HK1131340B - Compositions comprising alpha-ketoglutarate and their use for modulating muscle performance - Google Patents

Description

Compositions comprising alpha-ketoglutarate and their use for modulating muscle performance

Technical Field

The present invention relates to compositions for modulating muscle performance in mammals, including humans. Also relates to methods for modulating muscle performance in vertebrates (including mammals and birds), and to the preparation of compositions for preventing, alleviating or treating muscle performance in said vertebrates.

Background

Patients with end-stage renal disease (uremia) have a restricted physical constitution, which is often the cause of secondary problems, namely cardiac dysfunction, muscle abnormalities and even depression (Gutman et al, 1981; Painter, 1988, Kouidi et al, 1997; Shalom et al, 1984). To date, while muscle weakness is a common condition in dialysis patients, it remains an unexplained phenomenon and is therefore not easily treated, which severely impacts the patient's daily lives, whether work-related or recreational-related (Nakao et al 1982).

Recent studies on renal failure and uremic myopathies have focused on the training of patient health. Indeed, exercise training has been shown to significantly reduce muscle atrophy occurring in patients with end stage renal disease undergoing hemodialysis (Shalom et al, 1984; Painter, 1988; Eidemak et al, 1997; Kouidi et al, 1998). However, to date, little is known about the changes in muscle function and structure that occur as the disease progresses, or even the rehabilitation that motor training can have.

Muscle weakness (a symptom of uremia) is known to be associated with a definite loss of muscle strength (patients typically lose 140 newtons of quadriceps compared to control subjects; Fahal et al 1997). However, it is believed that this problem is not one of the disturbances in excitation-contraction coupling, as this is associated with the selective loss of force at low excitation frequencies that is not observed.

Muscle atrophy or loss is fiber-specific, in that uremia is primarily associated with loss of type II fibers, and indeed, in particular, with loss of type IIb fibers of the fast-shrinking glycolytic type (Fahal et al 1997), although atrophy of type I (slow-shrinking oxidized fibers) also occurs in some patients (Moore et al 1993).

Two recent studies have shown that people with developed muscles also have stronger bones. Studies from Johns Hopkins have shown that heart and lung performance is not related to stronger bone, but to muscle strength (Stewart et al 2002).

In addition, another study from Turkey showed that grip pressure (grip pressure) is closely and positively correlated with bone force mass (Sahin et al 2002).

It is therefore an object of the present invention to develop means and methods for treating or preventing any condition associated with reduced muscle performance, such as uremia, thereby enhancing muscle performance, which also avoid the problems associated with prior art means and methods. In this respect, the present invention meets this need and object.

Disclosure of Invention

The composition of the invention is characterized in that it comprises alpha-ketoglutarate (AKG) or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate or/and dipeptides of glutamine and other amino acids or/and tripeptides of glutamine and other amino acids or/and dipeptides and tripeptides of glutamine and other amino acids or/and mono-and divalent metal salts and other salts of alpha-ketoglutarate or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate and glucose in an amount of 0.1-5 mM.

Suitably, the composition comprises alpha-ketoglutarate (AKG) or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate or/and dipeptides of glutamine and other amino acids or/and tripeptides of glutamine and other amino acids or/and di-and tripeptides of glutamine and other amino acids or/and mono-and divalent metal salts and other salts of alpha-ketoglutarate or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate and optionally glucose in an amount of 0.1-5mM at a dose of 5-40mg/kg body weight. The composition is suitable for oral administration.

The composition may suitably also comprise alpha-ketoglutarate (AKG) or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate or/and dipeptides of glutamine and other amino acids or/and tripeptides of glutamine and other amino acids or/and di-and tripeptides of glutamine and other amino acids or/and mono-and divalent metal salts and other salts of alpha-ketoglutarate or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate and optionally glucose in an amount of 0.1-5mM for parenteral administration at a dose of 0.1-0.4mg/kg body weight or delivered by dialysis at a dose of 1-1000mg/kg body weight.

The present invention also provides a method for modulating muscle performance in a mammal comprising administering to a vertebrate in an appropriate amount and/or in an appropriate ratio for sufficient effect a composition comprising: alpha-ketoglutarate (AKG) or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate or/and dipeptides of glutamine and other amino acids or/and tripeptides of glutamine and other amino acids or/and dipeptides and tripeptides of glutamine and other amino acids or/and mono-and divalent metal salts and other salts of alpha-ketoglutarate or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate and additionally optionally glucose.

