Composition for improving hemoglobin-mediated oxygen delivery comprising alpha-ketoglutaric acid (alpha-KG) and 5-hydroxymethyl-2-furfural (5-HMF)

Abstract

The present invention relates to a pharmaceutical composition comprising α-ketoglutarate (α-KG) and 5-hydroxymethyl-2-furfural (5-HFM) for use in the treatment and / or prevention of diseases or conditions that benefit from increased hemoglobin-mediated oxygen delivery.

Description

[Technical field]

[0001] The present invention relates to a pharmaceutical composition comprising α-ketoglutarate (α-KG) and 5-hydroxymethyl-2-furfural (5-HMF) for the treatment and / or prevention of diseases or conditions that benefit from enhanced hemoglobin-mediated oxygen delivery. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0002] This description refers to several documents, including patent applications, patents, and instructions from various manufacturers. The disclosure of each of these documents is incorporated into the disclosure of this application, although it is not considered relevant to the patentability of this invention. In particular, all documents referenced in these documents are also included in the disclosure of this application to the same extent as if each of these documents was specifically incorporated by reference into the disclosure of this application.

[0003] Red blood cells are blood cells whose main function is to transport vital oxygen inhaled in the lungs through the blood vessels to the organs and tissues of the body. Red blood cells do this thanks to the red blood pigment they contain, hemoglobin (Hb).

[0004] Hemoglobin is a tetrameric protein that can reversibly bind four oxygen molecules through an allosteric mechanism. In blood, hemoglobin exists in balance between two allosteric structural states. In the "T" state (tension), hemoglobin is deoxygenated. In the "R" state (relaxation), hemoglobin is oxygenated.

[0005] The oxygen dissociation curve (ODC, or "oxyhemoglobin dissociation curve"), also called the oxygen binding curve of hemoglobin (Hb), is a graphical representation of the relationship between the oxygen partial pressure (PO2) and oxygen saturation (SO) of Hb, where SO2 corresponds to the percentage of oxygen-saturated Hb (oxyhemoglobin) in total hemoglobin. The typical sigmoidal shape of the ODC reflects the cooperativity of the reversible binding of four oxygen molecules to a tetrameric hemoglobin molecule. This means that the binding of one oxygen molecule induces a cooperative conformational change in Hb, changing it from a so-called "tense" state to a "relaxed" state, making the hemoglobin molecule more amenable to binding oxygen. The ODC assigns a specific oxygen saturation of hemoglobin to each oxygen partial pressure (PO2). A lower PO2 results in a lower saturation. In the central region of the curve, an increase in PO2 leads to an almost linear increase in oxygen saturation. At a certain PO2, Hb is saturated. Further increases in oxygen tension do not result in any significant further increase in oxygen saturation.

[0006] The most important parameters describing the ODC are the P50 value (the partial pressure of oxygen (PO2) at which 50% saturation of Hb with oxygen (O2) occurs, a measure of the O2 affinity of Hb) and the Hill coefficient, which represents the maximum slope of the ODC and is therefore a measure of the cooperativity of oxygen binding to Hb (2).

[0007] The course of the ODC curve is influenced by a number of factors (3), resulting in an increase in HbO2 affinity and an associated shift in ODC to the left, or a decrease in HbO2 affinity and an associated shift in ODC to the right (4); in general, a rightward shift favors oxygen release, and a leftward shift favors oxygen loading.

[0008] Known physiological factors that cause a rightward shift of the ODC are, for example, increased carbon dioxide concentration, low (acidic) pH, increased 2,3-diphosphoglycerate (2,3-DPG, also known as 2,3-biphosphoglycerate (2,3-BPG)) concentration, and / or elevated body temperature. This ultimately contributes to improved oxygen supply to metabolically active tissues, where oxygen and glucose are metabolized to carbon dioxide and organic acids.

[0009] Various diseases and conditions associated with insufficient supply of oxygen are known, and these diseases or conditions are usually due to either impaired oxygen uptake in the lungs or impaired supply of oxygenated blood. Apart from (patho)physiological causes, external factors such as low air pressure at high altitudes or low oxygen content in the air can cause insufficient oxygen supply and associated health complications. It is also known that an efficient oxygen supply is essential to maintain all physiological functions and hence physical and mental performance.

[0010] Influencing the allosteric balance of hemoglobin, i.e., its oxygen binding and releasing properties, is therefore a viable means for the treatment and / or prevention of various such diseases or conditions, as well as for improving physical and / or mental performance during exercise and work.

[0011] It has been suggested that shifting hemoglobin to a state of lower oxygen affinity, and the associated rightward shift in ODC, may be particularly beneficial in diseases and conditions in which tissues are under-oxygenated. It is believed that oxygen loading of Hb in the lungs is hindered by the reduced oxygen affinity, whereas oxygen release in peripheral tissues is generally promoted.

[0012] In contrast, affecting hemoglobin by stabilizing the R state toward a state of high oxygen affinity, and the associated leftward shift in ODC, is thought to be particularly advantageous for the treatment of sickle cell anemia. It has been shown that stabilizing the R state of sickle cell hemoglobin (Hb-S), which underlies this genetic disease, can inhibit red blood cell aggregation and the associated vaso-occlusion (sickling) associated with the polymerization-prone T state.

[0013] On the one hand, it has been argued in the past that substances that cause an increase in oxygen affinity and a concomitant leftward shift in ODC lead to improved oxygen loading but do not contribute to improved oxygen delivery in peripheral tissues, since oxygen release becomes more difficult. However, studies have revealed that the efficiency of oxygen release is mainly determined by the ODC curve. It has been shown that substances that, on the one hand, increase the oxygen affinity of HBs (i.e., cause a leftward shift in ODC) but, on the other hand, maintain the shape of the sigmoid ODC curve, contribute to improved overall oxygen delivery.

[0014] For example, the micronutrient 5-hydroxymethyl-2-furfural (5-HMF) was identified as an allosteric modulator of Hb that enhances its affinity for oxygen. In an animal model of severe hypoxia, it was detected that the enhanced HbO2 affinity by 5-HMF could protect against hemodynamic instability and maintain microvascular oxygenation (8). Another study showed that the enhanced HbO2 affinity induced by 5-HMF improved arterial oxygen saturation (SO2) and attenuated the increase in pulmonary artery pressure associated with hypoxia in pigs exposed to hypoxia (9). Similarly, the increased HbO2 affinity induced by 5-HMF was detected in human subjects and sickle cell anemia patients exposed to hypoxia (10).

[0015] Especially since the emergence and global spread of COVID-19 and its associated respiratory complications, there is a constant need for new, alternative and improved therapeutics for the treatment and / or prevention of diseases and conditions that would benefit from improved hemoglobin-mediated oxygen delivery. There is also an increasing demand for nutritional supplements to optimize and improve mental and physical performance during exercise and at work.

[0016] These problems are solved by the present invention according to the claims and the following specification and examples.

[0017] In a first aspect, the present invention relates to a pharmaceutical composition comprising α-ketoglutarate (α-KG) and 5-hydroxymethyl-2-furfural (5-HMF) for use in the treatment and / or prevention of diseases or conditions that benefit from enhanced hemoglobin-mediated oxygen delivery.

[0018] 5-Hydroxymethyl-2-furfural (5-HMF), also known as hydroxymethylfurfural (HMF), 5-(hydroxymethyl)furfural or 5-oxymethylfurfurol, is an aldehyde and furan compound that is formed, for example, in the thermal decomposition of sugars or carbohydrates and can be found in many foods, such as fruit juices or honey.

