herein, can be readily selected for use with the pharmaceutical compositions of the present invention. Thus, unit dosage forms suitable for parenteral or topical administration, such as injection solutions, gels, creams and ointments, which are adapted for controlled release are encompassed by the present invention.

[0159] Most controlled-release pharmaceutical products share the common goal of improving therapeutics over those achieved by their uncontrolled counterparts. Ideally, the use of an optimally designed controlled-release preparation in medical treatment is characterized by the use of a minimum amount of a therapeutic agent to cure or control a condition in a minimum amount of time. The advantages of controlled-release formulations include increased activity of the therapeutic agent over time, reduced dosing frequency, and increased patient compliance. In addition, controlled-release formulations can be used to influence the time of onset of action or other characteristics, such as the blood level of the therapeutic agent, and thus the occurrence of side effects can be influenced.

[0160] Most controlled release formulations are designed to initially release an amount of the therapeutic agent that rapidly produces the desired therapeutic effect, and gradually and continuously release other amounts of the therapeutic agent to maintain this level of therapeutic effect over an extended period of time. To maintain this constant level of therapeutic agent in the body, the therapeutic agent must be released from the dosage form at a rate that will replace the amount of therapeutic agent that is metabolized and excreted from the body.

[0161] Controlled release of the active agent can be stimulated by various inducers, such as pH, temperature, enzymes, water or other physiological conditions or compounds. The term "controlled release component" in the context of the present invention is defined herein as a compound or compounds, including but not limited to polymers, polymer matrices, gels, permeable membranes, liposomes or microspheres, or a combination thereof, that facilitates controlled release of the active agent.

[0162] In certain aspects, the formulations of the present invention may be, without limitation, short-acting, rapid-replenishing, and controlled-release formulations, such as sustained-release, delayed-release, and pulsatile-release formulations.

[0163] The term sustained release is used in its generally accepted sense to refer to a formulation of a therapeutic agent (e.g., a compound as defined in any of the aspects disclosed above)), which provides a gradual release of the therapeutically active agent over an extended period of time, and this may, although not necessarily, result in substantially constant levels of the therapeutic agent in the blood over an extended period of time. The period of time may be at least a month or more and should represent a release that is longer than the same amount of agent administered in the form of a bolus.

[0164] For sustained release, the compounds can be formulated with a suitable polymer or hydrophobic material that imparts sustained release properties to the compounds. As such, the compounds for use in the method of the present invention can be administered in the form of microparticles, for example, by injection, or in the form of plates or discs by implantation. In certain aspects, the compounds of the present invention are administered to a patient, alone or in combination with another pharmaceutical agent, using a sustained release formulation.

[0165] The term "sustained release" is used herein in its conventional sense to refer to a formulation of a therapeutic agent that provides an initial release of the therapeutic agent after some delay after administration of the therapeutic agent. The delay may be from about 10 minutes up to 12 hours. The term "pulsatile release" is used herein in its conventional sense to refer to a formulation of a therapeutic agent that provides release of the therapeutic agent in such a way as to obtain pulsed profiles of the therapeutic agent in the plasma after administration. The term immediate release is used in its conventional sense to refer to a formulation of a therapeutic agent that provides release of the therapeutic agent immediately after administration.

[0166] Additional formulations and dosage forms of the compositions of the present invention include dosage forms as described in US 6,340,475, US 6,488,962, US 6,451,808, US 5,972,389, US 5,582,837, and US 5,007,790; US 20030147952, 20030104062, 20030104053, 20030044466, 20030039688, and 20020051820; WO 2003035041, WO 2003035040, WO 2003035029, WO 200335177, WO 2003035039, WO 2002096404, WO 2002032416, WO 2001097783, WO 2001056544, WO 2001032217, WO 1998055107, WO 1998011879, WO 1997047285, WO 1993018755 and WO 1990011757.

[0167] The medicinal products according to the present invention are produced by methods known in the art, in particular by conventional mixing, coating, granulation, dissolution or lyophilization.

[0168] The present invention also relates to a compound, a combination of compounds or a pharmaceutical composition as defined in any of the above aspects of the present invention, in the broadest given definition or as set forth in any of the aspects presented above, for use as a medicine.

4. Methods and routes of administration

[0169] According to some aspects, the compound, pharmaceutical composition, or combination preparation as defined in any of the aspects disclosed above is administered together, simultaneously, or sequentially with another therapeutic agent. According to some versions of this aspect, the additional therapeutic agent comprises cilostazol, pentoxifylline, or a combination thereof.

[0170] In some aspects, administration of an effective amount of a compound, pharmaceutical composition, or combination preparation as defined in any of the aspects above is provided. Said compound, pharmaceutical composition, or combination preparation can be administered parenterally, such as intravenously, intraperitoneally, intramuscularly, intraarterially, intradermally, intrathecally, epidurally, or spinally or subcutaneously. Parenteral administration can be accomplished by bolus injection or intravenous infusion.

[0171] According to a particular aspect of the present invention, myo-inositol hexaphosphate (or a formulation containing myo-inositol hexaphosphate, such as SNF472) is administered by intravenous infusion. According to another particular aspect of the present invention, myo-inositol hexaphosphate is administered subcutaneously. According to another aspect, an inositol derivative or a myo-inositol hexaphosphate derivative, such as a compound of formula I wherein R ₃ and R ₇ = a radical of formula V wherein R ₁₃ is H and n is an integer from 2 to 200, and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II (or a sodium salt thereof), is administered by intravenous infusion. According to another aspect, an inositol derivative or a myo-inositol hexaphosphate derivative, for example a compound of formula I wherein R ₃ and R ₇ = a radical of formula V wherein R ₁₃ is H and n is an integer from 2 to 200 and all of R ₁ , R ₅ , R ₉ and R ₁₁ are a radical of formula II (or its sodium salt), is administered subcutaneously.

[0172] Alternatively, the compound, pharmaceutical composition, or combination preparation can be administered as a component of a solution or system for hemodialysis, hemofiltration, or peritoneal dialysis.

[0173] In the specific case of patients undergoing dialysis, a very suitable administration method consists of administering (for example, non-bolus administration) the inositol phosphate of the present invention through a dialysis machine (before or after the filter) instead of directly injecting the inositol phosphate of the present invention into the patient intravenously. Thus, the blood can be treated with the inositol phosphate of the present invention (for example, myo-inositol hexaphosphate) when it leaves the patient's body and circulates through the dialysis circuit, and when the blood containing the inositol phosphate of the present invention returns to the body.

[0174] Thus, according to some aspects, a compound, pharmaceutical composition, or combination preparation as defined in any of the aspects disclosed above is administered to a patient during hemodialysis. According to some aspects, a compound, pharmaceutical composition, or combination preparation as defined in any of the aspects disclosed above is administered to blood removed from a patient during hemodialysis, preferably prior to filtration thereof (i.e., the therapeutic agent is administered to the unfiltered blood of the patient in the dialysis circuit). According to some aspects, the compound is inositol hexaphosphate, in particular the sodium salt of myo-inositol hexaphosphate or a derivative thereof, namely a compound of formula I, wherein R ₃ and R ₇ = a radical of formula V, wherein R ₁₃ is H, and n is an integer from 2 to 200, and R ₁ , R ₅ , R ₉ and R ₁₁ all represent a radical of formula II (or a sodium salt thereof).

[0175] In the case of dialysis patients, administration of the inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) via a dialysis machine allows the blood to equilibrate with the dialysis fluid before being returned to the body; thus, although the inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) may bind ionic calcium, this fact is compensated for when the blood passes through the dialysis filter, thereby eliminating this side effect and significantly improving the safety profile. In addition, administration of the inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) concurrently with hemodialysis, in particular when administered into unfiltered blood taken from a patient during hemodialysis, allows for a reduction in the dose of the compound, with subsequent advantages in terms of reduced toxicity and minimization of unwanted side effects.

[0176] According to some aspects, the compound, pharmaceutical composition, or combination preparation as defined in any of the aspects disclosed above is administered to a patient being treated with hemodialysis, prior to dialysis treatment or after dialysis treatment.

[0177] In general, an effective dose of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) administered by the methods disclosed herein will depend on, for example, the relative potency of the compound in question, the severity of the disease being treated, and the species and body weight of the subject. In some aspects, an effective dose of an inositol phosphate of the present invention for a subject of a particular species (e.g., a human) can be calculated based on experimental data available for another or a control species (e.g., a rat).

[0178] Thus, for example, a dose of myo-inositol hexaphosphate administered as part of a regimen comprising administration of a 20 mg/kg dosage to a rat subject would be equivalent to administration of the same active agent at a dosage of 4.2 mg/kg to a human subject (i.e., a total dose of 300 mg myo-inositol hexaphosphate to a human subject weighing approximately 70 kg). Similarly, a dosage of 40 mg/kg to a rat subject would be equivalent to administration of myo-inositol hexaphosphate at a dosage of 8.4 mg/kg to a human subject, as defined previously. Dosages can be adjusted depending on the subjects' age, species, body weight, body surface area, renal clearance, gender, pathological condition, route of administration, concomitant administration of one or more other drugs, as well as a wide range of physiological and psychological factors using methods known in the art (Pan S, et al., Patient Prefer Adherence 2016; 10:549-560; Pai M, Pharmacotherapy 2012; 32:856-868; Hacker M, et al, Eds, "Pharmacology: Principles and Practice" (Academic Press; Burlington, MA, USA, 2009). As used herein, the term "mg/kg" refers to mg of inositol phosphate according to the present invention per kilogram of body weight (body mass) of the subject.