Preferably, in the method for modulating muscle performance in a mammal, the composition is administered at a dose of 5-40mg/kg body weight or 0.1-0.4mg/kg body weight, said composition optionally comprising glucose in an amount of 0.1-5 mM. Preferably, the composition is administered orally or parenterally to a vertebrate in need thereof selected from the group consisting of: rodents (such as mice, rats, guinea pigs or rabbits), farm animals (such as cows, horses, pigs, piglets), birds (such as hens and turkeys), free-living farm animals or birds, pets (such as dogs or cats), and humans.

Furthermore, the present invention provides a method for the preparation of a composition for preventing, reducing or inhibiting muscle performance in a vertebrate, said method comprising administering to a vertebrate in need thereof a composition comprising, in an appropriate amount or/and in an appropriate ratio, for sufficient effect: alpha-ketoglutarate (AKG) or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate or/and dipeptides of glutamine and other amino acids or/and tripeptides of glutamine and other amino acids or/and dipeptides and tripeptides of glutamine and other amino acids or/and mono-and divalent metal salts and other salts of alpha-ketoglutarate or/and glutamine or/and glutamic acid or/and ornithine alpha-ketoglutarate and additionally optionally glucose.

Preferably, in a method for the preparation of a composition for preventing, reducing or inhibiting muscle performance in a vertebrate, the composition is administered at a dose of 5-40mg/kg body weight or 0.1-0.4mg/kg body weight or 1-1000mg/kg body weight, said composition optionally comprising glucose in an amount of 0.1-5 mM. Preferably, the composition is administered orally or parenterally (especially by dialysis) to a vertebrate in need thereof selected from the group consisting of: rodents (such as mice, rats, guinea pigs or rabbits), farm animals (such as cows, horses, pigs, piglets), birds (such as hens and turkeys), free-living farm animals or birds, pets (such as dogs or cats), and humans.

The invention also relates to the use of said composition for the preparation of a medicament for preventing, reducing or inhibiting muscle performance in a vertebrate and for the preparation of a substance for modulating muscle performance in a mammal.

Preferably, the composition is a pharmaceutical composition containing a pharmaceutically acceptable carrier or/and additives for oral or parenteral administration.

The pharmaceutical composition is prepared in a therapeutically effective amount of 5-40mg/kg body weight/daily dose for oral compositions or 0.1-0.4mg/kg body weight/daily dose for parenteral compositions or 1-1000mg/kg body weight/daily dose for dialysis compositions, wherein optionally the amount of glucose is 0.1-5 mM.

Technical effects of the invention

The present invention provides a new and improved process for the preparation of a substance comprising an AKG component and glucose for improving muscle performance in a vertebrate in a situation where muscle performance is reduced compared to normal muscle performance in the subject or where muscle performance is enhanced compared to normal muscle performance.

The subject may be a vertebrate, such as a mammal or a bird, detailed examples of which are given below. Thus, the vertebrate may be those for which enhanced muscle performance is desired for medical or other reasons, such as enhanced muscle strength for athletes, exercisers, etc.

The method comprises the step of administering to the vertebrate a composition comprising alpha-ketoglutarate and glucose in a sufficient amount and/or in a sufficient proportion to obtain the desired effect. Muscle performance of a vertebrate (such as a mammal or bird) that does not have a reduction in muscle performance and to which the composition has not been administered is considered normal. Muscle performance is considered enhanced when the muscle performance is observed or measured to be enhanced as compared to the same mammal not administered the composition. An increase in muscle performance compared to the initial value of muscle performance in a vertebrate prior to administration of the composition can also be considered an increase.

The present invention also provides a method for modulating muscle performance in a mammal comprising administering to said mammal in need thereof a composition comprising AKG and optionally glucose to modulate muscle performance. The invention also provides the use of a composition comprising AKG and optionally glucose for the preparation of a composition for preventing, alleviating or treating a decrease in muscle performance.

Furthermore, the present invention provides the use of a composition comprising AKG and optionally glucose in the manufacture of a composition for modulating muscle performance in a vertebrate (including a mammal such as a human and an avian).

Drawings

FIG. 1 is a graph showing the decrease in isometric force (isometric force) of soleus muscles from 4-week-old rats during continuous electrical stimulation at 90 Hz.

Fig. 2 is a graph showing the contractile properties of soleus muscle in Uremic Buffer (n-8) or crenellated physiological solution (n-9) during continuous electrical stimulation at 40Hz (32mA, 1ms, < 90V).

Figure 3 illustrates the contractile properties of extensor longus in uremic buffer (n-7) or crenellated physiological solution (n-8) during continuous electrical stimulation at 90Hz (32mA, 1ms, < 90V).

