Compositions and methods using adenosylcobalamin

Abstract

Compositions effective for restoring mitochondrial function and other cellular functions and / or increasing mitochondrial energy in one or more cells contain adenosylcobalamin. Other aspects are directed to methods for improving physiological conditions associated with metabolic fatigue of one or more cells and / or reducing fatigue in an individual, treating, reducing the incidence, and / or reducing the severity of chronic diseases and / or mitochondrial-related diseases or conditions associated with altered mitochondrial function or reduced mitochondrial density.

Description

[Technical field]

[0001] [Background technology]

[0001] The present disclosure relates generally to compositions and methods for increasing cellular energy and / or treating or preventing mitochondrial-associated diseases or conditions, for example, by increasing intracellular ATP production, reducing oxidative stress, and / or enhancing mitochondrial function in an individual.

[0002] Adenosine triphosphate (ATP) is a complex organic compound that provides the energy to drive many processes in living cells, such as muscle contraction, nerve impulse propagation, and chemical synthesis. ATP is found in all living organisms and is often referred to as the "energy currency of the organism" for intracellular energy transfer. When consumed in metabolic processes, ATP is converted to either adenosine diphosphate (ADP) or adenosine monophosphate (AMP). ATP is also a precursor to DNA and RNA and is used as a coenzyme. ATP is also the substrate for adenylyl cyclase, most commonly in G protein-coupled receptor signaling pathways, where it is converted to the second messenger cyclic AMP, which releases calcium stores in the cell to initiate a calcium signal. This form of signaling is involved in the regulation of a variety of cellular processes, but is particularly important in brain function.

[0003]

[0003] Mitochondria are the main source of aerobic energy production in mammalian cells and are key organelles involved in cellular energy production. Mitochondrial dysfunction can lead to excessive fatigue and other symptoms that are common complaints in almost all chronic diseases. At the molecular level, impaired mitochondrial function can result in reduced efficiency of oxidative phosphorylation and reduced production of adenosine-5'-triphosphate (ATP). Clinical trials have demonstrated that L-carnitine, alpha-lipoic acid, coenzyme Q, and mitochondrial enzymes can reduce the production of oxidative phosphorylation and ATP. 10The use of oral supplements such as mitochondrial replacement therapy, ...

[0004]

[0004] Fatigue is a multidimensional sensation perceived as a decline in whole-body energy, and is considered to be unable to perform even simple tasks without exertion. Mild fatigue can be caused by a number of conditions, including depression and other psychological conditions, while moderate to severe fatigue involves the cellular energy system. At the cellular level, moderate to severe fatigue is associated with impaired mitochondrial function and reduced ATP production. Intractable fatigue lasting more than six months that is not reversed by sleep (chronic fatigue) is the most common complaint among patients presenting to general practice. Chronic fatigue is also an important secondary condition in many clinical diagnoses, and often precedes a patient's primary diagnosis.

[0005]

[0005] Furthermore, mitochondrial function is impaired as a result of aging and chronic disease due to oxidative damage to the mitochondrial membrane. As an example, individuals with chronic fatigue syndrome show evidence of oxidative damage to DNA and lipids, including oxidized blood markers and oxidized membrane lipids, indicative of excessive oxidative stress.

[0006] [Summary of the Invention]

[0006] In view of the experimental data disclosed later in this specification, the inventors believe that adenosylcobalamin (adenosyl B12) enhances the efficiency of mitochondrial energy production by increasing the ATP-synthase-dependent component of respiration, in contrast to other B12 isomers, and by enhancing ATP production stimulated by epibatine.

[0007]

[0007] Thus, in a general embodiment, the present disclosure provides a method for restoring mitochondrial function and other cellular functions and / or increasing mitochondrial energy in one or more cells, particularly by increasing ATP production and mitochondrial respiration, comprising administering an effective amount of adenosylcobalamin to an individual in need thereof.

[0008]

[0008] In one embodiment, the present disclosure provides a method for improving a physiological condition associated with metabolic fatigue of one or more cells and / or reducing fatigue in an individual, the method comprising administering an effective amount of adenosylcobalamin to an individual in need thereof.

[0009]

[0009] In another embodiment, the present disclosure provides a method for treating, reducing the incidence, and / or reducing the severity of a chronic disease, the method comprising administering an effective amount of adenosylcobalamin to an individual in need thereof.

[0010]

[0010] In one embodiment, at least a portion of the one or more cells is part of at least one body part selected from the group consisting of liver, kidney, brain, heart, intestine, pancreas, immune cells, and skeletal muscle.

[0011]

[0011] In another embodiment, the present disclosure provides a method for treating, reducing the incidence, and / or reducing the severity of a mitochondrial-associated disease, or a condition associated with altered mitochondrial function, or reduced mitochondrial density, the method comprising orally administering an effective amount of adenosylcobalamin to an individual in need thereof.

[0012]

[0012] The mitochondrial-related disease or condition can be selected from the group consisting of stress, physiological aging, obesity, slowed metabolic rate, metabolic syndrome, diabetes mellitus, diabetic complications, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stressor-related cognitive dysfunction, mood disorders, anxiety disorders, age-related neuronal cell death or dysfunction, chronic renal disease, renal failure, chronic heart failure, cardiac rehabilitation, orthopedic rehabilitation, wound healing, recovery from surgery, trauma, infection, cancer, hearing loss, macular degeneration, myopathy, and dystrophies, and combinations thereof.

[0013]

[0013] In a further embodiment, a method is provided for delaying the onset of metabolic decline, reducing oxidative stress, maintaining immune function, and / or maintaining cognitive function in healthy middle-aged or older adults, the method comprising orally administering an effective amount of adenosylcobalamin to a healthy middle-aged or older adult person.

[0014]

[0014] This also relates to a method for enhancing metabolism of reactive oxygen species and improving glucose control in an individual having at least one of obesity or diabetes, which method comprises orally administering an effective amount of adenosylcobalamin to the individual.

[0015] An advantage of one or more embodiments provided by the present disclosure is that they promote healthy aging of cells.

[0016] Another advantage of one or more embodiments provided by the present disclosure is that it helps combat the metabolic slowdown that accompanies aging.

[0017] Another advantage of one or more embodiments provided by the present disclosure is that they help increase fatty acid metabolism.

[0018] Yet another advantage of one or more embodiments provided by the present disclosure is that it helps the body metabolize fat and increase lean body mass.