[0019] Alpha-ketoglutaric acid (α-KG), also known as 2-oxoglutaric acid or 2-oxopentanedioic acid, is a dicarboxylic acid derived from n-pentane with an additional carbonyl group at the α-C atom. Alpha-KG forms colorless and nearly odorless crystals. Alpha-KG is the naturally occurring nitrogen-free moiety of the amino acids glutamine and glutamic acid. Alpha-KG exists as an anion (α-ketoglutarate) in the aqueous environment of cells and is an intermediate in energy metabolism during cellular ATP production via the citric acid cycle. Alpha-KG is a more potent radical scavenger (RONS) than an equivalent dose of vitamin C and acts as a nitrogen regulator in metabolism. For the purposes of this disclosure, the terms Alpha-ketoglutaric acid (α-KG) and Alpha-ketoglutaric acid are used interchangeably.

[0020] In the context of the present invention, it has been surprisingly found that the combination of 5-HMF and α-KG leads to the enhancement of the oxygen affinity of hemoglobin while maintaining the sigmoid ODC curve shape.As shown in the examples, the measured effect of the combination of 5-HMF and α-KG exceeds the effect of each of the substances individually, from which it can be concluded that the synergistic effect of the combination of 5-HMF and α-KG is related to the allosteric regulation of hemoglobin.This synergistic effect is the basis of the present invention.

[0021] The combination of 5-HMF and α-KG itself is already known and is commercially available as a dietary supplement (Sanopal®). For example, in a previous study by the present inventors, it was reported that administration of a combination of 5-HMF and α-KG in healthy subjects who exercised by cycling at a simulated altitude of 3500 m (FiO2 = 13.5%) caused an increase in peripheral oxygen saturation (SpO2) (12). In another study, it was reported that administration of a combination of 5-HMF and α-KG as preoperative oral micronutrient supplementation to lung cancer patients caused an improvement in maximum oxygen uptake (VO2max) and a reduction in postoperative (lobectomy with one-lung ventilation) hospital stay. In both studies, a possible cause underlying the observed effects was the known properties of the combination of 5-HMF and α-KG as a means of reducing oxidative stress, particularly tissue damage caused by oxidative stress. However, as first disclosed in connection with the present invention, the effect of the combination on the oxygen-binding properties of hemoglobin was completely unknown.

[0022] The term "enhancement of hemoglobin-mediated oxygen delivery" as used herein means that the combination of 5-HMF and α-KG causes a leftward shift in the hemoglobin oxygen saturation curve (ODC), which can be measured in the absence of the combination, compared to a reference (e.g., a reference blood sample). Various methods for measuring ODC are routinely known to those skilled in the art. For example, the ODC disclosed in the experimental section of this specification was determined using the in vitro method described in Woyke et al. (Ref:20).

[0023] Furthermore, it should be appreciated that preferably the leftward shift in ODC occurs while the shape of the sigmoid curve is maintained.

[0024] As used herein, the term "diseases that would benefit from enhanced hemoglobin-mediated oxygen delivery" includes all such diseases, disease symptoms and / or disorders that are characterized and / or caused by inadequate oxygen supply and / or that would benefit from improved hemoglobin-mediated oxygen delivery, for alleviation or cure.

[0025] As used herein, the term "conditions that would benefit from enhanced hemoglobin-mediated oxygen delivery" includes all of those conditions or disorders that are characterized and / or caused by a lack of oxygen supply and / or that, if left untreated, may lead to disease or pathological changes prior to the detectable occurrence of disease symptoms or pathological changes.

[0026] In any case, the term a disease or condition "benefits" from treatment and / or prevention is to be interpreted in the sense that the disease or condition is alleviated by the treatment, or the risk of developing such disease or condition is reduced or eliminated by prevention.

[0027] While the therapeutic / prophylactic uses disclosed herein are primarily intended for human (i.e., individual / patient) applications, the disclosure also includes veterinary applications, i.e., for animals, particularly horses, dogs, livestock (e.g., cows, pigs, goats, poultry).

[0028] It is understood that the pharmaceutical composition according to the present invention may have a pharma- ceutically acceptable formulation. Pharmaceutically acceptable formulations are well known in the art. For example, see Rowe et al. Handbook of Pharmaceutical Excipients (6 th See document 1999, 2002 edition, RC Rowe, PJ Sheskey, ME Quinn. Pharmaceutical Press, London, 2009.

[0029] The pharmaceutical compositions according to the invention may further comprise additives. These include any compound or composition advantageous for use according to the invention, including water, salts, binders, solvents, dispersants, buffers (particularly physiological buffers, such as Ringer's solution or phosphate buffered saline (PBS)), stabilizers, and other substances commonly used in connection with the formulation of drugs. The pharmaceutical compositions may also include preservatives and other additives, such as antimicrobial substances, antioxidants, complexing agents, and inert gases.

[0030] Particularly preferred additives are water, sweeteners (e.g. sugars, preferably sucrose, glucose or dextrose, or sugar substitutes), magnesium chloride, and / or acidity regulators, such as potassium hydroxide and / or sodium hydroxide. Trace elements, vitamins, amino acids and / or plant extracts can also be used as additives. Further particularly preferred additives are all substances that are required or beneficial for blood formation, such as, for example, vitamin B12 (cobalamin), vitamin B9 (folic acid) and / or vitamin B6, the trace elements iron and / or selenium, and the amino acid methionine.

[0031] According to a preferred embodiment, the composition according to the invention comprises α-KG and 5-HMF as the only combination of active ingredients (i.e., no other pharma- ceutical active ingredients). However, according to another embodiment, the pharmaceutical composition may comprise further active ingredients, particularly those that may be beneficial and advantageous for the respective intended use. Examples include anticoagulants (e.g., heparin), antihypertensives, antibiotics, viriostatics, anti-inflammatory substances (e.g., glucocorticoids), chemotherapeutic agents, immunosuppressants (e.g., cyclosporine or tacrolimus), proteins and / or peptides (including peptide hormones such as angiotensin).

[0032] The administration of the pharmaceutical composition according to the present invention is not limited to a particular method (mode of administration / form of dosage), and may be oral or parenteral, for example, intravenous, intraperitoneal, subcutaneous, intramuscular, topical, intranasal, intrabronchial, oral or intradermal, or any other well-known mode of administration. The administration may further be carried out in a suitable dosage form, for example, tablet, capsule, powder, granule, suppository, injection, cream or aerosol. The suitable route of administration and the dosage form and interval may be specifically selected depending on the disease or condition to be treated. Those skilled in the art recognize that the type of dosage depends on various factors, such as the height or weight, body surface area, age, sex and / or general health of the patient, but also on the specific drug to be administered, the duration and type of administration, and other medicines that may be administered simultaneously.

[0033] In a preferred embodiment, the composition according to the invention, as well as other additives or other active ingredients as defined above (e.g. heparin), are contained in and administered together with stored blood (i.e. whole blood, serum or plasma).

[0034] The preferred doses, routes and forms, as well as the preferred administration intervals, are specifically defined below.

[0035] In a second aspect, the present invention relates to the use of a composition comprising α-KG and 5-HMF as a dietary supplement for enhancing hemoglobin-mediated oxygen delivery.

[0036] The term "dietary supplement", also called "supplement", is well known and includes preparations that can be added to human or animal nutrition. The term therefore also expressly includes dietary supplements for use in animal feed.

[0037] It will be understood that the use of the composition according to the invention comprising alpha-KG and 5-HMF as a dietary supplement according to the second aspect of the invention described herein includes exclusively non-medical (i.e. non-therapeutic) uses. Thus, the composition may also contain further additives beneficial for the intended use as described for the pharmaceutical composition according to the first aspect of the invention, excluding the medicament active ingredient.