Figure 4 graphically illustrates the contraction characteristics of soleus muscle in uremic rats + test component (n-10), control rats + placebo (n-8), control rats + test component (n-4) incubated in crenellated physiological solution (30 min incubation) and subjected to electrical stimulation at 40Hz (32mA, 1ms, < 90V).

Figure 5 shows schematically the contraction characteristics at 90Hz, 1ms, <90V of the extensor digitorum longus of uremic rats + test components (n ═ 3), control rats + test components (n ═ 2) incubated in creneline buffer (incubation for 30 minutes) and uremic rats + test components (n ═ 3) incubated in uremic buffer (incubation for 30 minutes).

Detailed Description

Definition of

The term "muscle endurance" means herein the ability of the muscle group to undergo submaximal contractions over an extended period of time, as defined by Zachazewski (1996, cited in the review by Wheeler, t.exercisephysiology552 (1997)). Since it is difficult to define exactly the exact parameter list, since it is very dependent on the individual evaluation of the amount of muscle force produced and the point of exhaustion, one can imagine that an individual active at very low muscle activity levels may last for several hours without becoming fatigued, while another individual active close to maximum muscle activity may last only for a short period of time. Muscle endurance measurements rarely take into account the forces generated, focusing more on the time of muscle activity.

The term "muscle strength" means herein absolute strength and relative strength, independent of gender, as long as the muscles studied are the same size. Thus, in absolute terms, the larger the cross-sectional area of a muscle, the stronger the muscle, regardless of gender (Martin, reviewed in l.exercise Physiology552 (1997)). However, men tend to be stronger than women because they tend to have greater upper body muscle mass, with longer muscles having a larger cross-sectional area. When strength is considered relative to body weight or fat free body mass, it is referred to as relative muscle strength. Most commonly, the amount of exertion is divided by body mass, fat-free body mass, muscle cross-sectional area, limb volume, waist circumference, and height. By dividing the force by these factors, the difference between men and women is significantly reduced or eliminated. Researchers have studied female performance and have shown that when performance is expressed in weight per unit of body fat removed, then female performance equals or exceeds that of its male counterpart. For this reason, a woman's swimming ability is closer to his male counterpart than any other sport.

The term "muscle force" means herein the component of work performed by a muscle, calculated by the following formula: work is force x distance, expressed in SI units (N/g net weight), where distance is a measure between the fulcrum and the point of application of force. The force can be easily measured with a transducer and expressed in time to give an index of contraction over a sustained period of muscle activity. There is a close correlation between the force profile of the active muscle and the RMS recorded by electromyography. Thus, a non-invasive measurement of muscle activity can be obtained by EMG recordings and interpreted in the form of muscle force by RMS conversion.

The term "modulating muscle performance" means herein altering, altering or otherwise affecting muscle performance in a subject.

The term "improved muscle performance" means herein a change in muscle performance, wherein the change is compared to a subject of the same species that has not been treated or administered according to the present invention. If these changes are positive for the mammal, then the changes are considered to be improvement. Generally, the improvement in muscle performance is an enhancement in muscle performance.

The term "parenteral" is herein intended to mean administration by injection not via the digestive tract but via some other route, and "pharmaceutical composition" as used herein is intended to mean a therapeutically effective composition according to the invention.

As used herein, a "therapeutically effective amount" or "therapeutically effective" refers to an amount that provides a therapeutic effect for a given condition and administration regimen. This is the predetermined amount of active material that is calculated to produce the desired therapeutic effect in association with the required additives and diluents (i.e., carriers or application vehicles). Further, it means an amount sufficient to reduce, alleviate and/or prevent clinically significant deficits in the activity, function and response of the host. Alternatively, the therapeutically effective amount is sufficient to ameliorate a clinically significant condition in the host. It will be appreciated by those skilled in the art that the amount of a compound may vary depending on its particular activity. A suitable dose may contain a predetermined amount of the active composition in association with a desired diluent (i.e., carrier or additive) calculated to produce the desired therapeutic effect.

As used herein, "treatment" means treatment that is a cure (which may be a complete cure or a partial cure) of a decrease in muscle performance.

The term "reduced" as used herein means a decreased (i.e., less) or milder condition associated with decreased muscle performance.

The term "prevention" as used herein means to completely or partially arrest the onset or onset of a decrease in muscle performance.

The term "derivative" means herein a chemical substance derived from the original substance either directly or by modification or partial substitution.