[0019] An advantage of one or more embodiments provided by the present disclosure is that it helps maintain cardiac health.

[0020] Another advantage of one or more embodiments provided by the present disclosure is that they help maintain healthy LDL cholesterol and fatty acid concentrations in the blood.

[0021] Yet another advantage of one or more embodiments provided by the present disclosure is that they help reduce oxidative stress on the body.

[0022]

[0022] Additional features and advantages are described herein, and will be apparent from the drawings and detailed description that follow. [Brief description of the drawings]

[0023] 1-4 are graphs of data from the experimental examples disclosed herein. [Figure 1] Adenosyl B12 increases the ATP synthase-dependent component of respiration in stimulated human skeletal myotubes. Myotubes were treated for 3 h with the indicated concentrations of adenosyl B12 or without it (control). The oxygen consumption rate was then measured and myotubes were stimulated with epibatidine (10 μM). Statistical evaluation of the effect of adenosyl B12 on the ATP synthase-dependent component of respiration after stimulation was calculated by inhibiting mitochondrial ATP synthase with oligomycin (2.5 μg / ml). The graph shows the average of 16 cell assays. Results are expressed as mean ± SEM. * indicates a statistically significant difference with respect to control cells (white) at P < 0.05 (one-way ANOVA test). [Diagram 2]Methyl B12 does not increase the ATP synthase-dependent component of respiration in stimulated human skeletal myoblasts. Myotubes were treated for 3 hours with the indicated concentrations of methyl B12 or without it (control). The oxygen consumption rate was then measured and myotubes were stimulated with epibatidine (10 μM). Statistical evaluation of the effect of methyl B12 on the ATP synthase-dependent component of respiration after stimulation was calculated by inhibiting mitochondrial ATP synthase with oligomycin (2.5 mg / ml). The graph shows the average of 16 cell assays. Results are expressed as mean ± SEM. NS, not significant (one-way ANOVA test). [Diagram 3] Adenosyl B12, but not methyl B12, enhances epibatine-stimulated ATP production in human skeletal myotubes during acute treatment. Human myotubes were treated for 3 h with the indicated concentrations of adenosyl B12 or methyl B12. Statistical evaluation of the effect of B12 isoforms on epibatidine-induced ATP production. The graph shows the mean of three independent experiments. Results are expressed as mean ± SEM. * indicates a statistically significant difference compared to control cells (white) at P < 0.05 (one-way ANOVA test). [Figure 4] Chronic treatment of human skeletal myotubes with adenosyl B12, but not methyl B12, enhances epibatine-stimulated ATP production. Human myotubes were treated for 3 days with the indicated concentrations of adenosyl B12 or methyl B12. Statistical evaluation of the effect of B12 isoforms on epibatidine-induced ATP production. Graphs show the mean of three independent experiments. Results are expressed as mean ± SEM. * indicates statistically significant difference vs. control cells (white) at P < 0.05 (one-way ANOVA test). [Figure 5-1]Adenosyl B12 specifically displays a gene expression signature of oxidative phosphorylation genes in skeletal muscle. [FIG.5A] Skeletal muscle gene enrichment plot analysis comparing adenosyl B12-treated aged rats to methyl B12-treated aged rats shows enrichment for the oxidative phosphorylation gene set. [FIG.5B] Network representation of protein-protein interactions in skeletal muscle of adenosyl B12-treated aged rats shows that oxidative phosphorylation genes are differentially regulated. No network representation of oxidative phosphorylation genes is possible in methyl B12-treated animals. [Figure 5-2] Adenosyl B12 specifically displays a gene expression signature of oxidative phosphorylation genes in skeletal muscle. [FIG.5A] Skeletal muscle gene enrichment plot analysis comparing adenosyl B12-treated aged rats to methyl B12-treated aged rats shows enrichment for the oxidative phosphorylation gene set. [FIG.5B] Network representation of protein-protein interactions in skeletal muscle of adenosyl B12-treated aged rats shows that oxidative phosphorylation genes are differentially regulated. No network representation of oxidative phosphorylation genes is possible in methyl B12-treated animals. [Figure 6] Effect of Adenosyl B12 on activity time on the rod in aged rats. Results are expressed as mean ± SEM, n = 15. * indicates statistically significant difference vs. control cells (white) at P < 0.05 (Student's t-test). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024]

[0030] definition

[0031] Some definitions are provided below. However, definitions may be found in the "Embodiments" section below, and the heading "Definitions" above does not imply that such disclosure in the "Embodiments" section is not a definition.

[0025]

[0032] All percentages set forth herein are by weight of the total weight of the composition unless otherwise indicated. As used herein, "about," "approximately," and "substantially" are understood to refer to numbers within a numerical range, e.g., within -10% to +10% of the referenced number, preferably within -5% to +5% of the referenced number, more preferably within -1% to +1% of the referenced number, and most preferably within -0.1% to +0.1% of the referenced number. All numerical ranges herein should be understood to include all integers or fractions within the range. Furthermore, these numerical ranges should be interpreted as supporting claims directed to any number or subset of numbers within the range. For example, a disclosure of 1 to 10 should be interpreted as supporting ranges such as 1 to 8, 3 to 7, 1 to 9, 3.6 to 4.6, 3.5 to 9.9, etc.

[0026]

[0033] As used in this disclosure and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a component" or "the component" includes two or more components.

[0027]

[0034] The terms "comprise", "comprises", and "comprising" should be construed as inclusive rather than exclusive. Similarly, the terms "include", "including", and "or" should all be construed as inclusive unless such interpretation is clearly prevented by the context. However, the compositions disclosed herein may not include elements not specifically disclosed herein. Thus, disclosure of an embodiment using the term "comprising" includes disclosure of embodiments "consisting essentially of" and "consisting of" the specified component. A composition "essentially consisting of" contains at least 50% by weight of the referenced component, preferably at least 75% by weight of the referenced component, more preferably at least 85% by weight of the referenced component, and most preferably at least 95% by weight of the referenced component.

[0028]

[0035] The term "and / or" used in the context of "X and / or Y" should be interpreted as "X" or "Y" or "X and Y." Similarly, "at least one of X or Y" should be interpreted as "X" or "Y" or "X and Y." For example, "at least one of mental performance or muscular performance" should be interpreted as "mental performance or muscular performance" or "muscular performance" or "both mental performance and muscular performance."