[0038] It is well known that cells require oxygen to produce adenosine triphosphate (ATP), the metabolic energy source for all bodily functions. In the event of an insufficient oxygen supply, ATP production may cease, leading to fatigue and the associated loss or decline in performance. Increased physical and / or emotional / mental stress increases the need for ATP and therefore the oxygen required for ATP production.

[0039] Therefore, the inventors do not wish to be bound by theory, but believe that the provision of the nutritional combination of 5-HMF and α-KG by the use of the present invention leads to an increase in hemoglobin-mediated oxygen supply in the organism, which can contribute to maintaining and / or enhancing ATP production, and thus improving energy supply.The use of the composition according to the present invention, which comprises α-KG and 5-HMF as a dietary supplement, is considered to be particularly beneficial in preventing and / or reducing the loss of performance caused by physical and / or mental stress.Therefore, the use according to the present invention is considered to be useful for improving physical and / or mental performance.

[0040] A preferred embodiment according to the second aspect of the present invention relates to the use of the composition as a dietary supplement to improve physical and / or mental performance during sports activities, for example in competitive or recreational sports, or during mental stress, for example during study or work.The use to increase aerobic and / or endurance performance is particularly preferred.Possible applications expressly contemplated by the present invention are the use of the composition as a dietary supplement in endurance sports, especially long-distance sports or competitions, such as marathons, cycling, swimming, cross-country skiing, and combined sports, such as duathlons, triathlons, biathlons, etc.The use of the composition as a dietary supplement for animal, especially horse or dog competitions, is also contemplated.

[0041] A further preferred embodiment according to the second aspect of the invention relates to the use of the composition for reducing, delaying or preventing hypoxia-related performance impairment.

[0042] The term "hypoxia", also known as "oxygen deficiency" or "low oxygen", refers to an insufficient supply of oxygen affecting the entire body or a portion thereof. Arterial hypoxia is defined as a reduction in the partial pressure of oxygen in arterial blood and can be indirectly measured by a reduction in the oxygen saturation of arterial blood ("hypoxemia") using known methods (e.g., pulse oximetry).

[0043] Hypoxic conditions, i.e. conditions resulting in reduced performance associated with hypoxia, can be caused or promoted by a variety of known factors, for example, hypoxic conditions can occur during intense sporting activities, especially when the amount of oxygen taken in by breathing is not sufficient to cover the increased oxygen demand caused by physical activity.

[0044] Other possible causes of hypoxia are low oxygen content in the surrounding air available for breathing, due to air pollution, for example from exhaust fumes (smog) or fires, and low air pressure at high altitudes (e.g. when climbing), which results in insufficient oxygen loading of the haemoglobin in the lungs.

[0045] Thus, a particularly preferred embodiment according to the second aspect of the invention comprises (i) to reduce, delay or prevent performance impairment associated with hypoxia at altitude; and / or (ii) High altitude; preferably for improving physical and / or mental performance at altitudes above 2000m, more preferably above 2500m, even more preferably above 3000m. The present invention relates to the use of the composition as a dietary supplement.

[0046] It is to be understood that altitudes mentioned herein may preferably refer to altitude in metres (m) from the datum or height in metres (m) from sea level (SL).

[0047] In a preferred embodiment according to the first or second aspect of the present invention, α-KG and 5-HMF are in a mass ratio of 1:1 to 100:1, preferably 2:1 to 50:1, more preferably 2.25:1 to 25:1 in the (pharmaceutical) composition.

[0048] In the experiments disclosed herein, α-KG and 5-HMF were used in a mass ratio of 3:1. It should therefore be understood that mass ratios of α-KG to 5-HMF in the range of 2.5:1 to 15:1 are particularly preferred, with the range of 2.75:1 to 10:1 being even more preferred, and the range of 3:1 to 5:1 being most preferred. However, it is believed that the synergistic effect observed with the combination of α-KG and 5-HMF can be achieved to a similar (albeit possibly slightly weaker) degree with other mass ratios.

[0049] In a further preferred embodiment according to the first or second aspect of the present invention, the composition is administered at a dose of 5-500 mg / kg body weight of α-KG and 5-160 mg / kg body weight of 5-HMF, preferably at a dose of 25-450 mg / kg body weight of α-KG and 8-150 mg / kg body weight of 5-HMF, more preferably at a dose of 30-430 mg / kg body weight of α-KG and 10-145 mg / kg body weight of 5-HMF.

[0050] In another preferred embodiment according to the first or second aspect of the invention, the composition is administered at a dose of 30-420 mg / kg body weight of α-KG and 10-140 mg / kg body weight of 5-HMF, more preferably at a dose of 36-300 mg / kg body weight of α-KG and 12-100 mg / kg body weight of 5-HMF, even more preferably at a dose of 36-130 mg / kg body weight of α-KG and 12-43 mg / kg body weight of 5-HMF.

[0051] It is understood that the preferred doses described may alternatively be expressed as mg / ml blood volume, assuming a blood volume of approximately 5000 ml for a 70 kg body weight. For example, a dose of 5 mg 5-HMF per kg body weight may alternatively be expressed as a dose of 0.07 mg 5-HMF per ml of blood volume.

[0052] In the experiments disclosed herein, three different doses of α-KG and 5-HMF were tested, either as individual agents or in combination.

[0053] A "low dose" is 0.42 mg of α-KG and / or 0.14 mg of 5-HMF per ml of whole blood sample (corresponding to about 30 mg of α-KG or about 10 mg of 5-HMF per kg of body weight, assuming an average blood volume of 5000 ml per 70 kg of body weight); a "medium dose" is 1.8 mg of α-KG and / or 0.6 mg of 5-HMF per ml of whole blood sample (corresponding to about 129 mg of α-KG or about 43 mg of 5-HMF per kg of body weight, assuming an average blood volume of 5000 ml per 70 kg of body weight); and a "high dose" is 6 mg of α-KG and / or 2 mg of 5-HMF per ml of whole blood sample (corresponding to about 429 mg of α-KG or about 143 mg of 5-HMF per kg of body weight, assuming an average blood volume of 5000 ml per 70 kg of body weight).

[0054] For all three doses tested of the combination of α-KG and 5-HMF, and for 5-HMF alone, the experiments disclosed herein show a significant decrease in P50 (i.e., the oxygen partial pressure at which 50% of Hb is saturated with oxygen), from which we conclude an enhanced Hb oxygen binding affinity (see the decrease in P50 in FIG. 1a; also evident from the corresponding leftward shift in ODC in FIG. 3).

[0055] The experimental data also show that the combination of α-KG and 5-HMF, especially at low and moderate doses, results in a comparable increase in Hb oxygen binding affinity compared to 5-HMF alone (Figure 1a), but with a smaller decrease in the Hill coefficient (as a measure of the cooperativity of Hb oxygen binding) (Figure 1b). It can be concluded that the additional presence of α-KG maintains the cooperativity of Hb oxygen binding to a higher degree than 5-HMF alone (as evident from the Hill coefficient determined in Figure 1b; as evident from the maintenance of the sigmoidal curve shape of the ODC in Figure 3).

[0056] Thus, the combination of α-KG and 5-HMF appears to advantageously improve hemoglobin oxygenation, allowing for efficient oxygen release in target tissues.

[0057] It is to be understood that the terms "dose" and "dosage" may be used interchangeably within the scope of this disclosure.

[0058] In further preferred embodiments according to the first or second aspect of the invention, the composition is administered (a) daily, as a single dose or divided into two or more equal or different doses; and / or (b) for at least 2, 3, 4, 5, 6 or 7 consecutive days; and / or (c) within 12 hours prior to exposure to ischemic and / or hypoxic conditions.

[0059] In a further preferred embodiment according to the first or second aspect of the invention, the composition is administered parenterally or orally.

[0060] However, in different embodiments, the form, route, duration and interval of administration can be adapted individually.