[0179] In some aspects, a dose of inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) comprises from about 0.001 mg/kg to about 60 mg/kg of inositol phosphate, an inositol phosphate analog, an inositol phosphate derivative, or a combination thereof of the present invention. In some further aspects, a dose of inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.001 mg/kg to about 20.0 mg/kg, from about 20.0 mg/kg to about 40.0 mg/kg, or from about 40.0 mg/kg to about 60.0 mg/kg.

[0180] In some aspects, the dosage of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.001 mg/kg to about 1.0 mg/kg, from about 1.0 mg/kg to about 10.0 mg/kg, from about 10.0 mg/kg to about 20.0 mg/kg, from about 20.0 mg/kg to about 30.0 mg/kg, from about 30.0 mg/kg to about 40.0 mg/kg, from about 40.0 mg/kg to about 50.0 mg/kg, or from about 50.0 mg/kg to about 60.0 mg/kg.

[0181] In some aspects, the dosage of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.001 mg/kg to about 0.5 mg/kg, from about 0.5 mg/kg to about 1.0 mg/kg, from about 1.0 mg/kg to about 5.0 mg/kg, from about 5.0 mg/kg to about 10.0 mg/kg, from about 10.0 mg/kg to about 15.0 mg/kg, from about 15.0 mg/kg to about 20.0 mg/kg, from about 20.0 mg/kg to about 25.0 mg/kg, from about 25.0 mg/kg to about 30.0 mg/kg, from about 30.0 mg/kg to about 35.0 mg/kg, from about 35.0 mg/kg to about 40.0 mg/kg, from about 40.0 mg/kg to about 45.0 mg/kg or from about 45.0 mg/kg to about 50.0 mg/kg.

[0182] In some aspects, the dosage of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.001 mg/kg to about 0.25 mg/kg, from about 0.25 mg/kg to about 0.5 mg/kg, from about 0.5 mg/kg to about 0.75 mg/kg, from about 0.75 mg/kg to about 1.0 mg/kg, from about 1.0 mg/kg to about 2.50 mg/kg, from about 2.50 mg/kg to about 5.0 mg/kg, from about 5.0 mg/kg to about 7.5 mg/kg, from about 7.5 mg/kg to about 10.0 mg/kg, from about 10.0 mg/kg to about 12.5 mg/kg, from about 12.5 mg/kg to about 15.0 mg/kg, from about 15.0 mg/kg to about 17.5 mg/kg, about 17.5 mg/kg to about 20.0 mg/kg, about 20.0 mg/kg to about 22.5 mg/kg, about 22.5 mg/kg to about 25.0 mg/kg, about 25.0 mg/kg to about 27.5 mg/kg, about 27.5 mg/kg to about 30.0 mg/kg, about 30.0 mg/kg to about 32.5 mg/kg, about 32.5 mg/kg to about 35.0 mg/kg, about 35.0 mg/kg to about 37.5 mg/kg, about 37.5 mg/kg to about 40.0 mg/kg, about 40.0 mg/kg to about 42.5 mg/kg, about 42.5 mg/kg to about 45.0 mg/kg, from about 45.0 mg/kg to about 47.5 mg/kg, from about 47.5 mg/kg to about 50.0 mg/kg, from about 50.0 mg/kg to about 52.5 mg/kg, from about 52.5 mg/kg to about 55.0 mg/kg, from about 55.0 mg/kg to about 57.5 mg/kg, or from about 57.5 mg/kg to about 60.0 mg/kg.

[0183] In some aspects, the dosage of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.25 mg/kg to about 60.0 mg/kg, from about 0.5 mg/kg to about 60.0 mg/kg, from about 0.75 mg/kg to about 60.0 mg/kg, from about 1.0 mg/kg to about 60.0 mg/kg, from about 2.50 mg/kg to about 60.0 mg/kg, from about 5.0 mg/kg to about 60.0 mg/kg, from about 7.5 mg/kg to about 60.0 mg/kg, from about 10.0 mg/kg to about 60.0 mg/kg, from about 12.5 mg/kg to about 60.0 mg/kg, from about 15.0 mg/kg to about 60.0 mg/kg, from about 17.5 mg/kg to about 60.0 mg/kg, from about 20.0 mg/kg to about 60.0 mg/kg, from about 22.5 mg/kg to about 60.0 mg/kg, from about 25.0 mg/kg to about 60.0 mg/kg, from about 27.5 mg/kg to about 60.0 mg/kg, from about 30.0 mg/kg to about 60.0 mg/kg, from about 32.5 mg/kg to about 60.0 mg/kg, from about 35.0 mg/kg to about 60.0 mg/kg, from about 37.5 mg/kg to about 60.0 mg/kg, from about 40.0 mg/kg to about 60.0 mg/kg, from about 42.5 mg/kg to about 60.0 mg/kg, from about 45.0 mg/kg to approximately 60.0 mg/kg, from approximately 47.5 mg/kg to approximately 60.0 mg/kg, from approximately 50.0 mg/kg to approximately 60.0 mg/kg, from approximately 52.5 mg/kg to approximately 60.0 mg/kg, from approximately 55.0 mg/kg to approximately 60.0 mg/kg, or from approximately 57.5 mg/kg to approximately 60.0 mg/kg.

[0184] In some aspects, the dosage of an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) is from about 0.001 mg/kg to about 57.5 mg/kg, from about 0.001 mg/kg to about 55.0 mg/kg, from about 0.001 mg/kg to about 52.5 mg/kg, from about 0.001 mg/kg to about 50.0 mg/kg, from about 0.001 mg/kg to about 47.5 mg/kg, from about 0.001 mg/kg to about 45.0 mg/kg, from about 0.001 mg/kg to about 42.5 mg/kg, from about 0.001 mg/kg to about 40.0 mg/kg, from about 0.001 mg/kg to about 37.5 mg/kg, from about 0.001 mg/kg to about 35.0 mg/kg, from about 0.001 mg/kg to about 32.5 mg/kg, from about 0.001 mg/kg to about 30.0 mg/kg, from about 0.001 mg/kg to about 27.5 mg/kg, from about 0.001 mg/kg to about 25.0 mg/kg, from about 0.001 mg/kg to about 22.5 mg/kg, from about 0.001 mg/kg to about 20.0 mg/kg, from about 0.001 mg/kg to about 27.5 mg/kg, from about 0.001 mg/kg to about 25.0 mg/kg, from about 0.001 mg/kg to about 22.5 mg/kg, from about 0.001 mg/kg to about 20.0 mg/kg, from about 0.001 mg/kg to about 17.5 mg/kg, from about 0.001 mg/kg to about 15.0 mg/kg, from about 0.001 mg/kg to about 12.5 mg/kg, from about 0.001 mg/kg to about 10.0 mg/kg, from about 0.001 mg/kg to about 7.5 mg/kg, from about 0.001 mg/kg to about 5.0 mg/kg, or from about 0.001 mg/kg to about 2.5 mg/kg.

5. Indications

[0185] The compounds, pharmaceutical compositions, combination preparations, methods and routes of administration as defined in any of the aspects disclosed above can be used to increase tissue perfusion and/or oxygenation in a subject in need thereof.

[0186] As used herein, the terms "ischemia-related disease or condition" refer to any disease or condition associated with or resulting from an ischemic event or injury. Examples of ischemia-related diseases or conditions include, but are not limited to, cerebrovascular (e.g., stroke, transient ischemic attack (TIA), subarachnoid hemorrhage, vascular dementia), cardiovascular (e.g., myocardial infarction, angina), gastrointestinal (e.g., colitis), peripheral (e.g., acute limb ischemia), and cutaneous (e.g., cyanosis, gangrene) diseases or conditions.

[0187] According to some aspects, the compounds, pharmaceutical compositions, combination preparations, methods and routes of administration of the present invention can be used to treat or prevent ischemia and/or an ischemia-related disease or condition in a subject in need thereof.

[0188] As used herein, the term "renal failure" refers to a disease that causes a progressive loss of kidney function with a concomitant decrease in the glomerular filtration rate or index (GFR). Renal failure is also known as renal impairment or kidney disease. Kidney disease can be classified as (i) acute kidney injury (AKI), a progressive loss of kidney function that typically causes oliguria and fluid and electrolyte imbalance, and (ii) chronic kidney disease (CKD), a much slower loss of kidney function over months or years. There are five stages of CKD based on the degree of kidney function based on GFR: (a) stage 1, normal or high GFR (>90 mL/min), (b) stage 2, mild CKD, GFR=60-89 mi/min, (c) stage 3, moderate CKD, GFR=30-59 m/min, (d) stage 4, severe CKD, GFR=15-29 mL/min, and (e) stage 5, end-stage CKD, GFR<15 mL/min. Stage 5 requires dialysis or kidney transplantation to maintain health. AKI and CKD can occur simultaneously, which is known as exacerbation of chronic kidney disease.