The term "analog" as used herein means a compound that is structurally similar to another compound, but not necessarily an isomer. Analogs have similar functions, but differ in structure or evolutionary origin.

In the methods and uses for preparing the compositions of the present invention, a therapeutically effective amount of the active ingredient is provided. As is well known in the art, a therapeutically effective amount can be determined by one of ordinary skill in the medical or veterinary arts based on patient characteristics such as age, weight, sex, condition, complications, other diseases, and the like.

The invention includes a composition comprising alpha-ketoglutaric Acid (AKG) and glucose in an amount of 0.1-5 mM. In another embodiment, the amount of AKG is 5-40mg/kg body weight. This amount is suitable when the composition is an oral composition, i.e. delivered orally.

In yet another embodiment, the amount of AKG is 0.1-0.4mg/kg body weight. This amount is suitable when the composition is to be delivered as a parenteral composition, i.e. by injection.

In yet another embodiment, the amount of AKG is 1-1000mg/kg body weight/day. This amount is suitable when the composition is delivered by dialysis.

According to the present invention, it includes a method for modulating muscle performance in a vertebrate. The method comprises administering to the vertebrate a composition comprising AKG and optionally glucose in addition, in a sufficient amount and/or in a sufficient proportion to achieve the desired effect, for modulating muscle performance.

The modulation may be an enhancement in muscle performance compared to a subject prior to or untreated from treatment of the particular subject. Any modulation may alter muscle performance from a reduced level to a normal or even higher level. The modulation may also be modulating the level of normal muscle performance of a particular species to a level above normal muscle performance. The level of muscle performance deemed normal for that particular species will depend on the species itself, age and sex. Any modulation that results in a beneficial change in muscle performance (i.e., a change that is inclined to or above normal muscle performance) is considered an improvement.

In yet another embodiment, the modulation may be inhibition, prevention, or alleviation of a decrease in muscle performance. Inhibition may be complete or partial inhibition in a mammal (such as a human) in need thereof. Prevention may be prophylactic treatment of a mammal, such as a human, in need thereof.

In addition, the invention includes methods for inhibiting, preventing or alleviating a decrease in muscle performance in a vertebrate. The method comprises administering to a vertebrate in need thereof a composition comprising AKG and optionally further glucose for inhibiting, preventing or alleviating said reduction in muscle performance.

In further embodiments, the vertebrate is selected from the group consisting of a rodent (such as a mouse, rat, guinea pig, or rabbit), a farm animal (such as a cow, horse, pig, piglet), a bird (such as a hen and turkey), a free-moving farm animal or bird, and a pet (such as a dog or cat).

In an even further embodiment, the mammal is a human. The person may be a patient in need of treatment for reduced muscle performance, an athlete, a malnourished person, or an athlete, a fitness person, an astronaut, or a person in need thereof due to any condition.

AKG and optionally glucose in the disclosed methods can be administered in different ways depending on the species of vertebrate being treated, the state of the vertebrate in need of the method, and the particular indication being treated.

Dosage forms may include capsules or tablets (such as chewable or soluble tablets, e.g. effervescent tablets) as well as powders and other dry forms well known to those skilled in the art, such as pills, such as pellets and granules.

Administration may be in parenteral, rectal or oral form, as for example a food or feed supplement. Parenteral vehicles include sodium chloride solution, ringer's dextrose, dextrose and sodium chloride, emulsified ringer's oil, or fixed oil, which are compatible with, for example, different types of injection strategies (e.g., subcutaneous, intramuscular, intra-orbital, intracapsular, intraspinal, intrasternal, intravenous, etc.).

In yet another embodiment, administration may be by dialysis.

The food and feed supplement may also be emulsified. The active therapeutic ingredient may then be mixed with excipients that are pharmaceutically acceptable and compatible with the active ingredient. Suitable excipients are, for example, water, saline, dextrose, glycerol, ethanol, and the like, as well as combinations thereof. Furthermore, if desired, the composition may contain minor amounts of auxiliary substances which enhance the effectiveness of the active ingredient, such as wetting or emulsifying agents, pH buffering agents.