[0061] In a preferred embodiment of the first aspect of the invention, the disease or condition is (a) The following: (i) Impaired oxygen uptake in the lungs; (ii) impaired oxygen diffusion across the air-blood barrier; (iii) impaired hemoglobin oxygenation; (iv) decreased oxygen-carrying capacity of hemoglobin; (v) impaired oxygen transport due to blood flow restriction; (vi) microvascular oxygen saturation disturbances; and / or (vii) Insufficient oxygen supply A condition characterized by; (b) hypoxemia or anoxemia; (c) hypoxia or anoxia; (d) ischemia, preferably selected from gastrointestinal ischemia, cardiac ischemia, cerebral ischemia, renal ischemia, limb ischemia, neuronal ischemia, hypoxic-ischemic brain injury (hypoxic-ischemic encephalopathy) and ischemic neurological disorders; (e) a respiratory disease, preferably selected from inflammatory respiratory diseases, respiratory diseases associated with fungal, viral or bacterial infections, autoimmune-mediated respiratory diseases, idiopathic respiratory diseases, hyperproliferative respiratory diseases, cancer, asthma, chronic obstructive pulmonary disease (COPD), bronchitis, emphysema, pulmonary edema, acute respiratory distress syndrome (ARDS), bronchopulmonary dysplasia (BPD), pulmonary fibrosis, atelectasis, tuberculosis, pneumonia, pneumonitis, sinusitis, allergic rhinitis, pharyngitis, mucositis, stomatitis, bronchiectasis, lupus pneumonia, cystic fibrosis and respiratory failure; (f) altitude illness, preferably selected from acute mountain sickness (AMS), high altitude cerebral edema (HACE), high altitude pulmonary edema (HAPE), Da Costa syndrome and chronic mountain sickness (CMS); (g) COVID and / or one or more delayed / post-COVID complications thereof, preferably selected from delayed COVID symptoms and / or post-COVID syndrome; and / or (h) Chronic Fatigue Syndrome (CFS) is selected from.

[0062] The term "hypoxemia" refers to a decrease in the oxygen content (oxygen deficiency) in arterial blood.

[0063] The term "anoxia" refers to its most severe form, i.e. "hypoxia" in which the oxygen saturation in the blood is severely reduced (far below the physiologically required threshold). The term "anoxia" refers to the complete absence of oxygen.

[0064] The term "ischemia" refers to a reduction in blood flow, often painful, or a complete loss of blood flow to a tissue, body part, or organ, which may result in dysfunction. The most common cause of ischemia is a change in the form of narrowing or blockage of blood vessels. These may occur, for example, in the case of thrombosis or embolism. Narrowing is referred to as stenosis, such as in atherosclerosis and arterial occlusive disease (AOD). Functional stenosis may also occur, such as in Raynaud's syndrome or in the physiological response to circulatory shock. Ischemia impedes or stops the metabolism of cells. Ischemia caused by a restriction or blockage of blood flow is accompanied by a lack of oxygen in the affected area. If the supply to the nervous tissue is prolonged, a cascade occurs in which high intracellular calcium concentrations contribute to the uncontrollable release of the neurotransmitter glutamate, which ultimately damages the surrounding tissue cells. These processes may cause cell death (necrosis) and may result in infarction, for example in the case of ischemic heart disease, where a part of the myocardium does not receive an adequate blood supply, causing a heart attack. Pressure-related ischemia with accompanying tissue damage leads to pressure ulcers.

[0065] In a preferred embodiment of the first aspect of the invention, the hypoxemia is characterized by: (a) ventilation-perfusion mismatch; (b) impaired oxygen diffusion; (c) alveolar hypoventilation; (d) right-to-left shunt; and / or (e) impaired diffusion perfusion. is caused by.

[0066] In a preferred embodiment of the first aspect of the invention, the ischemia is caused by cardiac arrest, shock, carotid artery occlusion, hypotension, atherosclerosis, respiratory arrest, thoracic outlet syndrome, hypoglycemia, tachycardia, radiation therapy, chemotherapy, septic shock, heart failure, superior mesenteric artery syndrome, sickle cell disease, thalassemia, induced gravitational acceleration, extreme cold, increased stimulation of glutamate receptors, arteriovenous malformation, peripheral arterial occlusive disease, compression or rupture of a blood vessel supplying a tissue or organ, anemia and / or loss of consciousness.

[0067] According to a preferred embodiment of the first aspect of the present invention, the respiratory disease is characterised by reduced oxygen uptake in the lungs caused by a viral infection, wherein the virus causing the viral infection is preferably a coronavirus, preferably Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2), or a variant thereof.

[0068] "Chronic Fatigue Syndrome (CSF)", also known as "Myalgic Encephalomyelitis (ME)" or "ME / CFS", is a chronic disease whose main symptom is an abnormally rapid physical and mental fatigue, which in extreme cases can lead to widespread disability and the need for care. Despite the unknown causes and mechanisms of its development, the syndrome is internationally recognized as an independent disease pattern in which dysregulation of the nervous, immune and / or endocrine systems is observed. Studies have shown that CFS patients usually show marked metabolic disturbances in the energy balance of somatic cells, with limited ATP synthesis provided by mitochondria. As a result, there is chronically too little energy available in the body. This energy deficiency is caused, among other things, by nitrosative stress due to high levels of nitric oxide (NO). Due to the inhibition of the mitochondrial respiratory chain (ATP synthesis), cells with high energy demands, such as muscles, nerve cells, the immune system and cardiac muscle, are mainly affected. Considering the effect of the composition according to the invention determined in the context of the present invention, it is believed that the energy deficiency and associated disease symptoms (particularly fatigue / malaise) typically present in CFS patients can be therapeutically alleviated by the inventive use of the pharmaceutical composition according to the first aspect of the present invention.It is therefore believed that, in particular, the increase in oxygen supply in peripheral tissues has a beneficial effect on mitochondrial energy production (i.e. ATP synthesis), and thus at least partially alleviates the fatigue state associated with these diseases, which, apart from the lack of oxygen supply, is also caused by mitochondrial dysfunction (mitochondrial dysfunction and / or impaired ATP synthesis).

[0069] The term "long Covid", also called "post-COVID / long-COVID" or "post- / long-COVID", "long / post-COVID", "post-COVID", "post-COVID syndrome" and "post-COVID-19 condition according to ICD-10 GM", in English "post-acute sequelae of COVID-19 (PASC)", "chronic COVID syndrome (CCS)", "COVID-19 long-hauler" or "post-acute Covid-19 syndrome", refers to the delayed or prolonged health effects of novel coronavirus disease (COVID-19). As a rule, acute COVID-19 infection lasts up to four weeks, but it can last for several months, for example if hospitalization in an intensive care unit is required. However, long-term symptoms may persist beyond this period or may occur additionally, even if the disease course is mild or the infection is undetectable. In very rare cases, COVID-spreading symptoms also occur as a result of vaccination against the virus (post-vaccine syndrome). So far, there is no single, common definition of long-term effects. Symptoms observed include severe lung damage, inflammatory reactions, and changes in various organs, including shortness of breath, fatigue (post-COVID fatigue), confusion, and neurological disorders. The most commonly reported symptoms include, among others, respiratory disease, neurological disorders, mental health disorders, movement limitations, fatigue, and muscle weakness. "Long-term COVID symptoms" and "post-COVID symptoms" (also interchangeably referred to as "post-COVID syndrome") can have similar symptoms and are generally differentiated according to the duration of their presence into long-term COVID symptoms (continuation or new onset of symptoms for more than 4 weeks after acute infection) and post-COVID syndrome (symptoms persist or appear for more than 12 weeks).