[0029]

[0036] As used herein, the terms "example" and "such as," especially when followed by a list of terms, are merely exemplary and illustrative and should not be considered as exclusive or exhaustive. As used herein, a condition "associated with" or "linked with" another condition means that the conditions occur simultaneously, preferably that the conditions are caused by the same underlying condition, and most preferably that one of the specified conditions is caused by the other specified condition.

[0030]

[0037] The terms "food," "food product," and "food composition" refer to a product or composition intended for consumption by an individual, such as a human, and providing at least one nutrient to such an individual. A food product typically comprises at least one of protein, lipid, carbohydrate, and optionally one or more vitamins and minerals. The compositions of the present disclosure, including the many embodiments described herein, can comprise, consist of, or consist essentially of the elements disclosed herein, as well as any additional or optional ingredients, components, or elements described herein or not described herein that are useful in the diet.

[0031]

[0038] As used herein, the term "isolated" means separated from one or more other compounds or components with which the compound may otherwise be found, e.g., in nature. Preferably, for example, "isolated" means that the specified compound is separated from at least a portion of the cellular material with which it is typically found in nature. In one embodiment, an isolated compound is pure, i.e., free of other compounds.

[0032]

[0039] As used herein, an "effective amount" is an amount that prevents a deficiency, treats a disease or medical condition of an individual, or more generally, alleviates symptoms, manages the progression of a disease, or provides a nutritional, physiological, or medical benefit to an individual. The relative terms "improved," "increased," "enhanced," and the like refer to the effect of the compositions disclosed herein. As used herein, "promoting" refers to enhancing or inducing compared to a value prior to administration of the compositions disclosed herein.

[0033]

[0040] As used herein, the term "unit dosage form" refers to a physically discrete unit suitable as a unit of administration for human and animal subjects, each unit containing a predetermined amount of the composition disclosed herein in an amount sufficient to produce the desired effect, preferably together with a pharma- ceutically acceptable diluent, carrier, or vehicle. The specifications of the unit dosage form depend on the specific compound used, the effect to be achieved, and the pharmacodynamics associated with each compound in the host. In some embodiments, the unit dosage form can be a predetermined amount of active compound in a serving of food product, a predetermined amount of active compound in a sachet, a predetermined amount of active compound in a capsule or tablet, or a predetermined amount of active compound in a predetermined amount of liquid, preferably a therapeutically or prophylactically effective amount, or a predetermined fraction of a therapeutically or prophylactically effective amount.

[0034]

[0041] A "subject" or "individual" is a mammal, preferably a human. The term "elderly", in relation to humans, means a postnatal age of at least 60 years, preferably greater than 63 years, more preferably greater than 65 years, and most preferably greater than 70 years. The term "older adult", in relation to humans, means a postnatal age of 45 years or older, preferably greater than 50 years, and more preferably greater than 55 years, and includes elderly individuals.

[0035]

[0042] As used herein, "frailty" is defined as a clinically recognizable state of increased vulnerability and impaired ability to cope with routine or acute stressors due to age-related declines in reserve and function across multiple physiological systems. The presence of one or two of these criteria identifies the pre-frailty stage as being at high risk for progression to frailty.

[0036]

[0043] "Overweight" is defined in humans as a body mass index (BMI) between 25 and 30 kg / m 2 "Obesity" is defined in humans as a BMI of at least 30 kg / m 2 , for example 30 to 39.9 kg / m 2 "Weight loss" is defined as a reduction in total body weight. Weight loss can refer to a reduction in total body weight to improve one or more of, for example, health, fitness, or appearance.

[0037]

[0044] "Diabetes" includes both types I and II of the disease. Non-limiting examples of risk factors for diabetes include a waistline greater than 40 inches for men or greater than 35 inches for women, blood pressure equal to or greater than 130 / 85 mmHg, triglycerides greater than 150 mg / dL, fasting blood glucose greater than 100 mg / dL, or high density lipoprotein less than 40 mg / dL for men or less than 50 mg / dL for women.

[0038]

[0045] As used herein, the term "metabolic syndrome" refers to a combination of medical abnormalities that, when occurring together, increase the risk of developing cardiovascular disease and diabetes. Metabolic syndrome affects one in five people in the United States, with prevalence increasing with age. Some studies suggest that the prevalence in the United States is estimated at 25% of the population. According to the International Diabetes Foundation's consensus worldwide definition (2006), metabolic syndrome is central obesity in combination with any two of the following:

[0046] Elevated triglycerides: greater than 150 mg / dL (1.7 mmol / L) or treatments targeted to this lipid abnormality;

[0047] Reduction of HDL cholesterol: less than 40 mg / dL (1.03 mmol / L) in men and less than 50 mg / dL (1.29 mmol / L) in women, or treatment targeted to this lipid abnormality;

[0048] Elevated blood pressure: systolic BP > 130 or diastolic BP > 85 mmHg, or treatment of previously diagnosed hypertension; and

[0049] Elevated fasting plasma glucose: (FPG) greater than 100 mg / dL (5.6 mmol / L) or previously diagnosed type 2 diabetes.

[0039]

[0050] As used herein, "neurodegenerative disease" or "neurodegenerative disorder" refers to any condition in the central nervous system that results in a gradual loss of functional neurons. In one embodiment, the neurodegenerative disease is associated with age-related cell death. Non-limiting examples of neurodegenerative diseases include Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (also known as ALS and Lou Gehrig's disease), AIDS dementia, adrenoleukodystrophy, Alexander disease, Alpers disease, ataxia-telangiectasia, Batten disease, bovine spongiform encephalopathy (BSE), Canavan disease, corticobasal degeneration, Creutzfeldt-Jakob disease, dementia with Lewy bodies, fatal familial insomnia, prefrontal cortex syndrome, and pulmonary tuberculosis. These include temporotemporal lobar degeneration, Kennedy disease, Krabbe disease, Lyme disease, Machado-Joseph disease, multiple sclerosis, multiple system atrophy, neuroacanthocytosis, Niemann-Pick disease, Pick's disease, primary lateral sclerosis, progressive supranuclear palsy, Refsum disease, Sandhoff disease, diffuse myelinating sclerosis, spinocerebellar ataxia, subacute combined spinal degeneration, tabes dorsalis, Tay-Sachs disease, toxic encephalopathy, transmissible spongiform encephalopathy, and wobbly hedgehog syndrome.