[0189] In some aspects, the compounds, pharmaceutical compositions, combination formulations, methods, and routes of administration of the present invention can be used to increase tissue perfusion in a subject with a kidney disease. The kidney disease in the subject can be acute, chronic, or both. In some aspects, the subject is undergoing dialysis (e.g., peritoneal dialysis, hemodialysis). In another form of this aspect, the subject is undergoing hemodialysis. In some other aspects, the subject is not undergoing dialysis (e.g., a subject with CKD stages 1 through 4). In one version of this aspect, the subject is administered an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) at an effective dosage of about 0.001 mg/kg to about 60 mg/kg.

[0190] In some aspects, the compounds, pharmaceutical compositions, combination formulations, methods and routes of administration of the present invention can be used to treat or prevent ischemia and/or an ischemia-related disease or condition in a subject with a kidney disease. The kidney disease in the subject can be acute, chronic, or both. In one version of this aspect, the subject is undergoing dialysis (e.g., peritoneal dialysis, hemodialysis). In another form of this version, the subject is undergoing hemodialysis. In another version of this aspect, the subject is not undergoing dialysis (e.g., a subject with CKD stages 1 through 4). In one version of this aspect, the subject is administered an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) at an effective dosage of about 0.001 mg/kg to about 60 mg/kg.

[0191] In some aspects, the compounds, pharmaceutical compositions, combination formulations, methods, and routes of administration of the present invention can be used to improve the walking ability of a patient in need thereof. In some aspects, the compounds, pharmaceutical compositions, methods, and routes of administration of the present invention can be used to increase the maximum pain-free walking distance (MWD), the maximum pain-free walking time (MWT), or both in a subject in need thereof. In some aspects, the subject has a kidney disease. The kidney disease in the subject can be acute, chronic, or both. In one version of this aspect, the subject is undergoing dialysis (e.g., peritoneal dialysis, hemodialysis). In another form of this version, the subject is undergoing hemodialysis. In another version of this aspect, the subject is not undergoing dialysis (e.g., a subject with CKD stages 1 through 4). According to one version of this aspect, the subject is administered an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) at an effective dosage of about 0.001 mg/kg to about 60 mg/kg.

[0192] The compounds, pharmaceutical compositions, combination preparations, methods and routes of administration of the present invention are particularly useful for increasing tissue perfusion and/or oxygenation in the lower extremities, and especially for the treatment and prevention of peripheral arterial disease. Another condition that can be improved by using the inositol phosphates of the present invention is critical limb ischemia. According to certain aspects, the compounds, pharmaceutical compositions, combination preparations, methods and routes of administration as defined in any of the aspects disclosed above are for use in increasing tissue perfusion and/or oxygenation, especially in the lower extremities, and especially for the treatment and prevention of PAD and/or CLI. In some aspects, the subject is undergoing dialysis (e.g., peritoneal dialysis, hemodialysis). In some further aspects, the subject is undergoing hemodialysis. In some other aspects, the subject is not undergoing dialysis (e.g., a subject with CKD stages 1 to 4). According to one version of this aspect, the subject is administered an inositol phosphate of the present invention (e.g., myo-inositol hexaphosphate) at an effective dosage of about 0.001 mg/kg to about 60 mg/kg.

[0193] The following embodiments further illustrate the scope of the invention:

[0194] Embodiment 1. A compound of general formula I or a pharmaceutically acceptable salt thereof for use in increasing tissue perfusion and/or oxygenation in a subject in need thereof

[0195] where R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are independently selected from OH, a radical of formula II, III, IV, V, VI and VII:

[0196] where: n is an integer in the range from 2 to 200, and R ₁₃ is selected from H, methyl, ethyl and C ₃ -C ₁₀ alkyl;

[0197] provided that:

[0198] at least one of R ₁ , R ₃ , Rs, R ₇ , Rs and R ₁₁ is selected from a radical of formula II, III and IV, and

[0199] zero, one, two or three of R _{1 ;} R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are selected from a radical of formula V, VI and VII.

[0200] Embodiment 2. A compound for use according to embodiment 1, which is intended for the treatment or prevention of peripheral arterial disease.

[0201] Embodiment 3. A compound for use according to any of the previous embodiments, which is intended for the treatment or prevention of critical limb ischemia.

[0202] Embodiment 4. A compound for use according to any of the previous embodiments, which is intended for treating a subject undergoing dialysis, preferably hemodialysis.

[0203] Embodiment 5. A compound for use according to any of the previous embodiments, which is a sodium salt.

[0204] Embodiment 6: A compound for use according to any of the previous embodiments, wherein at least two, at least three, at least four, at least five, or at least 6 of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are selected from radicals of formulas V, VI and VII.

[0205] Embodiment 7. A compound for use according to the previous embodiment, wherein at least two, at least three, at least four, at least five, or at least six of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are a radical of formula V.

[0206] Embodiment 8. A compound for use according to the previous embodiment, wherein the compound of formula I is inositol hexaphosphate.

[0207] Embodiment 9. A compound for use according to the previous embodiment, which is a hexasodium salt.

[0208] Embodiment 10. A compound for use according to embodiment 7, wherein:

[0209] R ₇ is OSO ₃ ^- , and R ₁ , R ₃ , R ₅ , R ₉ and R ₁₁ are independently selected from OPO ₃ ^2- , OPSO ₂ ^2- or OSO ₃ ^- .

[0210] R ₉ , R ₅ and R ₁ are OSO ₃ ^2- , and R ₇ , R ₃ and R ₁₁ are OSO ₃ ^- ;

[0211] R ₉ , Rs and R ₁ are OSO ₃ ^- , and R ₇ , R ₃ and R ₁₁ are OPO ₃ ^2- ;

[0212] R ₃ , R ₁ and R ₁₁ are OSO ₃ ^- , and R ₉ , R ₇ and R ₅ are OPO ₃ ^2- ;

[0213] R ₃ , R ₁ and R ₁₁ are OPO ₃ ^2- , and R ₉ , R ₇ and R ₅ are OSO ₃ ^- ;

[0214] R ₇ and R ₁ are OPO ₃ ^2- , and R ₉ , R ₅ , R ₃ , and R ₁₁ are OPO ₃ ^- ;

[0215] R ₇ and R ₁ are OSO ₃ ^- , and R ₉ , R ₅ , R ₃ , and R ₁₁ are OPO ₃ ^2- ;

[0216] R ₇ and R ₅ are OPO ₃ ^2- , and R ₉ , R ₃ , R ₁ , and R ₁₁ are OSO ₃ ^- ; or,

[0217] R ₇ and R ₅ are OSO ₃ ^- , and R ₉ , R ₃ , R ₁ , and R ₁₁ are OPO ₃ ^2- .

[0218] Embodiment 11. A compound for use according to any of the previous embodiments, wherein the compound of formula I is characterized by the myo-inositol conformation.

[0219] Embodiment 12. A compound for use according to any one of Embodiments 1-6 wherein one, two, or three of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ , and R ₁₁ are selected from a radical of formula V, VI, and VII.

[0220] Embodiment 13. A compound for use according to the previous embodiment, wherein four of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are a radical of formula II and two of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are selected from a radical of formulas V, VI and VII.

[0221] Embodiment 14. A compound for use according to the previous embodiment, wherein four of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are a radical of formula II, and two of R ₁ , R ₃ , R ₅ , R ₇ , R ₉ and R ₁₁ are a radical of formula V.

[0222] Embodiment 15. A compound for use according to any one of embodiments 12-14, wherein:

(i) R ₁ , R ₅ , R ₉ and R ₁₁ represent a radical of formula II, and R ₃ and R ₇ are selected from a radical of formulas V, VI and VII,

(ii) R ₁ , R ₃ , R ₉ and R ₁₁ represent a radical of formula II, and R ₅ and R ₇ are selected from a radical of formulas V, VI and VII.

[0223] Embodiment 16. A compound for use according to the previous embodiment, wherein the radical selected from V, VI and VII is a radical of formula V.

[0224] Embodiment 17. A compound for use according to any one of embodiments 12-16, wherein the radical of formula V, VI or VI is characterized by n in the range of 2 to 200.

[0225] Embodiment 18. A compound for use according to the previous embodiment, wherein n is in the range of 9 to 30.

[0226] Embodiment 19. A compound for use according to the previous embodiment, wherein n is in the range of 15 to 30.

[0227] Embodiment 20. A compound for use according to embodiment 17, wherein n is in the range of 3 to 9.

[0228] Embodiment 21. A compound for use according to any one of Embodiments 12-20 wherein R ₁₃ is H.