[0070] Thus, similar to CFS, the pharmaceutical composition according to the first aspect of the invention and the associated enhanced hemoglobin-mediated oxygen delivery may also be expected to have a positive therapeutic and / or prophylactic effect not only in the treatment of acute COVID disease, but also in the treatment and / or prevention of "prolonged COVID symptoms" and / or the so-called "post-COVID syndrome", in particular in the treatment and / or prevention of the associated fatigue state.

[0071] In the context of the present invention, the term "COVID" co Rona vi rus d Coronavirus disease (COVID-19) generally refers to the coronavirus disease caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or its variants, preferably coronavirus disease-19 (COVID-19).

[0072] In a particularly preferred embodiment of embodiment (g) above, the protracted / late complications of COVID (e.g. protracted COVID symptoms and / or post-COVID syndrome) are characterized by one or more of the following symptoms: - physical and / or mental fatigue, preferably caused by a decrease in the capacity of the lungs and / or peripheral tissues to take up oxygen; - Reduced / impaired oxygen transport capacity and / or reduced / impaired oxygen loading of hemoglobin; and / or - Decreased / impaired mitochondrial energy production (ATP synthesis).

[0073] In a third aspect, the present invention relates to an in vitro method for enhancing the hemoglobin-mediated oxygen delivery capacity of blood comprising mixing a blood sample with a composition as defined in relation to the first or second aspect of the invention.

[0074] As used herein, the term "hemoglobin-mediated oxygen delivery capacity of blood" is interchangeable with the term "hemoglobin-mediated oxygen supply capacity of blood" and refers to the ability of red blood cells contained in blood to deliver or supply oxygen to tissues, organs and / or cells based on reversible oxygen binding to hemoglobin.

[0075] In a preferred embodiment, the method comprises: (i) contacting a blood sample, preferably a whole blood sample, containing red blood cells with a composition according to the invention under atmospheric conditions that favor oxygenation over oxygen release of hemoglobin contained in the red blood cells; and (ii) maintaining the blood sample under ambient conditions until further use. Includes.

[0076] In a preferred embodiment, the atmospheric conditions are characterized by an oxygen partial pressure of at least 30 mmHg.

[0077] In a fourth aspect, the present invention provides an in vitro method for reducing or preventing hypoxia-induced damage to an organ or tissue, the method comprising: (a) contacting an isolated organ or tissue in vitro with a preservation solution, the preservation solution comprising a composition-like composition as defined in relation to the first or second aspect of the invention; and (b) maintaining the organ or tissue in contact with the preservation solution in vitro until further use. and preferably (i) the contact of the organ or tissue with the preservation solution corresponds to complete immersion of the organ or tissue in the preservation solution; and / or (ii) the storage solution further comprises red blood cells, preferably plasma or whole blood; It concerns the method.

[0078] In a preferred embodiment of the method according to the third or fourth aspect of the invention, the α-KG and 5-HMF are (i) 0.07 to 7 mg of α-KG and 0.07 to 2.24 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.005 to 0.5 mg of α-KG and 0.005 to 0.160 mg of 5-HMF per gram of organ or tissue; Preferably: (i) 0.35 to 6.3 mg of α-KG and 0.11 to 2.1 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.025 to 0.45 mg of α-KG and 0.008 to 0.150 mg of 5-HMF per gram of organ or tissue; More preferably: (i) 0.42 to 6 mg of α-KG and 0.14 to 2.03 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.03 to 0.43 mg of α-KG and 0.01 to 0.145 mg of 5-HMF per gram of organ or tissue is applied at a concentration of

[0079] In another preferred embodiment of the method according to the third or fourth aspect of the invention, the α-KG and 5-HMF are (i) 0.42 to 5.88 mg of α-KG and 0.14 to 1.96 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.03 to 0.42 mg of α-KG and 0.01 to 0.14 mg of 5-HMF per gram of organ or tissue; More preferably: (i) 0.50 to 4.20 mg of α-KG and 0.17 to 1.4 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.04 to 0.30 mg of α-KG and 0.01 to 0.10 mg of 5-HMF per gram of organ or tissue; Even more preferably: (i) 0.5 to 1.8 mg of α-KG and 0.2 to 0.6 mg of 5-HMF per milliliter of blood sample or storage solution; or (ii) 0.04–0.13 mg of α-KG and 0.01–0.04 mg of 5-HMF per gram of organ or tissue is applied at a concentration of

[0080] In a fifth aspect, the present invention relates to the in vitro use of a composition as defined with respect to the first or second aspect of the invention for enhancing hemoglobin-mediated oxygen delivery to an organ or tissue.

[0081] In a preferred embodiment of the use according to the fifth aspect of the invention, (i) the organ or tissue is an organ or tissue isolated from the human or animal body; and / or (ii) the tissue is contained in a preservation solution (as defined in relation to the fourth aspect of the invention) and the composition contacts the organ or tissue such that the organ or tissue is completely immersed in the preservation solution.

[0082] In a preferred embodiment of the method according to the fourth aspect of the invention or the use according to the fifth aspect of the invention, the organ or tissue is a graft; the graft is preferably (i) kidney, liver, heart, pancreas, lung, skin, subcutaneous tissue, muscle and / or bone; and / or (ii) Free flap surgery, preferably free skin, skin-muscle and / or skin-muscle-bone flap surgery is selected from.

[0083] In a sixth aspect, the present invention relates to oxygen-enriched stored blood produced using the method according to the third aspect of the present invention.

[0084] In a seventh aspect, the present invention relates to a method for treating and / or preventing a disease or condition that would benefit from enhanced hemoglobin-mediated oxygen delivery, wherein a pharmaceutical composition comprising α-ketoglutarate (α-KG) and 5-hydroxymethyl-2-furfural (5-HMF) is administered in a pharma- ceutical effective amount to an individual suffering from or at risk of developing, or exposed to, such disease or condition.

[0085] It will be understood that the embodiments and preferred embodiments disclosed in relation to the first aspect of the invention, in particular those relating to the disease or condition to be treated, and those relating to the dosage and administration of the compositions of the invention and possible excipients, also apply, where applicable, to the method of the seventh aspect of the invention.

[0086] The term "oxygen saturation (SO2)" refers to the proportion of oxygen present in blood and the maximum oxygen capacity of blood as a percentage.

[0087] The term "peripheral oxygen saturation (SpO2)" refers to the oxygen saturation in the tissues and can be determined using various measurement methods known to those skilled in the art, such as non-invasive pulse oximetry.

[0088] The term "fraction of inspired oxygen" ("FiO2" is short for "fraction of inspired oxygen") is the percentage of oxygen in the inspired air.

[0089] The term "maximal oxygen uptake" (abbreviated "VO2max") refers to the maximum number of milliliters of oxygen the body can use per minute during exercise, typically expressed in milliliters of oxygen per minute (ml O2 / min). VO2max can be used as a criterion for assessing a person's endurance performance.

[0090] In an eighth aspect, the present invention provides a method for the preparation of a composition comprising: (a) enhancing hemoglobin-mediated oxygen delivery to mitochondria; and / or (b) enhancing mitochondrial energy production (preferably mitochondrial ATP synthesis); The present invention relates to a pharmaceutical composition for use in the treatment and / or prevention of diseases or conditions that would benefit from one or more of the effects of (a) and (b) above.

[0091] With regard to the eighth aspect of the invention, it is to be understood that the disease or condition may preferably be selected from one or more of the diseases or conditions disclosed in relation to the first aspect of the invention, and each of the preferred embodiments disclosed in relation to the first aspect of the invention, in particular with regard to the mixing ratios of the substances present in the composition, as well as possible doses, formulations and administration forms / types, where applicable, are also to be understood as being applicable to the pharmaceutical composition according to the eighth aspect of the invention and directly disclosed in relation to the latter.