[0040] As used herein, "cognitive function" refers to any mental process that involves symbolic activity, such as perception, memory, attention, speech comprehension, speech production, reading, imagery creation, learning, and reasoning, preferably at least memory.

[0041]

[0051] Methods for measuring cognitive function are well known and can include, for example, individual tests or battery tests for any aspect of cognitive function. One such test is the Prudhoe Cognitive Function Test by Margallo-Lana et al. (2003) J. Intellect. Disability Res. 47:488-492. Another test is the Mini Mental State Exam (MMSE), which is designed to assess orientation to time and place, memory, attention and calculation, recall, language use and comprehension, repetition, and complex commands. As used herein, "cognitive impairment" refers to any condition that reduces cognitive function. Non-limiting examples of cognitive impairment include delirium, dementia, learning disabilities, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD). "Stress-induced or stressor-related cognitive dysfunction" refers to cognitive dysfunction that is stress-induced or related to stress.

[0042]

[0052] As used herein, "mood disorder" (also known as affective disorder) refers to a disturbance of emotional state as described in the Diagnostic and Statistical Manual of Mental Disorders published by the American Psychiatric Association. Non-limiting examples of mood disorders include major depression, postpartum depression, dysthymia, and bipolar disorder. "Stress-induced or stressor-related mood disorder" refers to a disturbance of emotional state that is stress-induced or stress-related. Such mood disorders are sometimes referred to as reactive mood disorders and are distinguished from other mood disorders, such as "organic" mood disorders, which are due to a medical or health condition rather than a psychiatric illness.

[0043]

[0053] As used herein, "anxiety disorder" refers to a dysfunction of fear and anxiety, e.g., fear and anxiety that is out of proportion to the stressful situation or the anticipation of a stressful situation. Non-limiting examples of anxiety disorders include generalized anxiety disorder, panic disorder, panic disorder with agoraphobia, agoraphobia, social anxiety disorder, obsessive-compulsive disorder, and post-traumatic stress disorder. "Stress-induced or stressor-related anxiety disorder" refers to a dysfunction of fear and anxiety that is stress-induced or stress-related. Such anxiety disorders are sometimes referred to as reactive anxiety disorders and are distinguished from other anxiety disorders, e.g., "organic" anxiety disorders, which are caused by a medical or health condition rather than a psychiatric illness.

[0044]

[0054] As used herein, "metabolic fatigue" refers to a decrease in mitochondrial function in one or more cells (e.g., one or more of the liver, kidney, brain, heart, intestine, pancreas, immune cells, or skeletal muscle cells).

[0045]

[0055] Embodiment Vitamin B12 (also known as cobalamin) is a group of cobalt-containing water-soluble vitamins that cannot be synthesized by the human body and therefore must be obtained from food or synthesis by the gut microbiota. The vitamin B12 group can refer to several chemical forms of vitamin B12, depending on the upper axial ligand of the cobalt ion. These are cyanocobalamin (R=-CN), hydroxocobalamin (R=-OH), methylcobalamin (R=-CH3), and adenosylcobalamin (R=-5'-deoxyadenosyl).

[0046]

[0056] The pool of vitamin B12 in the human body is composed of several forms: cyanocobalamin, which is inactive and requires conversion to become active, and methylcobalamin and adenosylcobalamin, which are metabolically active forms of vitamin B12.

[0047]

[0057] Two enzymes, methionine synthase and methylmalonyl-CoA mutase, are known to utilize vitamin B12 as a cofactor. Methionine synthase is an intracellular enzyme that utilizes methylcobalamin to convert homocysteine ​​to methionine. This conversion provides S-adenosylmethionine (SAM) as a methylation donor, playing an important role in preventing the toxic accumulation of homocysteine. Low SAM and high homocysteine ​​levels observed with severe vitamin B12 deficiency impair myelination of peripheral nerves and the spinal cord. Methionine synthase also catalyzes the activation of 5-methyl-tetrahydrofolate to the biologically active tetrahydrofolate, which is required for one-carbon metabolism and DNA synthesis, thus contributing to the increase in red blood cells. Methylmalonyl-CoA mutase is a mitochondrial enzyme that utilizes adenosylcobalamin to convert methyl-malonyl-CoA to succinyl-CoA, which is subsequently incorporated into the TCA cycle. It is essential for regulating neurodevelopment during embryonic life, but not in adulthood, as evidenced by the degradation of branched-chain amino acids and odd-chain fatty acids.

[0048]

[0058] The adenosylcobalamin of the present invention may be in the form of a semi-synthetic derivative.

[0049]

[0059] In another embodiment, the adenosylcobalamin can be hydroxocobalamin and / or cyanocobalamin, which can be converted to adenosylcobalamin.

[0050]

[0060] Vitamin B12 deficiency

[0061] In one embodiment, the subject may have a vitamin B12 deficiency.

[0051]

[0062] The Recommended Dietary Allowance (RDA) for US adults was set by the Institute of Medicine at 2.4 μg per day, based on an average absorption from food of approximately 50% (National Academy of Sciences, Institute of Medicine (2000); Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin and Choline, Chapter 9, pp306-56). Daily requirements have been shown to vary with body size.

[0052]

[0063] The likelihood of vitamin B12 deficiency in humans can be defined according to serum vitamin B12 concentrations as follows: less than 148 pmol / L (less than 200 pmol / mL) indicates possible deficiency, 148-258 pmol / L (201-350 pmol / mL) indicates possible deficiency, and more than 258 pmol / L (more than 350 pmol / mL) indicates no likelihood of deficiency (BMJ, Best Practice, http: / / bestpractice.bmj.com / best-practice / monograph / 822 / basics.html). However, due to the lack of a gold standard for determining vitamin B12 levels and related complications for active and inactive vitamin B12, serum vitamin B12 assays are often combined with further biochemical assays or clinical evaluation based on the main symptoms to diagnose vitamin B12 deficiency.

[0053]

[0064] Additional assays that can be performed to provide a further indication of vitamin B12 deficiency include measuring the levels of holotranscobalamin, methylmalonic acid and / or homocysteine ​​in a sample isolated from the subject.

[0054]

[0065] Holotranscobalamin refers to vitamin B12 bound to its bioactive serum transporter, transcobalamin II. Holotranscobalamin levels can be measured using commercially available assays (e.g., ELISA assays). Low holotranscobalamin levels are associated with subclinical vitamin B12 deficiency.