[0229] Embodiment 22. A compound for use according to Embodiment 21 wherein R ₁ , R ₅ , R ₉ and R ₁₁ are a radical of formula II, and R ₃ and R ₇ are a radical of formula V.

[0230] Embodiment 23. A pharmaceutical composition for use as defined in any one of embodiments 1-4, comprising a compound as defined in any one of the previous embodiments, together with pharmaceutically acceptable excipients and carriers.

[0231] Embodiment 24. The pharmaceutical composition of the previous embodiment, wherein the compound is present in an amount of 20-90% (w/w) of the total composition.

[0232] Embodiment 25. The pharmaceutical composition of the previous embodiment, wherein the compound is present in an amount of 30-80% (w/w) of the total composition.

[0233] Embodiment 26. The pharmaceutical composition of the previous embodiment, wherein the compound is present in an amount of 40-70% (w/w) of the total composition.

[0234] Embodiment 27. The pharmaceutical composition of any one of embodiments 23-26, wherein the composition is in dry form for reconstitution with a suitable carrier.

[0235] Embodiment 28. The pharmaceutical composition according to any one of embodiments 23-26, wherein the composition is in solution, preferably in isotonic saline solution.

[0236] Embodiment 29. The pharmaceutical composition of any one of embodiments 23-27, which forms part of a hemodialysis, hemofiltration, or peritoneal dialysis solution.

[0237] Embodiment 30. The pharmaceutical composition of any one of embodiments 23-29, wherein the composition is for controlled release.

[0238] Embodiment 31. A compound for use according to any one of embodiments 1-22 or a pharmaceutical composition for use according to any one of embodiments 23-30, which is administered to a patient undergoing dialysis.

[0239] Embodiment 32. The compound for use according to the previous embodiment, wherein the dialysis is hemodialysis.

[0240] Embodiment 33. A compound or pharmaceutical composition for use according to any of embodiments 31-32, which is administered prior to dialysis.

[0241] Embodiment 34. A compound or pharmaceutical composition for use according to any one of embodiments 31-32, which is administered during dialysis.

[0242] Embodiment 35. A compound or pharmaceutical composition for use according to any one of embodiments 31-32, which is administered after dialysis.

[0243] Embodiment 36. A compound or pharmaceutical composition for use according to any one of embodiments 31-35, which is administered parenterally.

[0244] Embodiment 37. A compound or pharmaceutical composition for use according to the previous embodiment, wherein the parenteral administration is intravenous, subcutaneous, or intramuscular.

[0245] Embodiment 38. The compound or pharmaceutical composition for use according to the previous embodiment, wherein intravenous administration is performed by bolus injection or intravenous infusion.

[0246] Embodiment 39. A compound or pharmaceutical composition for use according to embodiment 34, which is administered to unfiltered blood taken from a patient.

[0247] Embodiment 40. A compound or pharmaceutical composition for use according to any of the previous embodiments, wherein the compound is administered to the subject at a therapeutically effective dosage of about 0.001 mg/kg to about 60 mg/kg.

[0248] Embodiment 41. A compound or pharmaceutical composition for use according to embodiment 40, wherein the compound is administered to the subject at a therapeutically effective dosage of about 15 mg/kg to about 45 mg/kg.

[0249] Embodiment 42. A combination preparation comprising: (i) (a) at least one compound according to any one of Embodiments 1-22 or (b) at least one pharmaceutical composition according to any one of Embodiments 23-30 and (ii) at least one additional therapeutic agent for use in providing human health.

[0250] Embodiment 43. The combination preparation of the previous embodiment, wherein the additional therapeutic agent is cilostazol, pentoxifylline, or a combination thereof.

[0251] Embodiment 44. A method for increasing tissue perfusion and/or oxygenation in a subject in need thereof, comprising administering a therapeutically effective amount of a compound or pharmaceutical composition of any one of embodiments 1-41 or a combination preparation of any one of embodiments 42-43 to the subject.

[0252] Embodiment 45. A method for treating or preventing ischemia and/or an ischemia-related disease or condition in a subject in need thereof, comprising administering a therapeutically effective amount of a compound or pharmaceutical composition of any one of embodiments 1-41 or a combination preparation of any one of embodiments 42-43 to the subject.

[0253] Embodiment 46. A method for improving walking ability in a subject in need thereof, comprising administering a therapeutically effective amount of a compound or pharmaceutical composition of any one of embodiments 1-41 or a combination preparation of any one of embodiments 42-43 to the subject.

[0254] Embodiment 47. A method for increasing maximum pain-free walking distance (MWD), maximum pain-free walking time (MWT), or both in a subject in need thereof, comprising administering a therapeutically effective amount of a compound or pharmaceutical composition of any one of embodiments 1-41 or a combination preparation of any one of embodiments 42-43 to the subject.

[0255] Embodiment 48. A method for treating or preventing peripheral arterial disease in a subject in need thereof, comprising administering a therapeutically effective amount of a compound or pharmaceutical composition of any one of embodiments 1-41 or a combination preparation of any one of embodiments 42-43 to the subject.

[0256] Embodiment 49. The methods of any one of embodiments 44-48, wherein the combination preparation is administered co-administered, simultaneously, or sequentially to the subject.

[0257] The present invention is further illustrated by the following examples, which should not be construed as limiting. The contents of all documents cited in this application are incorporated herein by reference in their entirety.

General procedures

1. Blood perfusion in the extremities

[0258] Blood perfusion in a given limb position and ischemic status (i.e., blood perfusion including perfusion unit, perfusion difference, and perfusion ratio) are assessed using laser Doppler perfusion imaging using a PeriCam PSI NR imaging system (Perimed AB, Järfälla, SE). Before measurement, subjects are anesthetized with 3% isoflurane delivered in 100% oxygen at a flow rate of 1 L/min. The perfusion difference and perfusion ratio are calculated by comparing baseline and any specified intermediate or final readings for each group. Blood flow is assessed around the C _max of the active agents tested (i.e., 15 minutes after treatment with SNF472, 20 minutes after treatment with IP4-BIS-PEG100, and 3-4 hours after treatment with cilostazol).

2. Walking ability tests

[0259] Quantitative assessment of walking ability (maximum pain-free walking time (MWT) and maximum pain-free walking distance (MWD)) was measured using a forced incremental treadmill running test. A two-lane rodent treadmill (LE8709TS; PanLab/Harvard Apparatus, Holliston, MA, USA) was used. The treadmill was set at a 15% incline and operated at a speed of 15 m/min (25 cm/s) for the first 5 min, then at a speed of 33 cm/s for the next 5 min, and finally at a speed of 40 cm/s for a maximum of 30 additional min.

[0260] Limb function is assessed around _Cmax of the active agents tested (i.e., 15 minutes after treatment with SNF472 and 3-4 hours after treatment with cilostazol). Subjects are acclimated to a treadmill prior to testing according to an established protocol. Subjects who do not comply with the protocol are excluded from the test.

[0261] Subjects continue to run for 40 minutes or until exhaustion (i.e., they remain on the shock grid for five uninterrupted seconds) during the test. The MWD and MWT are then calculated for each animal.

3. Tissue and blood collection, calcification tests

[0262] Subjects are anesthetized by inhalation of isoflurane. Blood is obtained by exsanguination via cardiac puncture. Subjects are then sacrificed and their tissues (e.g., right and left femoral arteries, aorta) are collected. Blood and tissues are processed and analyzed for calcium content.

[0263] Calcium content in the tissue sample is quantified by inductively coupled plasma optical emission spectrometry (ICP-OES) using an Optima 7300 DV ICP-OES System (PerkinElmer, Inc., Waltham, MA, USA) according to the manufacturer's instructions. Myo-inositol hexaphosphate levels in plasma are quantified using LC-MS/MS chromatographic methods described in the art. See WO2013050603.

Example 1

Prevention of limb ischemia

Effect on blood perfusion

[0264] The prophylactic effects of SNF472, IP4-4,6-bisPEG100 sodium salt, and cilostazol on blood perfusion were tested using a rat model for 12 days. Limb ischemia was induced in subjects starting on D1, except for the sham control group, in which ischemia was not induced. Subjects with induced ischemia were then treated with placebo and active agent formulations on D1 through D12 to assess their effects on preventing limb ischemia. Observations were made at multiple points during treatment from D1 through D12. All subjects were weighed daily before treatment.

1. Induction of limb ischemia

[0265] Fifty-four male Sprague Dawley (SD) rats (Envigo Corp., Huntingdon, UK) weighing approximately 250-275 g were used. Subjects were fed an AO4 diet (Scientific Animal Food &Engineering; Carpe Bio, Amersfoort, The Netherlands). Subjects were divided into 5 groups of 8-10 rats each as follows:

[0266] Group 1 - Control (control group with sham intervention)

[0267] Group 2a Placebo - Physiological saline solution

[0268] Group 2b Placebo - 5% sodium carboxymethylcellulose (CMC) solution

[0269] Group 3 - SNF472 (Na ₆ IP ₆ )

[0270] Group 4 - Sodium salt IP4-4,6-bisPEG100

[0271] Group 5 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ )

[0272] Limb ischemia was induced in subjects in Groups 2a-5 by subcutaneous administration of 120,000 IU/kg vitamin D ₃ (cholecalciferol, Duphafral D ₃ 1000; Zoetis Inc., Parsippany, NJ, USA) in saline (2 mL/kg) every day from D1 to D3. Subjects in _the sham control Group 1 were administered saline (2 mL/kg) subcutaneously every day from D1 to D3 without vitamin D ₃ .