Parenteral administration may be provided in various forms, such as liquid or lyophilized formulations or other dry formulations. It may include diluents having various pH and ionic strength containing various buffers (e.g. Tris-HCl, acetate, phosphate), additives to prevent adsorption to surfaces (such as albumin or gelatin), detergents (e.g. tween 20, tween 80, pluronic F68, bile acid salts), solubilizing agents (e.g. glycerol, polyethylene glycerol), antioxidants (e.g. ascorbic acid, sodium metabisulfite), preservatives (e.g. thimerosal, benzyl alcohol, parabens), bulking substances or tonicity adjusting agents (e.g. lactose, mannitol), covalent binding of polymers (such as polyethylene glycol) to the composition, complexation with metal ions, or incorporation of the substance into or onto particle preparations of polymeric compounds (such as polylactic acid, polyglycolic acid, hydrogels, etc.), or into or onto liposomes, microemulsions, micelles, hydrogels, etc, Monolayer or multilayer carriers, erythrocyte shells or spheroplasts.

In one embodiment, in any of the methods according to the present invention, the composition is administered in the form of a beverage or a dry composition thereof.

The beverage comprises an effective amount of glutamine, a glutamine derivative or metabolite, a glutamine analogue or a water-soluble non-toxic salt thereof or a mixture thereof and a nutritionally acceptable water-soluble carrier such as minerals, vitamins, carbohydrates, fat and protein. If the beverage is provided in dry form, all of these ingredients are provided in dry form. The provided beverage for consumption further comprises water. The final beverage solution may also have a controlled osmotic pressure and acidity, such as a buffer according to the general recommendations above.

The pH range is preferably about 2-5 (especially about 2-4) to prevent bacterial and fungal growth. Sterilized beverages having a pH of about 6-8 may also be used.

The beverages may be provided alone or in combination with one or more therapeutically effective ingredients.

Use of a composition comprising AKG and optionally glucose. The invention includes the use of a composition comprising AKG and optionally glucose.

One use is for the manufacture of a composition for preventing, reducing or treating a decrease in muscle performance in a vertebrate, including a human.

Another use is for the preparation of a composition for modulating muscle performance in vertebrates, including mammals (such as humans) and birds.

In another embodiment of the foregoing, the composition is a pharmaceutical composition. The pharmaceutical compositions may be used in the methods and uses disclosed herein together with pharmaceutically acceptable carriers and/or additives such as diluents, preservatives, solubilizers, emulsifiers, adjuvants and/or carriers.

In addition, as used herein, a "pharmaceutically acceptable carrier" is well known to those skilled in the art and may include, but is not limited to, 0.01-0.05M phosphate buffer or 0.8% saline. In addition, these pharmaceutically acceptable carriers may be aqueous or non-aqueous solutions, suspensions and emulsions. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oils (such as olive oil) and injectable organic esters (such as ethyl oleate). Aqueous carriers include water, alcoholic/aqueous solutions, emulsions or suspensions, including saline and buffered media. Parenteral vehicles include sodium chloride solution, ringer's dextrose, dextrose and sodium chloride, emulsified ringer's oil or fixed oils. Preservatives and other additives may also be present such as, for example, antimicrobials, antioxidants, chelating agents, inert gases and the like.

Additional embodiments include uses wherein the composition is an oral composition.

Yet another embodiment includes the use wherein the composition is a parenteral composition.

Yet another embodiment comprises the use of a therapeutically effective amount of AKG and optionally glucose. Examples of therapeutic amounts for oral administration, parenteral administration (such as injection and dialysis) are given in the dosage section of the administered or pharmaceutical composition.

The AKG composition prepared, which may be a pharmaceutical composition or a food or feed supplement, may optionally comprise a carrier and/or an amount of a second or further active ingredient that affects muscle endurance.

The above effects may contribute to the enhancement of muscle performance.

AKG can be administered in different doses depending on the route of administration used.

In one embodiment of the above method, AKG is administered in an amount of 5-40mg/kg body weight, i.e. 5, 10, 15, 20, 25, 30, 35, 40mg/kg body weight. This amount of AKG is suitable when the composition is administered orally.

In another embodiment, AKG is administered in an amount of 0.1-0.4mg/kg body weight, i.e., 0.1, 0.2, 0.3, or 0.4mg/kg body weight. This amount of AKG is suitable when the composition is administered parenterally.

In another embodiment, AKG is administered in an amount of 1-1000mg/kg body weight.

In still other embodiments, the optional glucose is administered in an amount of 0.1-5mM, i.e., 0.1, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5 mM.

In one embodiment, the composition comprises 0.1-0.4mg/kg body weight AKG and 0.1-5mM glucose. The compositions are suitable for parenteral administration.

In another embodiment, the composition comprises AKG in an amount of 5-40mg/kg body weight and glucose in an amount of 0.1-5 mM. The composition is suitable for oral administration.

In yet another embodiment, the composition comprises AKG in an amount of 1-1000mg/kg body weight and glucose in an amount of 0.1-5 mM. The composition is suitable for dialysis.