[0055]

[0066] Methylmalonic acid (MMA) accumulates due to decreased activity of the vitamin B12-dependent enzyme methylmalonyl-CoA mutase, and therefore high levels of MMA are associated with vitamin B12 deficiency.

[0056]

[0067] In one embodiment, the individual has high blood levels of methylmalonic acid.

[0057]

[0068] Homocysteine ​​accumulates due to decreased activity of the vitamin B12-dependent enzyme methionine synthase. Low and high levels of homocysteine ​​are associated with vitamin B12 deficiency. However, assays for homocysteine ​​levels can be confounded by folate deficiency.

[0058]

[0069] Adenosylcobalamin may be provided, for example, in the form of a tablet, liquid (e.g., for oral ingestion or use in a nasal spray or injection), or a transdermal patch. For example, adenosylcobalamin may be available as a dietary supplement, either by itself or in combination with other supplements.

[0059]

[0070] Oral supplementation typically involves the daily administration of 1-10 μg up to 100 μg-2000 μg of adenosylcobalamin depending on the form. When administered in the form of an oral dietary supplement, the daily dose provides 1-10 μg, preferably 1-2 μg, of adenosylcobalamin. When administered in the form of a supplement, the daily dose provides 100 μg-2000 μg, preferably 250 μg-1 mg, of adenosylcobalamin.

[0060]

[0071] The present invention may involve administering to a subject a probiotic supplement comprising adenosylcobalamin-producing bacteria.

[0061]

[0072] The probiotic supplement may comprise any probiotic microorganism(s) that beneficially impacts the host subject by improving the intestinal bacterial balance and enhancing vitamin B12 uptake in the host subject. The probiotic microorganism may be selected from the group comprising the genera Bifidobacterium, Lactobacillus, Streptococcus, Enterococcus and Saccharomyces, or mixtures thereof.

[0062]

[0073] Oral supplementation with adenosyl vitamin B12 can be in the form of a food or beverage product. The food or beverage product can include a probiotic supplement containing vitamin B12-producing bacteria or other probiotics that can enhance existing microorganisms in the gut that produce vitamin B12 in situ.

[0063]

[0074] Typically, a physician will determine the actual dosage that will be most appropriate for an individual subject, which will vary with the age, weight, and response of the particular patient. The dosage is sufficient to provide the required amount of active adenosyl vitamin B12.

[0064] method

[0075] Without being bound by theory, it is believed that various types of stress cause mitochondrial stress damage, which reduces the ability of mitochondria to perform many roles essential to overall cellular function.The methods disclosed herein can be useful for treating conditions involving stress damage to mitochondria.This damage can be manifested in any of a number of ways, including but not limited to mitochondrial disease.

[0065]

[0076] Mitochondrial diseases arise when the function of proteins or RNA molecules normally present in mitochondria is altered, either by genetic or spontaneous mutations in mitochondrial or nuclear DNA. However, problems in mitochondrial function may only affect certain tissues, due to factors occurring during development and growth that are not yet fully understood. Even when considering tissue-specific isoforms of mitochondrial proteins, it is difficult to explain the variable patterns of organ systems affected in clinical mitochondrial disease syndromes.

[0066]

[0077] Mitochondrial diseases result from damage to mitochondria, specialized compartments present in all cells of the body except red blood cells. Mitochondria are responsible for producing over 90% of the energy the body needs to sustain life and support development. When mitochondria stop functioning, less energy is produced in cells. Cells become damaged and even die. When this process is repeated throughout the body, the whole system begins to break down, which severely threatens the person's life. Mitochondrial diseases primarily affect children, but are increasingly being seen in adults as well.

[0067]

[0078] Mitochondrial diseases are most likely to cause damage to cells of the brain, heart, liver, skeletal muscle, kidneys, and the endocrine and respiratory systems.

[0068]

[0079] Many symptoms in mitochondrial disease are non-specific. Symptoms may also show a temporary course with periodic exacerbations. A review article on mitochondrial medicine mentions episodic migraine states, as well as muscle pain, gastrointestinal symptoms, tinnitus, depression, chronic fatigue, and diabetes, among the various manifestations of mitochondrial disease. In patients with mitochondrial disease, clinical symptoms typically occur when energy demands are high in association with physiological stressors such as illness, hunger, excessive exercise, and extreme environmental temperatures. In addition, psychological stressors often trigger symptoms, perhaps due to the brain's energy demands being so high that the patient cannot perform sufficient ATP production.

[0069]

[0080] Depending on which cells are affected, symptoms can include loss of motor control, muscle weakness and pain, gastrointestinal problems and difficulty swallowing, poor growth, heart disease, liver disease, diabetes, respiratory complications, seizures, visual / hearing problems, lactic acidosis, developmental delays, and susceptibility to infections.

[0070]

[0081] Mitochondrial disorders include, but are not limited to, Alpers disease, Barth syndrome, beta-oxidation disorders, carnitine deficiency, carnitine-acyl-carnitine deficiency, chronic progressive external ophthalmoplegia syndrome, coenzyme Q10 deficiency, complex I deficiency, complex II deficiency, complex III deficiency, complex IV deficiency, complex V deficiency, CPT I deficiency, CPT II deficiency, creatine deficiency syndrome, cytochrome c oxidase deficiency, glutaric aciduria type 2, Kearns-Sayre syndrome, lactic acidosis, LCHAD (long-chain acyl-CoA dehydrogenase deficiency), Leber's hereditary optic neuropathy, Leigh disease, lethal infantile cardiomyopathy, cardiomyopathy, Luft's disease, MAD (medium-chain acyl-CoA dehydrogenase deficiency), mitochondrial cytopathy, mitochondrial DNA depletion, mitochondrial encephalomyopathy, lactic acidosis, and stroke-like symptoms, mitochondrial encephalopathy, mitochondrial myopathy, mitochondrial recessive ataxia syndrome, muscular dystrophies, myoclonic epilepsy and ragged-red fiber disease, myoneurogenic gastrointestinal encephalopathy, neuropathy, ataxia, retinitis pigmentosa, and ptosis, Pearson's syndrome, POLG mutations, pyruvate carboxylase deficiency, pyruvate dehydrogenase deficiency, SCHAD (short-chain acyl-CoA dehydrogenase deficiency), and very long-chain acyl-CoA dehydrogenase deficiency.