[0273] Subjects in Groups 1 and 2a were administered physiological saline (2 mL/kg) subcutaneously every day from D1 to D12. Subjects in Group 2b were administered 5% sodium CMC in aqueous solution (5 mL/kg) orally every day from D1 to D12.

[0274] Vitamin _D3 administration induced ischemia in the hind limbs of subjects in groups 2a, 2b, and 3-5, as assessed by laser Doppler perfusion imaging.

2. Relief of ischemia and effect on blood perfusion in the extremities - analysis using laser Doppler imaging

[0275] Limb blood perfusion was induced in subjects in Groups 3 and 4 by subcutaneous administration of 20 mg/kg SNF472 (Na ₆ IP ₆ ; free base: 600 g/mol) and sodium salt of IP4-4,6-bisPEG100 (free base 696.27 g/mol), respectively, in physiological saline (2 ml/kg) every day from D1 to D12. Limb blood perfusion was induced in subjects in Group 5 by oral administration of 20 mg/kg _cilostazol ( _C20H27N5O2 , free base 369.46 g/mol; lot number LRAB9590, Sigma- _Aldrich Corp., St. Louis, MO, _USA ) in 5% sodium CMC in aqueous solution (5 mL/kg) each day from D1 to D12.

[0276] Limb ischemia status was assessed on days D0 (baseline), D6, and D12 in all rats using laser Doppler perfusion imaging. The perfusion difference and perfusion ratio were calculated by comparing the baseline at D0 and the readings at D6 and D12 for each group. Specifically, the perfusion difference and perfusion ratio were calculated by comparing the readings of Group 1 (control) and (a) Groups 2a and 2b placebo (i.e., normal saline, 5% sodium CMC in aqueous solution), (b) Group 3 (SNF472), (c) Group 4 (IP4-4,6-bisPEG100 sodium salt), and (d) Group 5 (cilostazol).

[0277] On days when blood perfusion tests were performed using active agents (i.e., D6, D12), dosages were administered as follows:

[0278] Group 3 - SNF472 (Na ₆ IP ₆ ): 15 minutes before reading

[0279] Group 4 - Sodium salt IP4-4,6-bisPEG100: 20 minutes before taking readings

[0280] Group 5 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ): 3-4 hours before taking readings

[0281] According to the scheme above, the active agents will have a maximum serum concentration (C _max ) at the time the test reading is taken.

[0282] Subjects in Group 1 did not show any significant changes in perfusion parameters.

[0283] Administration of SNF472 and IP4-4,6-bisPEG100 sodium salt attenuated the decline in blood perfusion in the hind limbs of subjects in Groups 3-5 (i.e., those receiving vitamin _D3 ) compared to both placebo and cilostazol. These results suggest that SNF472 and IP4-4,6-bisPEG100 are more effective than cilostazol in increasing blood perfusion in the hind limbs of treated subjects. See Figs. 7, 8, and 9.

3. Calcium content and calcification - ICP-OES

[0284] SNF472 demonstrated efficacy against vascular calcification, as SNF472 inhibited aortic calcification (31±16%) following daily subcutaneous administration of 20 mg/kg compared to placebo. Cilostazol was inactive against calcification at the same dose. See Fig. 10.

4. Pharmacokinetics

[0285] Subjects in groups 1, 2, 3, and 5 were sacrificed after exsanguination. They were then autopsied and the aortas were collected. The tissues were lyophilized for 24 h and weighed. The lyophilized tissues were then treated with a 1:1 mixture of HNO ₃ :HClO ₄ in a dry bath incubator for 2-4 h at 180°C. The treated tissues were then diluted with Milli-Q ultrapure water (MilliporeSigma (Merck KGaA), Burlington, MA, USA) to a final volume of 10 ml. The calcium content of the aorta sample was quantified by ICP-OES.

[0286] Subjects in groups 1, 2, 3, 4, and 5 were anesthetized and had their blood drawn at D12. Approximately 8-10 mL of total blood was collected from each subject and divided into plasma (K3EDTA, approximately 6 mL of blood) and serum (approximately 2-3 mL of blood) collection tubes. Plasma was stored in one 600 µL aliquot and several other 500 µL aliquots. Serum was divided into two aliquots.

[0287] Plasma myo-inositol hexaphosphate levels in subjects in Group 3 (i.e., measured at approximately C _max , 15 min after the last dose of SNF472) were quantified using LC-MS/MS chromatographic methods described in the art. See WO2013050603.

[0288] All subjects in Group 3 were adequately exposed to SNF472. Plasma levels 15 minutes after the last subcutaneous administration at D12 were 15,587±ng/mL (24±12 µM). Plasma levels in rats following daily administration of 20 mg/kg were comparable to those found in hemodialysis patients receiving SNF472 at 4.2 mg/kg intravenously.

Example 2

Prevention of limb ischemia

Effect on maximal pain-free walking time (MWT) and pain-free walking distance (MDT)

[0289] The prophylactic effects of SNF472 and cilostazol on blood perfusion, walking ability, and tissue calcification were tested using a rat model for 24 days. Additionally, the effects of combination treatment with SNF472 and cilostazol on blood perfusion were tested. Limb ischemia was induced in subjects from D1 to D3, except for the sham control group in which ischemia was not induced. Subjects with induced ischemia were then treated with placebo and active agent formulations from D1 to D12 to assess their effects on preventing (a) limb ischemia and tissue calcification and (b) walking ability impairment. Treatment was discontinued for all subjects from D13 to D24. Observations were made at multiple points during the treatment phase and the post-treatment phase from D1 to D24. All subjects were weighed daily before treatment.

1. Induction of limb ischemia

[0290] Fifty-four male Sprague Dawley (SD) rats (Envigo Corp., Huntingdon, UK) weighing approximately 250-275 g were used. Subjects were fed an AO4 diet (Scientific Animal Food &Engineering; Carpe Bio, Amersfoort, The Netherlands). Subjects were divided into 5 groups of 8-10 rats each as follows:

[0291] Group 1 - Control (control group with sham intervention)

[0292] Group 2a Placebo - Physiological saline solution

[0293] Group 2b Placebo - 5% sodium salt solution CMC

[0294] Group 3 - SNF472 (Na ₆ IP ₆ )

[0295] Group 4 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ )

[0296] Group 5 - Cilostazol + SNF472

[0297] Limb ischemia was induced in subjects in Groups 2a-5 by subcutaneous administration of 120,000 IU/kg vitamin D ₃ (cholecalciferol, Duphafral D ₃ 1000; Zoetis Inc., Parsippany, NJ, USA) in saline (2 mL/kg) every day from D1 through D3. Subjects in _the sham control Group 1 were administered saline (2 mL/kg) subcutaneously every day from D1 through D3 without vitamin D ₃ .

[0298] Subjects in Groups 1 and 2a were administered physiological saline (2 mL/kg) subcutaneously each day from D1 to D12. Subjects in Group 2b were administered 5% sodium CMC in aqueous solution (5 mL/kg) orally each day from D1 to D12.

2. Relief of ischemia and effect on blood perfusion in the extremities - analysis using laser Doppler imaging

[0299] Subjects in Groups 1 and 2a were administered 2 mL/kg of physiological saline daily from D1 to D12 by the subcutaneous route. Subjects in Group 2b were administered 5 mL/kg orally of 5% sodium salt of CMC in aqueous solution daily from D1 to D12. In addition, subjects in Group 3 were administered 20 mg/kg SNF472 (Na ₆ IP ₆ , free base: 600 g/mol), sodium salt (free base 696.27 g/mol) in physiological saline (2 mL/kg) daily from D1 to D12 by the subcutaneous route. Subjects in Group 4 were orally administered 20 mg/kg _cilostazol ( _C20H27N5O2 , _free base 369.46 g/ _mol ; lot number LRAB9590, Sigma-Aldrich Corp., St. Louis, MO, USA) in 5% sodium CMC in aqueous solution (5 mL/kg) daily from D1 to D12. Subjects in Group 5 were administered (i) ₂₀ mg/kg cilostazol ( _C20H27N5O2 , _{free base} 369.46 g/mol; lot number LRAB9590, Sigma-Aldrich Corp., St. Louis, MO, USA) in 5% sodium salt of CMC in aqueous solution (5 ml/kg) orally followed by (ii) 20 mg/kg _SNF472 ( _Na6IP6 , free base: 600 g/mol) in physiological saline (2 ml/kg) subcutaneously. Administration was performed daily from D1 to D12.