As noted above, the present invention relates to means and methods for treating, alleviating or preventing any condition associated with reduced muscle performance in a vertebrate, such as a mammal (including a human) or a bird. Disorders associated with reduced human muscle performance are patients with renal disease (such as hemodialysis patients, peritoneal dialysis patients, pre-dialysis patients and transplant patients), osteoporosis patients, elderly (such as 70 years and older), injured/operated/bedridden infants (such as premature infants, SGAs (small for gestational age)), athletes (such as cyclists, weightlifting athletes), spacemen (such as astronauts in and/or from space).

In birds, it will be readily understood by those of ordinary skill in the art that the methods and pharmaceutical compositions of the present invention are particularly useful for administration to any vertebrate, such as a bird or mammal (e.g., a human), in need thereof. Vertebrates include, but are not limited to, turkeys, hens or chickens and other animals suitable for broiler and free-range activity, or mammals (including, but not limited to, domestic animals such as feline or canine subjects), farm animals (such as, but not limited to, bovine, equine, caprine, ovine, and porcine subjects), wild animals (whether wild or in zoos), research animals (such as mice, rats, rabbits, goats, sheep, pigs, dogs, cats, and the like).

Examples

Example 1

It is well known that muscle endurance can be influenced by Na-channel and Na, K-pump concentrations. Indeed, changes in channel or pump concentration and/or gating/pump characteristics with age, disease or drug administration can affect the contractile performance of muscles quite significantly and often reversibly. The lower panel shows the effect of T3 administration to induce rapid upregulation of Na-channels previously upregulated by Na, K-pumps, noting the rapid fatigue rate associated with greater Na-influx and insufficient Na, K-pump capacity in these muscles (Harrison & Clausen, 1998).

This example is illustrated in figure 1, which figure 1 illustrates the process of isometric force decline of soleus muscles from 4 week old rats treated with control (. smallcircle.) and 3, 5, 3' -triiodothyroxine (T3) for 48 hours (●) during continuous electrical stimulation at 90Hz until-80% of maximal force is lost. T3 was administered subcutaneously (day 1: 0.4. mu.g/g body weight/day; day 2: 0.2. mu.g/g body weight/day) while sham-injected (sham injected) with an equal volume of solvent to the control. The time required for the peak isometric contraction force to drop by 75%, defined as endurance, was determined by reading the intercept of the dashed line with the curve showing force versus the x-axis. The force decline curves for soleus muscle for control and T3-treated 48-hour rats can be fitted with the following single exponential equation, respectively: y ═ exp (0.42x +4.51), r ═ 0.98 and y ═ exp (0.60x +4.52), r ═ 0.99. Each point represents the mean ± sem observed on 8-18 muscles (vertical line). P <0.001 compared to the mean of controls; p < 0.01; p <0.05 (unpaired t-test).

Animal feeding

Animals were housed in a thermostatically regulated chamber containing equipment for measuring the contractile force of isolated rat skeletal muscles, i.e. computer, analog-to-digital converter, amplifier, differential amplifier, stimulator, 5% oxygen and 5% carbon dioxide.

The thermostated chamber was maintained at 30 ℃, containing a glass diffuser (for effective oxygenation of the buffer), a fixed seat/stimulation seat (with two needles to fix the fibula section in situ and 2 silver stimulation electrodes).

Results

Changes in muscle performance

The above measurements were recorded and the results are shown in figure 1. The results show that AKG administration to uremic rats gives a better slow-shortening soleus muscle endurance profile (AKG effect) compared to the same muscle from sham-operated control rats administered placebo (cren-linder buffer).

Indeed, soleus muscles from uremic rats administered AKG and incubated in crenellated ringer's buffer performed similarly to soleus muscles from sham-operated control rats administered placebo and incubated under the same conditions.

In control rats, administration of AKG improved the endurance of soleus muscle incubated in crenellated buffer compared to placebo (figure 1) (AKG effect).

Administration of AKG (clintheri's buffer) to uremic rats gave better endurance characteristics (buffer action) of the slow-twitch soleus muscle compared to the same muscle from uremic rats administered AKG (uremic buffer).

Administration of AKG (clinkeri buffer) to uremic rats gave better endurance characteristics (buffer action) of the extensor digitorum longus muscle (EDL) of the rapid-contraction type than the same muscle incubated in uremic buffer from uremic rats administered AKG.

EDL muscle appears to be relatively unaffected by incubation time in uremic buffer (n ═ 2).