[0071]

[0082] Thus, one aspect of the present disclosure is a composition in unit dosage form comprising adenosylcobalamin in an amount effective for the treatment or prevention of at least a condition selected from the group consisting of stress (e.g., early life stress and / or its effects), obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stressor-related cognitive dysfunction, mood disorder (e.g., stress-induced or stressor-related mood disorder), anxiety disorder (e.g., stress-induced or stressor-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not due to a specific neurodegenerative disease), trauma, infection (e.g., ICU), or cancer.

[0072]

[0083] Another aspect of the present disclosure is a method for treating at least one condition in an individual having at least one condition selected from the group consisting of stress (e.g., early life stress and / or its effects), obesity, metabolic rate reduction, metabolic syndrome, diabetes mellitus, cardiovascular disease, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stressor-related cognitive dysfunction, mood disorder (e.g., stress-induced or stressor-related mood disorder), anxiety disorder (e.g., stress-induced or stressor-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not caused by a specific neurodegenerative disease), trauma, infection (e.g., ICU), or cancer. The method comprises administering to the individual a composition comprising a therapeutically effective amount of adenosylcobalamin.

[0073]

[0084] A further aspect of the present disclosure is a method for preventing at least one condition selected from the group consisting of stress, obesity, metabolic rate reduction, metabolic syndrome, diabetes mellitus, cardiovascular disease, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stress-related cognitive dysfunction, mood disorder (e.g., stress-induced or stress-related mood disorder), anxiety disorder (e.g., stress-induced or stress-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not caused by specific neurodegenerative disease), trauma, infection (e.g., in ICU), or cancer. The method comprises administering a composition comprising a prophylactically effective amount of adenosylcobalamin to an individual at risk of at least one condition.

[0074]

[0085] In one embodiment of these methods, the hyperlipidemia treated or prevented comprises hypertriglyceridemia. In one embodiment of these methods, the hyperlipidemia treated or prevented comprises an increase in free fatty acids. In one embodiment of these methods, the age-related neuronal cell death or dysfunction treated or prevented is by administration of the composition to a middle-aged or elderly person, such as an elderly person.

[0075]

[0086] The stress to be treated or prevented can be early life stress, i.e. stress experienced during the period from birth to 5 years of age. Early life stress has been reported to have a significant adverse effect on cognitive performance, including psychological parameters such as increased incidence or susceptibility to depression, anxiety, and abnormal risk-taking behavior. Higher incidence of attention deficit / hyperactivity disorder (ADHD), post-traumatic stress disorder (PTSD), and major depression has been reported in individuals who have experienced early life stress.

[0076]

[0087] Another aspect of the present disclosure is a method of delaying the onset of metabolic decline, reducing oxidative stress, maintaining immune function, and / or maintaining cognitive function in a healthy middle-aged or elderly individual, comprising administering to the healthy middle-aged or elderly individual an effective amount of adenosylcobalamin.

[0077]

[0088] Another aspect of the present disclosure is a method of improving mitochondrial function in an individual, such as a middle-aged or elderly individual, comprising administering to the individual an effective amount of adenosylcobalamin.

[0078]

[0089] Yet another aspect of the present disclosure is a method of enhancing metabolism of reactive oxygen species and improving glucose control in an individual having at least one of obesity or diabetes, comprising administering to the individual an effective amount of adenosylcobalamin.

[0079]

[0090] Another aspect of the present disclosure is a method of improving mitochondrial function (preferably benefiting at least one of metabolism or muscle strength) in an individual, such as a middle-aged or elderly individual, comprising administering an effective amount of adenosylcobalamin to the individual.

[0080]

[0091] Yet another aspect of the present disclosure is a composition comprising adenosylcobalamin in an amount effective for weight management. "Weight management" for adults (e.g., at least 18 years after birth) means that the individual has about the same body mass index (BMI) after 1 week of taking the composition, preferably after 1 month of taking the composition, more preferably after 1 year of taking the composition, compared to the BMI when the individual started taking the composition. "Weight management" for young individuals means that the BMI is about the same percentile for individuals of the corresponding age after 1 week of taking the composition, preferably after 1 month of taking the composition, more preferably after 1 year of taking the composition, compared to the BMI percentile when the individual started taking the composition. In some embodiments, the individual undergoing weight management is an overweight individual who is preventing obesity.

[0081]

[0092] In a related embodiment, a method for weight management in an individual comprises administering to the individual a composition comprising an effective amount of adenosylcobalamin.

[0082]

[0093] The composition can improve physical endurance (e.g., the ability to perform physical tasks such as exercise, physical labor, sports activities, etc.), inhibit or delay physical fatigue, increase blood oxygen levels, enhance energy in healthy individuals, enhance work capacity and endurance, reduce muscle fatigue, improve recovery from exercise, reduce stress, enhance cardiomyocyte function, improve sexual performance, increase muscle ATP levels, and / or reduce blood lactate. "Endurance" refers to the time to fatigue when exercising at a given workload, typically at an intensity less than 80% of V02max. In some embodiments, the composition is administered in an amount that increases mitochondrial activity, increases mitochondrial biogenesis, and / or increases mitochondrial mass.

[0083]

[0094] A further aspect of the present disclosure is a composition comprising adenosylcobalamin in an amount effective to increase or maintain at least one of mitochondrial function or metabolic rate. In a related embodiment, a method of increasing or maintaining at least one of mitochondrial function or metabolic rate in an individual comprises administering to the individual a composition comprising an effective amount of adenosylcobalamin.

[0084]

[0095] Yet another aspect of the present disclosure is a composition in unit dosage form, comprising adenosylcobalamin in an amount effective for treating, preventing or managing at least one of mitochondrial-associated disease, conditions associated with altered mitochondrial function or reduced mitochondrial density.In a related embodiment, a method for treating an individual with at least one of mitochondrial-associated disease, conditions associated with altered mitochondrial function or reduced mitochondrial density comprises administering to the individual a composition comprising an effective amount of adenosylcobalamin.In another related embodiment, a method for preventing at least one of mitochondrial-associated disease, conditions associated with altered mitochondrial function or reduced mitochondrial density in an individual at risk comprises administering to the individual a composition comprising an effective amount of adenosylcobalamin.

[0085]

[0096] Another aspect of the present disclosure is a composition in unit dosage form comprising adenosylcobalamin in an amount effective to improve or maintain cognitive function. In a related embodiment, a method of improving or maintaining cognitive function in an individual comprises administering to the individual a composition comprising adenosylcobalamin.