[0300] On days when blood perfusion tests were performed using active agents (i.e., D6, D12, and D18), doses were administered as follows:

[0301] Group 3 - SNF472 (Na ₆ IP ₆ ): 15 minutes before reading

[0302] Group 4 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ): 3 - 4 hours before taking readings

[0303] Group 5 - Cilostazol + SNF472: 15 minutes for SNF472 and 3-4 hours for cilostazol before readings

[0304] According to the scheme above, the active agents will have a maximum plasma concentration (C _max ) at the time the test reading is taken.

[0305] Limb ischemia status was assessed during treatment and after treatment was interrupted (i.e., D0, D6, D12, D18) in all rats using laser Doppler perfusion imaging. Perfusion difference and perfusion ratio were calculated by comparing baseline and any of the readings at D6, D12, and D18 for each group. Specifically, perfusion difference and perfusion ratio were calculated by comparing the readings of Group 1 (control) and (a) Groups 2a and 2b placebo (saline, 5% sodium CMC), (b) Group 3 (SNF472), (c) Group 4 (cilostazol), and (d) Group 5 (cilostazol/SNF472 combination).

[0306] SNF472 alone or in combination with cilostazol attenuated rat limb ischemia. In this model, cilostazol treatment alone was ineffective. The effects of SNF472 on blood perfusion persisted even 6 days after treatment cessation. See Fig. 11.

3. Effects on walking ability - treadmill running test

[0307] Maximal pain-free walking time (MWT) and maximal pain-free walking distance (MWD) were assessed in groups 1-4 using a treadmill running test (8 to 10 animals per group and time point) at D0, D5, D10, and D17.

[0308] Subjects were acclimated to the treadmill for two days prior to testing. On the first day, subjects were exercised for 5 to 10 minutes at a treadmill speed gradually increased from 15 m/min to 24 m/min. On the second day, subjects were first exercised at 15 m/min for 5 minutes. They were then exercised at 19.8 m/min (33 cm/sec) for an additional 5 minutes. Finally, subjects were exercised at 24 m/min (40 cm/sec) for a maximum of 30 additional minutes. Pre-intervention pain-free walking time and distance were obtained. Animals that did not meet protocol requirements were excluded from the test.

[0309] Limb function (MWT and MWD) was assessed in Groups 1, 2, and 3 using a treadmill running test 15 minutes after the respective daily administration. Limb function in Group 4 was assessed 3-4 hours after treatment.

[0310] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the treadmill test reading is taken.

[0311] Subjects continued running for 40 minutes or until exhaustion (i.e., they remained on the shock grid for five uninterrupted seconds). MWD and MWT were then calculated for each animal.

[0312] SNF472 and cilostazol improved the walking ability of rats compared to vehicle (+54% MWD and +46% MWT). See Fig. 13. Moreover, the effect of SNF472 on improving walking ability was maintained even 5 days after treatment cessation (D17). In contrast, cilostazol lost its beneficial effects immediately after treatment cessation. See Fig. 13.

4. Calcium content and calcification - ICP-OES

[0313] Subjects were anesthetized and bled on D24. Subjects were sacrificed after exsanguination. They were then necropsied and aortas were collected. Tissues were lyophilized for 24 h and weighed. Lyophilized tissues were then processed with 1:1 HNO3:HClO4 in a dry bath incubator for 2-4 h at 180°C. Processed tissues were then diluted with Milli-Q ultrapure water (MilliporeSigma (Merck KGaA), Burlington, MA, USA) to a final volume of 10 ml. Calcium content of tissue samples was quantified by ICP-OES.

[0314] SNF472 demonstrated efficacy against vascular calcification, as SNF472 inhibited aortic calcification (41±9%) following daily subcutaneous administration at a dose of 20 mg/kg compared to placebo. Cilostazol showed no activity against calcification at the same dose. See Fig. 14.

Example 3

Treatment of limb ischemia

[0315] The effects of SNF472 and cilostazol on blood perfusion, walking ability, and tissue calcification after the onset of limb ischemia were tested using a rat model for 13 days. Limb ischemia was induced in all groups from D1 to D3 except for the sham control group, in which ischemia was not induced. Subjects with induced ischemia were administered placebo and active agent formulations from D5 to account for the development of ischemia before treatment. Subjects were treated from D5 to D13 to assess the effects of treatment on limb ischemia, walking ability, and tissue calcification. Observations were made at multiple points during treatment from D1 to D13. All subjects were weighed daily before treatment.

1. Induction of limb ischemia

[0316] Sixty-six male Sprague Dawley (SD) rats (Envigo Corp., Huntingdon, UK) weighing approximately 250-275 g were used. Subjects were fed an AO4 diet (Scientific Animal Food &Engineering; Carpe Bio, Amersfoort, The Netherlands). Subjects were divided into 5 groups of 8-14 rats each as follows:

[0317] Group 1 - Control (control group with sham intervention)

[0318] Group 2 - D5 Initial Ca level

[0319] Group 3a Placebo - Physiological saline solution

[0320] Group 3b Placebo - 5% sodium CMC solution

[0321] Group 4 - SNF472 (Na ₆ IP ₆ )

[0322] Group 5 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ )

[0323] Limb ischemia was induced in subjects in Groups 2-5 by subcutaneous administration of 120,000 IU/kg _vitamin D ₃ (cholecalciferol, Duphafral D ₃ 1000; Zoetis Inc., Parsippany, NJ, USA) in saline (2 ml/kg) every day from D1 to D3. Subjects in Group 1 were administered saline (2 ml/kg) subcutaneously every day from D1 to D3 without vitamin D ₃ .

2. Relief of ischemia and effect on blood perfusion in the extremities - analysis using laser Doppler imaging

[0324] Subjects in Group 1 were administered 2 mL/kg of saline daily from D1 through D13 via the subcutaneous route. Subjects in Group 2 were administered 2 mL/kg of saline daily from D1 through D5 via the subcutaneous route. On D5, four subjects in Group 1 and all subjects in Group 2 were sacrificed to determine their baseline Ca values.

[0325] Subjects in Groups 3a and 3b were administered 2 mL/kg of normal saline daily from D5 to D13 via the subcutaneous route. Subjects in Group 3b and 3b were administered 5 mL/kg of 5% sodium CMC in aqueous solution orally daily from D5 to D13. In addition, subjects in Groups 4 were administered 40 mg/kg of SNF472 (Na ₆ IP ₆ , free base: 600 g/mol) in normal saline (2 mL/kg) daily from D5 to D13 via the subcutaneous route.

[0326] Subjects in Group 5 were orally administered 40 mg/kg cilostazol ( _C20H27N5O2 , free _base 369.46 g/mol; lot number _LRAB9590 , Sigma-Aldrich Corp., St. Louis, MO, _USA ) in 5% sodium CMC in aqueous solution (5 mL/kg) daily from D5 through D13. The 40 mg/kg cilostazol dose in rats is comparable to the therapeutic dose of 8.4 mg/kg in patients with PAD.

[0327] On days when blood perfusion tests were performed using active agents (i.e. D5, D13), dosages were administered as follows:

[0328] Group 4 - SNF472 (Na ₆ IP ₆ ): 15 minutes before reading

[0329] Group 5 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ): 3 - 4 hours before taking readings

[0330] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the test reading is taken.

[0331] Limb function and ischemic status were assessed using laser Doppler perfusion imaging in all groups at D0 and D5 and in groups 1, 3a, 3b, 4, and 5 at D13. The perfusion difference and perfusion ratio were calculated by comparing baseline and the readings at D5 and D13 for each group. Specifically, the perfusion difference and perfusion ratio were calculated by comparing the readings of group 1 (control) and: (a) groups 3a and 3b placebo (saline, 5% sodium CMC); (b) group 4 (SNF472); and (c) group 5 (cilostazol).

[0332] VitD3 administration caused a decrease in hindlimb blood perfusion in groups 3a, 3b, 4, and 5 (measured at D5 immediately prior to treatment administration). At D13, only animals receiving SNF472 showed a significant improvement in hindlimb blood perfusion compared to D5 prior to treatment. Animals receiving placebo or cilostazol at D13 did not show any improvement or resolution of ischemia compared to D5. See Fig. 16.

3. Effects on walking ability - treadmill running test

[0333] Maximal pain-free walking time (MWT) and maximal pain-free walking distance (MWD) were assessed in all groups at D0 and in groups 1, 3a, 3b, 4, and 5 at D6 and D11 using a treadmill running test (8 to 12 animals per group and time point). Prior to the test, subjects were acclimated to the treadmill for two days. On the first day, subjects were exercised for 5 to 10 minutes at a treadmill speed gradually increased from 15 m/min to 24 m/min. On the second day, subjects were initially exercised at 15 m/min for 5 minutes. They were then exercised at 19.8 m/min (33 cm/s) for an additional 5 minutes. Finally, subjects were exercised at a speed of 24 m/min (40 cm/s) for a maximum of 30 additional minutes. Preoperative pain-free walking time and distance were obtained. Animals that did not meet protocol requirements were excluded from the test.