Explanation of the invention

In terms of endurance, AKG administration during sustained contraction appears to have a beneficial effect on muscle performance.

The buffer components in which the muscle is incubated have an AKG-independent effect on muscle function.

Example 2

Uremic patients with chronic renal failure have excessive muscle fatigue that affects not only their professional life but also their recreational life. In addition, these patients suffer from muscle atrophy that is predominantly type II (rapid-shrink) fibers. These patients are known to have problems with anemia, but this does not account for muscle fiber loss or muscle weakness, and again, the data shows that local metabolic factors are not affected (normal ATP and phosphocreatine concentrations in uremic rats at rest) and that mitochondria are not abnormal.

However, one difference that does exist in patients with chronic renal failure is the difference in insulin resistance. The symptoms can be partially ameliorated by hemodialysis, but are not associated with hyperparathyroidism, which affects only insulin secretion. In these patients, basal glucose transport is normal, as is the amount of insulin-sensitive transport protein. However, it appears that muscle blood flow in these patients is reduced, which will affect glucose delivery and uptake by the muscle fibers.

In view of the above, it is clear that we are solving two separate aspects-above all the energy supply problem, since insulin resistance will reduce the glucose available to the deep muscle bed of the body. Here we expect that AKG will provide the muscle fibres directly with an energy component suitable for the TCA cycle before the critical energy production step. This hypothesis is very consistent with our observed response in control rats fed AKG compared to control rats given placebo. The second aspect is the problem of incubation media (bathing media) because uremic buffers cause a loss of strength (endurance) that can be reversed by incubation in creutzfeldt-lind buffer. Indeed, it has been documented that incubating uremic muscles in creutzfeldt-linder buffer restores very much of the lost endurance; in other words, we take the form of a dialysis treatment, observing a similar recovery of muscle function after dialysis as compared to before dialysis in human subjects.

According to a second aspect, it now appears from studies from isolated muscle that the loss of endurance is indeed buffer-related, i.e. related to the accumulation of compound ions etc. that occur in renal failure. We know that K ions can and do affect the sarcolemma, as do Ca ions, and therefore in our study we selectively removed these ions from the composition of uremic buffers. In contrast, the concentration of bicarbonate, urea and phosphate was increased compared to the kresoxim-methyl buffer. It is known that the membrane is excited by the operation of the Na, K-pump, and that accumulation of phosphate alone inhibits normal Na, K-pump operation, resulting in loss of excitation, and therefore loss of strength (endurance) faster than the isolated muscle of the control rats observed. The accumulation of phosphate leads to a disturbance in the excitability of the muscle fibres, affecting initially those fibres that are most dependent on the pump (i.e. fast fibres) and over a period of time leading to fibre damage/atrophy.

Results

1. As a result of the reduced blood flow in muscles, insulin resistance affects type I and II diabetes, arterial hypertension or renal failure patients, etc., in which AKG can be used as an energy-supplementing substance, especially in muscles which, unlike ion-coupled exchange systems, it diffuses more readily than glucose (requiring transporters), and possibly as a substance that increases the capillary bed of strong muscles.

2. AKG has been shown to reduce plasma phosphate levels, which alleviates the effects of phosphate on the sarcolemma (and other cells) and delays the development of uremia, if not stopped.

3. Administration of AKG to athletes will enhance their endurance performance in a natural and non-addictive manner over a period of days to weeks.

Figure 2 is a graph illustrating the plot of soleus muscle control (cre-linder buffer) -curve I, soleus muscle control (uremic buffer) -curve II, showing minimal effect of uremic buffer on muscle endurance of slow-contracting soleus muscles, noting that the first fatigue signal begins to appear after prolonged continuous stimulation (30-40 seconds). This finding supports the disclosed data from patients in whom slow muscles are less affected than fast muscles during chronic renal failure.

Figure 3 is a graph showing the rapid effect of uremic buffer on muscle endurance of the fast-contracting extensor digitorum longus of the extensor digitorum longus control group (crenulata buffer) -curve I, and of the extensor digitorum longus control group (uremic buffer) -curve II. Note that the first fatigue signal begins to be displayed only after 2-4 seconds of continuous stimulation.

FIG. 4 illustrates the observable effect of AKG administration to control soleus muscle in endurance improvement-curve I; curve I relates to uremic rats + test components, curve II to control rats + placebo and curve III to uremic rats + test components. It was noted that uremic rats administered AKG showed similar slightly improved endurance compared to the muscles of placebo-administered control rats.