[0086]

[0097] In one embodiment, the individual does not have a cognitive disorder, for example, the composition can enhance cognitive function in a subject with normal cognitive function.

[0087]

[0098] The compositions disclosed herein can also be used to treat any of a variety of additional diseases and conditions in which defective or reduced mitochondrial activity is involved in the pathophysiology of the disease or condition, or in which increased mitochondrial function produces the desired beneficial effect.Non-limiting examples of such conditions include male infertility associated with reduced sperm motility, macular degeneration, and other age-related and genetic eye disorders and hearing loss (e.g., age-related hearing loss).

[0088]

[0099] In each of the compositions and methods disclosed herein, adenosylcobalamin can be administered in a composition that is preferably a food product, including a food additive, a food ingredient, a functional food, a dietary supplement, a medical food, a nutraceutical, or a dietary supplement.

[0089] Dietary interventions and products

[0100] The term "dietary intervention" refers to an external agent given to a subject that results in a change in the subject's diet. In one embodiment, the dietary intervention is a high-calorie diet. In another embodiment, the dietary intervention is a high-protein and / or high-carbohydrate diet. In another embodiment, the dietary intervention is a diet supplemented with vitamins and minerals.

[0090]

[0101] In a preferred embodiment, the dietary intervention is a diet supplemented with adenosylcobalamin.

[0091]

[0102] In another preferred embodiment, the dietary intervention is a diet supplemented with vitamin B12, in particular hydroxocobalamin and / or cyanocobalamin, which can be converted to adenosylcobalamin.

[0092]

[0103] The diet may be adjusted for the subject's initial weight.

[0093]

[0104] Dietary intervention may include the administration of at least one dietary product. Dietary product may be, for example, a meal replacement product or a nutritional supplement product that may enhance the appetite of a subject. Dietary product may include food products, beverages, pet food products, dietary supplements, nutraceuticals, food additives, or nutritional formulas. Examples of oral nutritional supplements include Boost and Meritene, Nestle products.

[0094]

[0105] In one embodiment, the composition further comprises a medium chain triglyceride, such as one or more of caproic acid, caprylic acid, capric acid, and lauric acid. In one embodiment, the composition further comprises a phospholipid, such as a phosphatidylcholine.

[0095]

[0106] In one embodiment, the composition further comprises a protein source, preferably a purified protein (i.e. isolated from the natural food component in which the protein is produced). The protein content of the composition is preferably from 20 to 99% by weight of the composition, such as from 20 to 90% by weight of the composition, for example from 30 to 80% by weight of the composition, such as from 40 to 80% by weight of the composition, for example from 50 to 80% by weight, for example from 40 to 70% by weight of the composition.

[0096]

[0107] Non-limiting examples of proteins or sources thereof suitable for use in the composition include hydrolyzed, partially hydrolyzed, or non-hydrolyzed proteins or protein sources. These may be derived from any known or other suitable source, such as milk (e.g., casein, whey), animal (e.g., meat, fish), grain (e.g., rice, corn), or vegetable (e.g., soy, pea). Combinations of sources or proteins may be used. Non-limiting examples of proteins or sources thereof include intact pea protein, intact pea protein isolate, intact pea protein concentrate, milk protein isolate, milk protein concentrate, casein protein isolate, casein protein concentrate, whey protein concentrate, whey protein isolate, sodium or calcium caseinate, whole milk, partially or fully skimmed milk, yogurt, soy protein isolate, and soy protein concentrate, and combinations thereof. Combinations of sources or proteins may be used. Preferred proteins include pea protein, whey protein, soy protein, and casein. Casein proteins may include, for example, sodium caseinate and calcium caseinate.

[0097]

[0108] The protein source may be provided by individual amino acids, polypeptides containing amino acids, or mixtures thereof. In many muscle growth, muscle maintenance, and / or muscle building treatments, the specific amino acids that are beneficial are, for example, L-arginine, L-glutamine, lysine, and branched chain amino acids (i.e., leucine, isoleucine, and valine; particularly leucine and isoleucine). These specific amino acids may be provided as protein sources or may be added to the main source of protein. Thus, the protein source in the composition may include one or more branched chain amino acids (leucine, isoleucine, and valine), one or both of L-arginine and L-glutamine, and lysine. In a preferred embodiment, the composition includes whey protein and / or casein protein together with one or more individual amino acids, for example, one or more (or all) of leucine, isoleucine, and L-arginine.

[0098]

[0109] The composition may be administered at least 1 day per week, preferably at least 2 days per week, more preferably at least 3 or 4 days per week (e.g., every other day), most preferably at least 5 days per week, 6 days per week, or 7 days per week. The administration period may be at least 1 week, preferably at least 1 month, more preferably at least 2 months, most preferably at least 3 months, e.g., at least 4 months. In one embodiment, administration is at least daily, e.g., the subject may be administered one or more times per day. In some embodiments, administration continues for the remainder of the individual's life. In another embodiment, administration is administered until there is no detectable symptom of the medical condition. In certain embodiments, administration is administered until there is a detectable improvement in at least one symptom, and in further cases, administration is continued to maintain remission.

[0099]

[0110] The compositions disclosed herein can be administered to a subject orally, enterally, or parenterally.Non-limiting examples of parenteral administration include intravenous, intramuscular, intraperitoneal, subcutaneous, intraarticular, intrasynovial, intraocular, intrathecal, topical, and by inhalation.Non-limiting examples of the form of the composition therefore include natural foods, processed foods, natural fruit juices, concentrates and extracts, injections, microcapsules, nanocapsules, liposomes, plasters, inhalation forms, nasal sprays, nasal drops, eye drops, sublingual tablets, and sustained release formulations.

[0100]

[0111] The compositions disclosed herein may be used in any of a variety of formulations for therapeutic administration. More specifically, the pharmaceutical compositions may include suitable pharma-ceutically acceptable carriers or diluents and may be formulated as solid, semi-solid, liquid or gaseous formulations, such as tablets, capsules, powders, granules, ointments, liquids, suppositories, injections, inhalants, gels, microspheres and aerosols. Thus, administration of the compositions may be accomplished in a variety of ways, including oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal and intratracheal administration. The active agent may be systemic after administration or may be localized by the use of topical administration, intramural administration or the use of an implant that acts to retain an effective dose at the implantation site.

[0101]

[0112] In pharmaceutical dosage forms, the compounds may be administered as pharma- ceutically acceptable salts. They may also be used in appropriate association with other pharma- ceutically active compounds. The following methods and additives are merely illustrative and in no way limiting.