[0334] Limb function (MWT and MWD) was assessed in Groups 1, 3a, 3b, and 4 using a treadmill running test 15 minutes after the respective daily administration. Limb function in Groups 5 was assessed 3-4 hours after treatment.

[0335] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the test reading is taken.

[0336] Subjects continued running for 40 minutes or until exhaustion (i.e., they remained on the shock grid for five continuous seconds). MWD and MWT were then calculated for each animal.

[0337] SNF472 improved the walking ability of rats compared to vehicle (+49% MWD) even when treatment was started 5 days after ischemia induction, whereas cilostazol was not effective under the same conditions and at a therapeutic dose (40 mg/kg/day). See Fig. 16.

4. Calcium content and calcification - ICP-OES

[0338] Four subjects from Group 1 and all subjects from Group 2 were sacrificed on D5 to determine their baseline Ca values. The remaining subjects from Group 1 and all subjects from Groups 3a, 3b, 4, 5, and 6 were sacrificed on D13.

[0339] Subjects were sacrificed after exsanguination. They were then autopsied and their right and left femoral arteries were collected. The tissues were lyophilized for 24 h and weighed. The lyophilized tissues were then processed using a 1:1 mixture of HNO ₃ :HClO ₄ in a dry bath incubator for 2-4 h at 180°C. The processed tissues were then diluted with Milli-Q ultrapure water (MilliporeSigma (Merck KGaA), Burlington, MA, USA) to a final volume of 10 ml. The calcium content of the tissue samples was quantified by ICP-OES.

[0340] SNF472 demonstrated efficacy against vascular calcification at D13. Compared to placebo, SNF472 inhibited femoral artery calcification by approximately (30%) after daily subcutaneous administration at a dose of 40 mg/kg. See Fig. 17.

5. Pharmacokinetics

[0341] Subjects in groups 1, 3a, 3b, 4, and 5 were anesthetized and had their blood drawn on D13. Approximately 8-10 mL of total blood was collected from each subject and divided into plasma (K3EDTA, approximately 6 mL of blood) and serum (approximately 2-3 mL of blood) collection tubes. Plasma was stored in one 600 µL aliquot and several other 500 µL aliquots. Serum was divided into two aliquots.

[0342] Plasma myo-inositol hexaphosphate levels in subjects in Cohort 4 (i.e., measured at approximately C _max , 15 minutes after the last dose of SNF472) were quantified using LC-MS/MS chromatographic methods described in the art. See WO2013050603.

[0343] All subjects in Group 4 were adequately exposed to SNF472. Plasma levels 15 minutes after the last subcutaneous administration at D13 were 40,078±15,024 ng/mL (60.7±22.8 μM). Plasma levels in rats following daily administration of 20 mg/kg were comparable to levels found in hemodialysis patients receiving SNF472 at 8.4 mg/kg intravenously.

Example 4

Drug interactions SN472

[0344] The compatibility of SNF472 with other drugs regularly administered to subjects with renal impairment was analyzed.

[0345] Wistar rats (Charles R ₁ ver Labs, Inc., Wilmington, MA, USA) were treated with the following: (i) SNF472 administered subcutaneously (s.c.), (ii) SNF472 + sevelamer (po), (iii) SNF472 (s.c.) + cinacalcet (po), (iv) SNF472 (s.c.) + Vit D (s.c.), (v) SNF472 (s.c.) + sodium thiosulfate (s.c.), and SNF472 (s.c.) + ibandronate (s.c.). No significant differences were observed between administration of SNF472 alone or co-administered with any other study drugs.

Example 5

Prevention of limb ischemia

Dose-dependent effects of SNF472 on maximal walking time (MWT) and distance (MDT)

[0346] The prophylactic effects of several different doses of SNF472 and the highest tolerated dose of cilostazol on blood perfusion, walking ability, and tissue calcification were tested over 12 days using a rat model. Limb ischemia was induced in subjects at D1 through D3, excluding the sham control group. Animals with induced ischemia were then treated with placebo and active agent formulations at D1 through D12 to assess their effects on preventing (a) limb ischemia and tissue calcification and (b) walking ability impairment. Observations were made at multiple time points during the D1 through D12 treatment phase. All subjects were weighed daily prior to treatment.

1. Induction of limb ischemia

[0347] One hundred and two male Sprague Dawley (SD) rats (Envigo Corp., Huntingdon, UK) weighing approximately 250-275 g were used. Subjects were fed an AO4 diet (Scientific Animal Food &Engineering; Carpe Bio, Amersfoort, The Netherlands). Subjects were divided into 5 groups, 8 to 12 animals per group, as follows:

[0348] Group 1 - Control (control group with sham intervention);

[0349] Group 2a Placebo - Physiological saline solution, subcutaneous;

[0350] Group 2b Placebo - 5% sodium CMC solution, orally;

[0351] Group 3 - SNF472 (Na ₆ IP ₆ ) at a dose of 1 mg/kg, subcutaneously;

[0352] Group 4 - SNF472 (Na ₆ IP ₆ ) at a dose of 7.5 mg/kg, subcutaneously;

[0353] Group 5 - SNF472 (Na ₆ IP ₆ ) at a dose of 15 mg/kg, subcutaneously;

[0354] Group 6 - SNF472 (Na ₆ IP ₆ ) at a dose of 30 mg/kg, subcutaneously;

[0355] Group 7 - SNF472 (Na ₆ IP ₆ ) at a dose of 45 mg/kg, subcutaneously;

[0356] Group 8 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ) at a dose of 45 mg/kg orally.

[0357] Limb ischemia was induced in subjects in Groups 2-8 by subcutaneous administration of 120,000 IU/kg vitamin D ₃ (cholecalciferol, Duphafral D ₃ 1000; Zoetis Inc., Parsippany, NJ, USA) in saline (2 ml/kg) every day from D1 to D3. Subjects in _the sham control Group 1 were administered saline (2 ml/kg) subcutaneously every day from D1 to D3 without vitamin D ₃ .

[0358] Subjects in Groups 1 and 2a were administered physiological saline (2 mL/kg) subcutaneously every day from D1 to D12. Subjects in Group 2b were administered 5% sodium CMC in aqueous solution (5 mL/kg) orally every day from D1 to D12.

2. Relief of ischemia and effect on blood perfusion in the extremities - analysis using laser Doppler imaging

[0359] Subjects in Groups 1 and 2a were administered 2 mL/kg of physiological saline daily from D1 to D12 via the subcutaneous route. Subjects in Group 2b were orally administered 5 mL/kg of 5% sodium salt of CMC in aqueous solution every day from D1 to D12. In addition, subjects in Groups 3, 4, 5, 6, and 7 were administered 1 mg/kg, 7.5 mg/kg, 15 mg/kg, 30 mg/kg, and 45 mg/kg of SNF472 (Na ₆ IP ₆ , free base: 600 g/mol), sodium salt (free base 696.27 g/mol), respectively. SNF472 was administered in physiological saline (2 mL/kg) every day from D1 to D12 via the subcutaneous route. Subjects in Group 8 were orally administered 45 mg/kg _cilostazol ( _C20H27N5O2 , _free base 369.46 g/ _mol ; lot number LRAB9590, Sigma-Aldrich Corp., St. Louis, MO, USA) in 5% sodium CMC in aqueous solution (5 mL/kg) daily from D1 to D12.

[0360] On days when blood perfusion tests were performed using active agents (i.e., D6 and D12), dosages were administered as follows:

[0361] Groups 3, 4, 5, 6 and 7 - SNF472 (Na ₆ IP ₆ ): 15 minutes before reading;

[0362] Group 8 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ): 3-4 hours before taking readings.

[0363] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the test reading is taken.

[0364] Limb ischemia status was assessed during the treatment period (i.e., at D0, D6, and D12) using laser Doppler perfusion imaging in all rats. Perfusion difference and perfusion ratio were calculated by comparing baseline and either D6 or D12 readings for each group. Specifically, perfusion difference and perfusion ratio were calculated by comparing readings from Group 1 (control) and (a) Groups 2a and 2b placebo (saline, 5% sodium CMC), (b) Groups 3, 4, 5, 6, and 7 (SNF472), and (c) Group 8 (cilostazol).

[0365] SNF472 attenuated rat limb ischemia in a dose-dependent manner. Treatment with cilostazol alone was not effective in this model.

3. Effect on walking ability - treadmill running test

[0366] Maximal pain-free walking time (MWT) and maximal pain-free walking distance (MWD) were assessed in groups 1-8 using a treadmill running test (8-12 animals per group and time point) at D0, D5, and D10.

[0367] Subjects were acclimated to the treadmill for two days prior to testing. On the first day, subjects were exercised for 5 to 10 minutes at a treadmill speed gradually increased from 15 m/min to 24 m/min. On the second day, subjects were initially exercised at 15 m/min for 5 minutes. They were then exercised at 19.8 m/min (33 cm/s) for an additional 5 minutes. Finally, subjects were exercised at 24 m/min (40 cm/s) for a maximum of 30 additional minutes. Pre-operative pain-free walking time and distance were obtained. Animals that did not meet protocol requirements were excluded from the test.