[0092] Unless otherwise defined, 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 invention pertains. In case of conflict, the present patent specification, including definitions, shall control.

[0093] It will be understood that the definitions and embodiments of the invention set out above in relation to the first aspect of the invention also apply, where possible, to the second, third, fourth and further aspects of the invention.

[0094] With respect to the embodiments described in this specification, and particularly the claims, it is intended that any embodiment referred to in a dependent claim may be combined with any embodiment of any claim (independent or dependent) from which that dependent claim depends. In the case of independent claim 1 specifying three alternatives A, B and C, dependent claim 2 specifying three alternatives D, E and F, and claim 3 dependent on claims 1 and 2 and specifying three alternatives G, H and I, for example, it should be understood that the present specification expressly discloses embodiments corresponding to the combinations A,D,G;A,D,H;A,D,I;A,E,G;A,E,H;A,E,I;A,F,G;A,F,H;A,F,I;B,D,G;B,D,H;B,D,I;B,E,G;B,E,H;B,E,I;B,F,G;B,F,H;B,F,I;C,D,G;C,D,H;C,D,I;C,E,G;C,E,H;C,E,I;C,F,G;C,F,H;C,F,I. This also applies to combinations of the different options recited in the different subclaims.

[0095] Similarly, if a dependent claim refers back to several preceding claims, any combination of the subject matter covered by those claims is considered to be explicitly disclosed, even if no alternatives are set out in the independent and / or dependent claims. For example, in the case of independent claim 1, dependent claim 2 referring back to claim 1, and dependent claim 3 referring back to both claims 2 and 1, the combination of the subject matter of claims 3 and 1 is directly and unambiguously disclosed as well, for example, as the combination of the subject matter of claims 3, 2 and 1. If there is a further dependent claim 4 relating to any of claims 1-3, the combinations of the subject matter of claims 4 and 1, claims 4, 2 and 1, claims 4, 3 and 1, and claims 4, 3, 2 and 1 are directly and unambiguously disclosed.

[0096] The present invention is herein described, by way of example only, with reference to the accompanying drawings, for the purposes of illustrating preferred embodiments of the invention. [Brief description of the drawings]

[0097] [Figure 1] The effect of the substances at different doses (mean ± SD) is shown. a) Mean P50 of all subjects at baseline and at different doses. b) Mean HC values ​​of all subjects at baseline and at different doses. [Diagram 2] Dose-dependence of P50 changes. Light grey lines show individual P50 values, black lines correspond to P50 values ​​of all subjects (mean ± SD), dark grey lines are linear regression functions whose characteristics (P50 and R2 values) are shown in the graphs. a) Dose-dependence of 5-HMF+α-KG. b) Dose-dependence of 5-HMF. c) Dose-dependence of α-KG. [Diagram 3] The mean ODC for 5-HMF (black) and 5-HMF+α-KG (grey) is shown, expressed as the cross of the mean P50 and mean HC across all subjects. EXAMPLES

[0098] The examples illustrate the invention.

[0099] Example 1: Experimental Overview Shown in Examples 2 to 4 The present study deals with the investigation of the possible influence of the active ingredients 5-HMF, α-KG and the combination of 5-HMF and α-KG on the oxygen binding curve of hemoglobin. The oxygen binding curve was determined using a new in vitro method (described in Ref. 20:Woyke S, Stroehle M, Brugger H, Strapazzon G, Gatterer H, Mair N, Haller T. High-throughput determination of oxygen dissociation curves in a microplate reader - A novel, quantitative approach. Physiol Rep. 2021 Aug;9(16):e14995. doi: 10.14814 / phy2.14995. PMID: 34427400) on human whole blood samples. Three different doses (low, medium and high) were used. A strong dose-dependent increase in Hb-O2 affinity was observed with 5-HMF alone and with the combined addition of 5-HMF and α-KG. α-KG alone also increased Hb-O2 affinity, but to a lesser extent. As Hb-O2 affinity increased, the sigmoidal shape of the oxygen binding curve was better preserved by the combination of 5-HMF and α-KG than by 5-HMF alone. This effect, observed for the first time in this study, suggests a previously unknown synergistic effect of the combination of α-KG and 5-HMF on the allosteric regulation of hemoglobin, possibly due to α-KG-mediated stabilization of the known highly reactive 5-HMF.

[0100] Example 2: Materials and Methods Venous blood was collected from 20 healthy subjects (10 women, 10 men) aged 18–40 years. Subjects were non-smokers, not pregnant or lactating, had no known hemoglobinopathies, and had no history of recent illness, trauma, blood loss, or travel to high altitudes (>3000 m) for several days. Blood gas analysis was performed (ABL800flex, Radiometer) immediately after collection, and samples were kept on ice and processed within 8 hours.

[0101] 5-HMF and α-KG were provided in different carrier solutions (water, glucose, glucose and phosphate, acid, water and sodium hydroxide, water and phosphate, glucose and sodium hydroxide) and control solutions consisted of carrier solution alone without 5-HMF or α-KG. Ten μl of each of the three different concentrations of 5-HMF or α-KG and 5-HMF+α-KG were added to 100 μl of whole blood and gently mixed by resuspension.

[0102] The maximum recommended daily dose of 5-HMF in Shanpal® (Cyl, Austria) is 720 mg. Assuming that its total distribution in the blood volume of a 70 kg person is approximately 5000 ml (16), we defined a low dose of 5-HMF as 0.14 mg / ml (700 mg / 70 kg). The medium dose of 5-HMF (0.6 mg / ml, 3,000 mg / 70 kg) was chosen to correspond to the usual dose of Aes-103, a known anti-sickle cell drug that contains 5-HMF (10). The high dose of 5-HMF (2 mg / ml, 10,000 mg / 70 kg) corresponds to a dose of 5-HMF (143 mg / kg) that has been shown in animal studies to cause adverse effects such as elevated serum γ-globulin levels and relative spleen weight (17, 18). The low dose of α-KG was 0.42 mg / ml (2,100 mg / 70 kg), the medium dose was 1.8 mg / ml (9,000 mg / 70 kg), and the high dose was 6 mg / ml (30,000 mg / 70 kg). This means that when the two active ingredients, α-KG and 5-HMF, were used in combination, they were present in a mass ratio of 3:1 (α-KG:5-HMF).

[0103] ODC measurement For each of the three stated test concentrations of the active ingredients alone (α-KG or 5-HMF) or in combination (α-KG:5-HMF), the ODC (as duplicates) was determined using the patent pending in vitro method for high-throughput determination of oxygen dissociation curves (ODC) described by Woyke et al. (20) (Ref. 20: Woyke, S., Stroehle, M., Brugger, H., Strapazzon, G., Gatterer, H., Mair, N., & Haller, T. (2021). High-throughput determination of oxygen dissociation curves in a microplate reader - A novel, quantitative approach. Physiological Reports, 9, e14995. https: / / doi.org / 10.14814 / phy2.14995);see also "ASCENION GmbH - Novel gas flow system for high throughput determination of oxygen dissociation curves (ODC)" at https: / / www.ascenion.de / en / technoloqv-offers / novel-qas-flow-svstem-for-high-throughput-determination-of-oxygen-dissociation-curves-odc.

[0104] This real-time, high-throughput analysis allows for the first time accurate and efficient ODC measurements of whole blood samples using a simple adaptation of a standard microplate reader and a specially modified (gas-perfused) 96-well microliter measurement plate. To ensure stable conditions during the ODC measurement, the measurement setup was set up in an environmentally controlled closed device (EC box). This box is temperature-controlled (37 °C) and contains the experimental setup consisting of a humidifier, the cover of the microplate reader, tubes and valves. A modified 96-well plate (where a special coating treatment ensures a thin but adherent layer of red blood cells for optimal fluorescence measurements) was used, allowing a constant gas flow over all wells in a serpentine gas flow channel. The measurement plate is integrated into a gas system consisting of gas bags, gas hoses, humidifiers, gas mixers and peristaltic pumps.