Figure 5 illustrates the effect of AKG resistance to the extensor digitorum longus of uremic rats compared to placebo-treated uremic rats and control rats; curve I relates to uremic rats + test components (cre-linder buffer), curve II to uremic rats + test components (uremic buffer), and curve III to control rats + test components. It was noted that the performance of uremic rats administered AKG was improved compared to uremic rats administered placebo.

Conclusion

1) AKG is an oral formulation that improves muscle function in 21 days or less.

2) Oral administration of AKG has a specific effect on the contractile performance of muscles and/or fibres, which is clearly different between fast and slow contraction muscles.

3) AKG acts to reduce "peripheral" fatigue that occurs when muscles contract at their normal operating frequency over an extended period of time.

4) Maximum isometric contraction force (N): treatment with oral AKG (21 days or less) resulted in a 28% increase in the force produced in fast-twitch muscles and a 12% increase in the force produced in slow-twitch muscles compared to untreated controls.

5) Rate of force generation: AKG oral treatment (21 days or less) reduced the rate of force production by the fast-contracting muscles by about 8.5% and the rate of force production by the slow-contracting muscles by about 130%.

Mode of action of AKG

AKG acts on muscle contraction performance at multiple levels; improve fatigue resistance, increase strength and reduce the rate of strength generation.

Membrane potential: one obvious explanation for the improvement of fatigue resistance is that the AKG increases the activated membrane bound Na⁺，K⁺-the amount of ATPase. These units are responsible for maintaining the membrane resting potential and thus for fiber excitability. In experiments where pump activity was increased or inhibited, fatigue resistance was significantly improved or reduced, respectively.

Increase in muscle capacity: an increase in fiber size and the number of contractile units per fiber will greatly improve muscle isometric contraction. In this regard, one may assume that AKG has an anabolic effect on muscle hypertrophy. Alternatively, one may consider AKG to affect the cross-bridge circulation such that an improvement in force production is achieved. It is known that changes in e.g. RLC phosphorylation can affect not only the force generated, but also the rate of force generation in a muscle-specific manner.

Variation of fiber type: alternatively, one can predict the change in fiber type with AKG treatment. Indeed, an increase in MHC class I fiber components or Ca⁺⁺The change of the ATPase isozymes to the isozymes of the slow contraction type fibers will demonstrate a slower rise time of both muscles. E.g. slower Ca⁺⁺Release is the reason that the rise to peak force is slow.

Claims (16)

1. Use of alpha-ketoglutarate (AKG) or a salt of a mono-or divalent metal salt thereof selected from AKG and ornithine-AKG for the manufacture of a medicament for preventing, reducing or inhibiting a reduction in muscle performance in a subject.

2. The use according to claim 1, wherein the subject is selected from the group consisting of mouse, rat, guinea pig, rabbit, cow, horse, pig, piglet, hen, turkey, dog, cat, and human.

3. Use according to claim 1, wherein the medicament is a pharmaceutical composition comprising a pharmaceutically acceptable carrier and/or additive in addition to the active ingredient according to claim 1.

4. Use according to claim 3, wherein the composition is an oral composition, a parenteral composition or a composition to be administered by dialysis.

5. Use according to claim 4, wherein the medicament is administered orally at a dose of 5-40mg/kg body weight/day.

6. Use according to claim 4, wherein the medicament is administered parenterally in a dose of 0.1-0.4mg/kg body weight/day.

7. Use according to claim 4, wherein the medicament is administered by dialysis at a dose of 1-1000mg/kg body weight/day.

8. Use according to claim 3, wherein the composition further comprises glucose in an amount of 0.1-5 mM.

9. Use according to any one of claims 1 to 8, wherein the medicament is for the treatment of muscle fatigue.

10. Use according to claim 9, wherein the muscle fatigue is muscle fatigue in uremic patients.

11. The use according to claim 10, wherein the uremic patient is a uremic patient suffering from chronic renal failure.

12. Use according to any one of claims 1 to 8, wherein the medicament is for the treatment of muscle atrophy.

13. Use according to claim 12, wherein the muscle atrophy is muscle atrophy in uremic patients.

14. The use according to claim 13, wherein the uremic patient is a uremic patient suffering from chronic renal failure.

15. Use according to any one of claims 1 to 8, wherein the medicament is for a subject selected from: renal patients, hemodialysis patients, peritoneal dialysis patients, pre-dialysis patients, transplant patients, osteoporosis patients, elderly, injured/operated/bedridden infants, premature infants, small for gestational age, athletes, astronauts in and/or returning from space.

16. The use according to claim 15, wherein the elderly is elderly at least 70 years of age.