[0368] Limb function (MWT and MWD) was assessed in Groups 1, 2, 3, 4, 5, 6, and 7 using a treadmill running test 15 minutes after the respective daily administration. Limb function in Group 8 was assessed 3-4 hours after treatment.

[0369] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the treadmill test reading is taken.

[0370] Subjects continued running for 40 minutes or until exhaustion (i.e., they remained on the shock grid for five continuous seconds). MWD and MWT were then calculated for each animal.

[0371] SNF472 and cilostazol improved the walking ability of rats compared to vehicle. Moreover, the effect of SNF472 on improving walking ability was dose dependent.

4. Calcium content and calcification - ICP-OES

[0372] Subjects were anesthetized and bled on D12. Subjects were sacrificed after exsanguination. They were then autopsied and cardiac and aortic arteries were collected. Tissues were lyophilized for 24 h and weighed. Lyophilized tissues were then processed with a 1:1 mixture _of HNO3: _HClO4 in a dry bath incubator for 2-4 h at 180°C. Processed tissues were then diluted with Milli-Q ultrapure water (MilliporeSigma (Merck KGaA), Burlington, MA, USA) to a final volume of 10 ml. Calcium content of tissue samples was quantified by ICP-OES.

[0373] SNF472 has been shown to be effective against cardiac and vascular calcification, such as cardiac arterial and aortic calcification, in a dose-dependent manner. Cilostazol was not active against calcification at the same dose.

Example 6

Prevention of limb ischemia in a rat model of adenine-induced uremia

Effect of SNF472 on blood perfusion in the extremities

[0374] The prophylactic effects of SNF472 and cilostazol doses on blood perfusion and tissue calcification were tested using a rat model of chronic kidney disease for 21 days.

[0375] Uremia and limb ischemia were induced in animals from D1 to D21, except for the sham-intervention control group, in which neither uremia nor ischemia was induced. Animals with induced ischemia were then treated with placebo and active agent formulations from D1 to D21 to assess their effects on the prevention of (a) limb ischemia and (b) tissue calcification. Observations were made at multiple points during the treatment phase from D1 to D21. All subjects were weighed each day before treatment.

1. Induction of uremia and limb ischemia

[0376] Sixty-eight male Sprague Dawley (SD) rats (Envigo Corp., Huntingdon, UK) weighing approximately 250-275 g were used and were fed a high phosphorus pelleted diet (SM R, 10 mm pellets, 1.06% Ca, 1.03% P) (SSNIFF Spezialdiaeten, Soest, Germany) ad libitum. Subjects were divided into 6 groups, 8 to 12 animals in each group, as follows:

[0377] Group 1 - Control (control group with sham intervention)

[0378] Group 2a Placebo - Physiological saline, subcutaneously, once daily;

[0379] Group 2b Placebo - Physiological saline, subcutaneous, Alzet pump, 4 weeks;

[0380] Group 3 - SNF472 (Na ₆ IP ₆ ) at a dose of 30 mg/kg subcutaneously once daily;

[0381] Group 4 - SNF472 (Na ₆ IP ₆ ) at a dose of 45 mg/kg, subcutaneous, once daily;

[0382] Group 5 - SNF472 (Na ₆ IP ₆ ) at a total dose of 400 mg/4 weeks (100 mg/week), subcutaneously, Alzet pump implanted at D1 for 4 weeks prior to adenine administration;

[0383] Group 6 Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ) at a dose of 45 mg/kg/day, orally, once daily.

[0384] Uremia and limb ischemia were induced in groups 2–6 with a daily dose of adenine (500 mg/kg suspended in 1% carboxymethylcellulose, administered orally) for the first 10 days, followed by a dose of α-calcidol (100 ng/kg in olive oil, administered orally) three times a week from D11 to D19 to accelerate and ensure uniformity of the development of cardiovascular calcification and ischemia.

[0385] On day 21, animals were sacrificed and blood and tissue samples were collected. Serum creatinine (#OSR6178) and urea (#OSR6134) were determined using the appropriate Beckman Coulter assay kit (Beckman Coulter, Inc., Brea, CA, USA).

[0386] Animals in the sham-intervention control group 1 were given 1% carboxymethylcellulose solution (5 ml/kg) orally every day from D1 to D10, and then olive oil was given orally three times a week from D11 to D19. Neither uremia nor ischemia was induced in the sham-intervention control group.

2. Relief of ischemia and effect on blood perfusion in the extremities - analysis using laser Doppler imaging

[0387] Subjects in Groups 2a were administered saline (2 mL/kg) subcutaneously twice daily for 21 days. Subjects in Group 2b were administered saline subcutaneously using an Alzet pump after 4 weeks.

[0388] In addition, animals in Groups 3 and 4 were administered subcutaneously twice daily 30 mg/kg and 45 mg/kg SNF472 (Na ₆ IP ₆ , free base: 600 g/mol), sodium salt (free base 696.27 g/mol), respectively. SNF472 was administered in physiological saline (2 ml/kg) twice daily from D1 to D21 via the subcutaneous route. Animals in Group 5 were administered SNF472 at a dose of 400 mg/week dissolved in physiological saline via an Alzet subcutaneous pump implanted for 4 weeks.

_[ 0389] Animals in Group 6 were orally administered 45 mg/kg cilostazol ( _C20H27N5O2 , free base 369.46 g/ _mol ; lot number LRAB9590, Sigma-Aldrich Corp., St. Louis, MO, _USA ) in 5% sodium CMC in aqueous solution (5 mL/kg) daily from D1 to D21.

[0390] On days when blood perfusion tests were performed using active agents (i.e., D10, D17, and D21), dosages were administered as follows:

[0391] Groups 3 and 4 - SNF472 (Na ₆ IP ₆ ): 15 minutes before reading;

[0392] Group 6 - Cilostazol (C ₂₀ H ₂₇ N ₅ O ₂ ): 3-4 hours before taking readings.

[0393] According to the above scheme, the active agents will have a maximum plasma concentration (C _max ) at the time the test reading is taken.

[0394] Limb ischemia status was assessed during treatment (i.e., D0, D10, and D17 and D21) in all rats using laser Doppler perfusion imaging. Perfusion difference and perfusion ratio were calculated by comparing baseline and D10, D17, and D21 readings for each group. Specifically, perfusion difference and perfusion ratio were calculated by comparing readings from Group 1 (control) and (a) placebo groups 2a and 2b, (b) groups 3, 4, and 5 (SNF472), and (c) group 6 (cilostazol).

[0395] SNF472 attenuated rat limb ischemia in a dose-dependent manner in uremic rats. Treatment with cilostazol alone was not effective in this model.

3. Calcium content and calcification - ICP-OES

[0396] Subjects were anesthetized and bled on D21. Subjects were sacrificed after exsanguination. They were then autopsied and cardiac and aortic arteries were collected. Tissues were lyophilized for 24 h and weighed. Lyophilized tissues were then processed with a 1:1 mixture of _HNO3 : _HClO4 in a dry bath incubator for 2-4 h at 180°C. Processed tissues were then diluted with Milli-Q ultrapure water (MilliporeSigma (Merck KGaA), Burlington, MA, USA) to a final volume of 10 ml. Calcium content of tissue samples was quantified by ICP-OES.

[0397] SNF472 has been shown to be effective against cardiac and vascular calcification in uremic rats, such as cardiac and aortic arterial calcification, in a dose-dependent manner following daily subcutaneous dosing and/or delivery via the Alzet pump compared to placebo. Cilostazol was not active against calcification at the same dose.

Claims (12)

1. The use of hexasodium salt of myo-inositol hexaphosphate or a pharmaceutical composition containing said salt together with pharmaceutically acceptable excipients and carriers for the treatment or prevention of peripheral arterial disease. 2. Use according to item 1, intended for the treatment or prevention of critical limb ischemia. 3. The use according to claim 1, wherein the pharmaceutical composition is an intravenous aqueous composition of hexasodium salt of myo-inositol hexaphosphate (SNF472). 4. The use according to any one of claims 1-3, wherein the compound or composition is administered to a subject with renal insufficiency. 5. The use according to any one of claims 1-4, wherein the compound or composition is administered to the subject during dialysis. 6. The use according to claim 5, wherein the compound or composition is administered to the subject during hemodialysis. 7. The use according to claim 6, wherein the compound or composition is administered into unfiltered blood taken from the subject. 8. Use according to claim 1, wherein the compound or composition is administered parenterally. 9. Use according to paragraph 8, wherein parenteral administration is intravenous, subcutaneous or intramuscular. 10. The use according to paragraph 1 for improving the ability to walk in a subject in need thereof. 11. The use of claim 1 for increasing the maximum pain-free walking distance (MWD), maximum pain-free walking time (MWT), or both in a subject in need thereof. 12. The use according to any one of claims 1-11, wherein the compound is administered to the subject at an effective dosage of about 0.001 to about 60 mg/kg.