[0105] Blood samples were placed into individual wells using a special stamping technique, such that a membrane consisting of several cell layers was formed. The oxygen saturation of each well was measured using dual-wavelength spectroscopy, and the oxygen partial pressure was measured using the fluorescence lifetime of a commercially available oxygen sensor at the inlet and outlet ports of the measurement plate. An oxygen ramp from approximately 20% to 0% by volume was created so that dissociation curves could be measured. Measurements were performed every minute on a conventional fluorescence plate reader, and up to 92 whole blood samples could be analyzed within approximately 25 minutes.

[0106] In the ODC experiments, a gas mixture containing a PCO2 of 40 mmHg was used and the temperature was kept constant at 37°C.

[0107] Data processing and statistical analysis: Statistical analysis showed no significant differences for the carrier solutions, so the results for the different carrier solutions at each concentration level were averaged for the final analysis.

[0108] Curve fitting, P 50Calculations of mean and Hill coefficient (HC) and graphs were generated using Excel (Microsoft 2016). Statistical analysis was performed using IBM SPSS Statistics 25. Unpaired t-tests were used to analyze baseline differences between substance compositions. P-values ​​between different doses and substance compositions were analyzed using ANOVA with repeated measures design. 50 and HC differences were determined.

[0109] Post-hoc t-tests (Bonferroni correction) and ANOVA with repeated measures design were used for individual substances or combinations of the two substances to identify the location and magnitude of changes. Linear regression analysis was used to determine the concentration dependence of each substance. P<0.05 was considered significant. Data are expressed as mean ± SD.

[0110] Example 3: Results The mean age of the subjects was 29.6 ± 3.0 years. Hemoglobin concentration (14.5 ± 1.3 g / dl) and hematocrit (44.4 ± 4.0%) were normal in this population, but pH was slightly lower than 7.40 (7.35 ± 0.04), which corresponds to higher carbon dioxide levels in venous blood compared with arterial blood (48.4 ± 7.4 mmHg).

[0111] At the start of the study, the P50 value was 25.1±1.3, with a slight sex difference (women 26.0±1.0 mmHg vs men 24.3±0.9 mmHg; P=0.001), and the HC value was 2.61±0.21.

[0112] ANOVA with repeated measures design showed that the total substance ( P < 0.001), dose ( P < 0.001) and interaction (substance × dose, P < 0.001) effects were significant for both P50 and HC ( Fig. 1 ).

[0113] 5-HMF significantly altered P50 (P<0.001) (Figures 1 and 2) and HC (P<0.001) (Figure 1b) in a dose-dependent manner. α-KG alone also altered P50 (P<0.001) and HC (P=0.012), but to a numerically significantly less extent than 5-HMF (Figure 1).

[0114] The addition of α-KG to 5-HMF showed general dose and substance effects compared with 5-HMF alone for P50 and HC (P<0.001), particularly on P50 at the high dose (P=0.028) (Fig. 1a) and HC at the low and medium doses (P<0.001 and P=0.015) (Fig. 1b).

[0115] For all substances and combinations, the dose dependence of P50 followed a linear course (Figure 2).

[0116] Overall, changes in P50 and HC change the shape of the ODCs in a specific way (Figure 3). At low doses and in the control solution, the typical sigmoid shape of the ODCs is clearly visible. At medium doses, the sigmoidal features are maintained, but due to the higher HC, the ODCs of the 5-HMF+α-KG combination have a more maintained sigmoidal shape compared to 5-HMF alone. At high doses, both ODCs lose the typical sigmoidal shape due to extremely low P50 and HC values.

[0117] Example 4: Observations The most important finding of this study is that the combined administration of 5-HMF and α-KG increased HbO2 affinity in human whole blood. A linear dose-dependence was observed for all substances and combinations, although to different degrees (i.e., α-KG showed the least effect).

[0118] Interestingly, HC differed between substance combinations, especially at low and intermediate doses, indicating changes in the shape of the ODC in addition to the P50-related shifts (Figure 3).

[0119] A notable finding of this study was the observed combined effect of 5-HMF and α-KG on Hill coefficient (HC). Changes in HC value directly affect the sigmoid shape of ODC, which is important for hemoglobin oxygen transport (2, 19). An acute increase in ODC promotes hemoglobin oxygen transport (2), because a smaller change in PO2 induces a larger change in SpO2, which means that O2 loading at the lung level and O2 unloading at the tissue level are promoted.

[0120] As the data disclosed herein demonstrate, the combination of 5-HMF and α-KG, particularly at low and medium doses, can maintain the sigmoid curve shape of the ODC while increasing HbO2 affinity compared to 5-HMF alone (Figure 3).

[0121] This effect, observed for the first time here, suggests a previously unknown synergistic effect of the combination of α-KG and 5-HMF on the allosteric regulation of hemoglobin, possibly due to α-KG-mediated stabilization of the known highly reactive 5-HMF.

[0122] conclusion 5-HMF induces a strong dose-dependent increase in HbO2 affinity. The combination of 5-HMF and α-KG exhibits higher HC values ​​at a lower P50 compared to 5-HMF alone, thereby maintaining the sigmoidal shape of the oxygen binding curve.

[0123] Example 5: Experimental Testing Procedures The physiological effect of the compositions of the invention on which the invention is based is manifested, inter alia, in a measurable increase in physical performance (muscle strength and / or endurance) compared to matched control individuals administered a placebo. Exemplary assessment methods known in the art and routinely usable for these and similar purposes, without any intended limitation, are muscle strength measurements (e.g., brachial or respiratory strength measurements), the so-called 1-minute sit-stand test, comparison of running / walking distance traveled within a certain time (e.g., in the form of the so-called 6-minute walk test, also known as "6MWT"). The effects observed in the in vitro test methods disclosed herein (see Examples 1-4) can also be evaluated using other common direct and / or indirect measurement methods for determining arterial blood oxygen saturation (qualitative and / or quantitative), exemplary methods including, inter alia, pulse oximetry measurements, blood gas analysis (e.g., using spirometery), heart rate variability analysis (HRV analysis), and near-infrared spectroscopy (NIRS). Various commercially available test assays are also available that can be used to measure and quantify ATP levels in vitro in samples obtained from patients / subjects (e.g., Luminescent ATP Detection Assay Kit by abcam (www.abcam.com)). At the time of filing this patent application, various clinical studies are planned or being performed that will further demonstrate the above effects, particularly the direct effects on humans.

[0124] It is understood that the examples and embodiments described herein are for illustrative purposes only, and that various modifications and variations to the examples and embodiments, as well as combinations of features otherwise described in this application, are readily apparent to those skilled in the art and fall within the scope of protection of the disclosure and claims of the invention described herein. All patents and patent applications referred to in this specification are incorporated in the disclosure of this application.

[0125] All patents, patent applications, publications, and documents referenced herein are hereby incorporated by reference in their entirety. Reference to the above patents, patent applications, publications, and documents is not an admission that any of said patents, patent applications, publications, and documents is relevant prior art, nor is it an admission as to the contents or dates of such publications or documents. Such references do not indicate a search for the relevant disclosures. Statements as to the dates or contents of documents are based on the information available and do not constitute an admission as to their accuracy or correctness.

[0126] Modifications can be made to the above content without departing from the basic aspects of the technology. However, although the technology has been described in considerable detail with reference to one or more specific embodiments, those skilled in the art will recognize that modifications can be made to the embodiments specifically disclosed in this application. However, these modifications and improvements are within the scope of the present invention.

[0127] The technology described herein by way of example may be conveniently practiced in the absence of any element not specifically disclosed herein. For example, in each instance, the terms "comprising," "consisting of," and "consisting essentially of" and "consisting of" may be replaced with one of the other two terms.

[0128] The terms and expressions used are intended to be descriptive rather than limiting, and the use of such terms and expressions is not intended to exclude equivalents of the features shown and described or portions thereof, but rather allows for various modifications within the scope of the claimed technology. The terms "method" and "procedure" are used interchangeably herein.

[0129] The terms "a" or "an" can refer to one or more of the element it modifies (e.g., "a carrier" can mean one or more carriers), unless the context is clear that one element or more than one element is being described.

[0130] As used herein, the term "approximately" or "about" refers to a value within 10% (i.e., plus or minus 10%) of the underlying parameter, and the use of the term "approximately" or "about" at the beginning of a series of values ​​modifies each of the values ​​(i.e., "about 1, 2, and 3" refers to about 1, about 2, and about 3). For example, a weight of "approximately 100 grams" can include weights between 90 grams and 110 grams. When a list of values ​​is provided herein (e.g., about 50%, 60%, 70%, 80%, 85%, or 86%), the list includes all intermediate values ​​and fractions thereof (e.g., 54%, 85.4%).

[0131] Thus, while the present technology has been specifically disclosed by representative embodiments and optional features, it should be understood that modifications and variations of the concepts disclosed herein may be made by those skilled in the art, and such modifications and variations are deemed to be within the scope of the technology.

[0132] Particular implementations of the technology are set forth in the claims.

[0133] [Table 1] [Table 2]

Claims

1. A pharmaceutical composition comprising α-ketoglutaric acid (α-KG) and 5-hydroxymethyl-2-furfural (5-HMF) for the treatment and / or prevention of diseases or conditions that would benefit from enhanced hemoglobin-mediated oxygen delivery.

2. A composition comprising α-KG and 5-HMF for use as a dietary supplement for enhancing hemoglobin-mediated oxygen delivery.

3. 3. The composition of claim 1, wherein α-KG and 5-HMF are present in the composition in a mass ratio of 1:1 to 100:1, preferably 2:1 to 50:

1.

4. 3. The composition of claim 1 or 2, wherein the composition is administered at a dose of 5 to 500 mg / kg body weight of α-KG and 5 to 160 mg / kg body weight of 5-HMF.

5. The composition comprises: (a) daily, either as a single dose or divided into two or more equal or different doses; and / or (b) for at least five consecutive days; and / or (c) within 12 hours prior to exposure to ischemia and / or hypoxia The composition of claim 1 or 2, wherein the composition is administered

6. The disease or condition is (a) The following: (i) impaired oxygen uptake in the lungs; (ii) impaired oxygen diffusion through the air-blood barrier; (iii) impaired hemoglobin oxygenation; (iv) decreased oxygen-carrying capacity of hemoglobin; (v) impaired oxygen transport due to blood flow restriction; (vi) microvascular oxygen saturation impairment; and / or (vii) Insufficient oxygen supply A condition characterized by: (b) hypoxemia or anoxemia; (c) hypoxia or anoxia; (d) ischemia, preferably selected from gastrointestinal ischemia, cardiac ischemia, cerebral ischemia, renal ischemia, limb ischemia, neuronal ischemia, hypoxic-ischemic brain injury (hypoxic-ischemic encephalopathy), and ischemic neurological disorders; (e) a respiratory disease, preferably selected from inflammatory respiratory diseases, respiratory diseases associated with fungal, viral or bacterial infections, autoimmune-mediated respiratory diseases, idiopathic respiratory diseases, hyperproliferative respiratory diseases, cancer, asthma, chronic obstructive pulmonary disease (COPD), bronchitis, emphysema, pulmonary edema, acute respiratory distress syndrome (ARDS), bronchopulmonary dysplasia (BPD), pulmonary fibrosis, atelectasis, tuberculosis, pneumonia, pneumonitis, sinusitis, allergic rhinitis, pharyngitis, mucositis, stomatitis, bronchiectasis, lupus pneumonia, cystic fibrosis, and respiratory failure; (f) altitude sickness, preferably selected from acute mountain sickness (AMS), high altitude cerebral edema (HACE), high altitude pulmonary edema (HAPE), Da Costa syndrome, and chronic mountain sickness (CMS); (g) COVID and / or one or more late / post-COVID complications thereof, preferably selected from prolonged COVID symptoms and / or post-COVID syndrome; and / or (h) Chronic Fatigue Syndrome (CFS) The composition of claim 1, selected from:

7. Hypoxemia is (a) Ventilation-perfusion mismatch; (b) impaired oxygen diffusion; (c) alveolar hypoventilation; (d) right-to-left shunt; and / or (e) Diffuse perfusion disorder The composition of claim 6, which is caused by:

8. 8. The composition of claim 6 or 7, wherein the ischemia is caused by cardiac arrest, shock, carotid artery occlusion, hypotension, atherosclerosis, respiratory arrest, thoracic outlet syndrome, hypoglycemia, tachycardia, radiation therapy, chemotherapy, septic shock, heart failure, superior mesenteric artery syndrome, sickle cell disease, thalassemia, induced g-forces, extreme cold, increased stimulation of glutamate receptors, arteriovenous malformation, peripheral arterial occlusive disease, compression or rupture of blood vessels supplying a tissue or organ, anemia, and / or loss of consciousness.

9. 8. The composition of claim 6 or 7, wherein the respiratory disease is characterized by reduced oxygen uptake in the lungs caused by a viral infection, and the virus causing the viral infection is preferably a coronavirus, preferably severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or a variant thereof.

10. 10. An in vitro method for enhancing the hemoglobin-mediated oxygen delivery capacity of blood, comprising mixing a blood sample with the composition of claim 1.

11. 1. An in vitro method for reducing or preventing hypoxia-induced damage to an organ or tissue, the method comprising: (a) contacting an isolated organ or tissue in vitro with a preservation solution, the preservation solution comprising the composition of claim 1; and (b) maintaining the organ or tissue in contact with the preservation solution in vitro until further use of the organ or tissue. and preferably (i) contact of the organ or tissue with the preservation solution corresponds to complete immersion of the organ or tissue in the preservation solution; and / or (ii) the storage solution further comprises red blood cells, preferably plasma or whole blood; method.

12. α-KG and 5-HMF (i) 0.07 to 7 mg of α-KG and 0.07 to 2.24 mg of 5-HMF per ml of blood sample or preservative solution; or (ii) 0.005 to 0.5 mg of α-KG and 0.005 to 0.160 mg of 5-HMF per gram of organ or tissue The method according to claim 10 or 11, wherein the concentration is

13. 10. In vitro use of a composition according to claim 1 or 2 to enhance hemoglobin-mediated oxygen delivery to an organ or tissue.

14. the organ or tissue is a transplant; The graft is preferably (i) kidney, liver, heart, pancreas, lung, skin, subcutaneous tissue, muscle and / or bone; and / or (ii) free flap surgery, preferably free skin, skin-muscle, and / or skin-muscle-bone flap surgery The method of claim 11, wherein the

15. Contains α-ketoglutaric acid (α-KG) and 5-hydroxymethyl-2-furfural (5-HMF), (a) enhancing hemoglobin-mediated oxygen delivery to mitochondria; and / or (b) enhancing mitochondrial energy production (preferably mitochondrial ATP synthesis); A pharmaceutical composition for the treatment and / or prevention of diseases or conditions that would benefit from one or more of the effects of (a) and (b) above.

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