[0102]

[0113] In oral formulations, the compounds can be used alone or in combination with suitable additives for producing tablets, powders, granules or capsules, for example, conventional additives such as lactose, mannitol, corn starch or potato starch, binders such as crystalline cellulose, functional derivatives of cellulose, gum arabic, corn starch or gelatin, disintegrants such as corn starch, potato starch or sodium carboxymethylcellulose, lubricants such as talc or magnesium stearate, and, if desired, diluents, buffers, wetting agents, preservatives and flavouring substances.

[0103]

[0114] The compounds can be formulated as injectable preparations by dissolving, suspending or emulsifying in aqueous or non-aqueous solvents, such as vegetable oils or other similar oils, synthetic aliphatic acid glycerides, esters of higher aliphatic acids, or propylene glycol, together with conventional additives, such as solubilizers, isotonicity agents, suspending agents, emulsifying agents, stabilizers, and preservatives, if desired.

[0104]

[0115] The compounds can be utilized in aerosol formulations to be administered via inhalation. For example, the compounds can be placed into pressurized acceptable propellants, such as dichlorodifluoromethane, propane, and nitrogen.

[0105]

[0116] In addition, the compound can be prepared as a suppository by mixing with various bases such as emulsifying bases or water-soluble bases.The compound can be administered rectally by suppository.The suppository can include vehicles such as cocoa butter, carbowax, and polyethylene glycol, which melt at body temperature but solidify at room temperature.

[0106]

[0117] Unit dosage forms for oral or rectal administration, such as syrups, elixirs, and suspensions, may be provided, with each dosage unit, e.g., teaspoon, tablespoon, tablet, or suppository, containing a predetermined amount of the composition. Similarly, unit dosage forms for injection or intravenous administration may contain the compound in a composition as a solution in sterile water, saline, or another pharma- ceutically acceptable carrier, with each dosage unit, e.g., mL or L, containing a predetermined amount of a composition containing one or more of the compounds.

[0107]

[0118] [Example]

[0119] The following non-limiting examples present scientific data that develops and supports the concept that administration of adenosylcobalamin increases cellular energy production, thereby improving the function of various tissues.

[0108]

[0120] Example 1

[0121] Materials and Methods

[0122] To test the effect of adenosyl-B12 and methyl-B12 on human myotubes, oxygen consumption rate and ATP production were measured in human myotubes differentiated from primary adult muscle cells (HSMM). HSMM were purchased from Lonza (https: / / bioscience.lonza.com). HSMM were isolated from muscle tissue of the upper arm or leg of healthy donors and used after the second passage. HSMM were seeded in SKM-M medium (ZenBio) in 96-well plates at a density of 12000 cells per well. Myotubes were differentiated from HSMM by growing the cells for 4 days in DMEM / F-12 (Gibco) containing 2% horse serum. From the second day of growth, the medium was B12-free.

[0109]

[0123] Oxygen consumption was measured using an XF96 instrument (Seahorse Biosciences, North Billerica, MA, USA). After differentiation, respiration rates were measured every 7 min at 37°C. Myotubes were stimulated with 10 μM epibatidine. Oligomycin (2.5 μg / ml) was then added to measure the ATP synthase-dependent component of respiration. ATP synthase-dependent respiration was calculated as the difference in respiration rate before and after the addition of oligomycin.

[0110]

[0124] ATP measurements were performed using myotubes infected with adenovirus expressing luciferase (from Sirion biotech). Luminescence was measured with a Cytation 3 cell imaging reader (Biotek). 48 h after infection, relative changes in ATP were measured with a luminometer in standard medium containing 145 mM NaCl, 5 mM KCl, 1 mM MgCl2, 1 mM CaCl2, 10 mM glucose and 10 mM Hepes, pH 7.4. Luciferin (5 μM) was added to drive ATP-dependent responses and basal luminescence was normalized as 100%. ATP production in myotubes was stimulated by adding epibatidine.

[0111]

[0125] For treatment, compounds were added directly to cell or myotube cultures 3 h (acute treatment) or 3 days (chronic treatment) before measurements. Quantification was performed using a custom module analysis based on Excel (Microsoft) and GraphPad Prism 7.02 (GraphPad) software.

[0112]

[0126] result

[0127] As shown in Figure 1, adenosyl B12 increases the ATP synthase-dependent component of respiration in stimulated human skeletal myotubes, whereas methyl B12 (Figure 2) does not increase the ATP synthase-dependent component of respiration in stimulated human skeletal myotubes.

[0113]

[0128] Similarly, as shown in FIG. 3, adenosyl B12, but not methyl B12, enhances epibatine-stimulated ATP production in human skeletal myotubes during acute treatment.

[0114]

[0129] Furthermore, chronic treatment of human skeletal myotubes with adenosyl B12, but not methyl B12, enhances epibatine-stimulated ATP production (Figure 4).

[0115]

[0130] Example 2

[0131] Materials and Methods

[0132] Male Wistar rats aged 3 or 19 months were treated with adenosylcobalamin (adenosyl B12) or methylcobalamin (methyl-B12) at a dose of 1 mg / kg by subcutaneous injection three times a week for 5 months. Control animals were injected with the same volume of saline. After 5 months of treatment, adult rats were defined as 8-month-old rats, and aged (old) animals were defined as 24-month-old rats. months was defined as aged rodents.

[0116]

[0133] Total RNA was extracted from tibialis anterior using Agencourt RNAdvance tissue kit. For gene set enrichment and network analysis, RNA quantity was measured with Ribogreen and RNA quality was confirmed with fragment analyzer using standard sensitivity RNA analysis kit. All target cRNA was synthesized using IVT+ kit and fragmented according to Affymetrix protocol based on Eberwine T7 procedure. Briefly, 100 ng total RNA was used to generate double-stranded cDNA, followed by in vitro transcription and biotin labeling of cRNA. Gene set enrichment analysis (GSEA) of skeletal muscle shows significant differentially expressed pathways (FIG. 5A). In network analysis (FIG. 5B), nodes with interaction scores >0.9 are represented in different grey levels and aggregated based on biological function.

[0117]

[0134] To measure muscle performance (Figure 6), coordinated movement was measured using a rotarod apparatus. Rats were placed on the rotating rod and the rotation speed was gradually increased from 4 rpm to 40 rpm over 600 s until the rat could no longer cope with the rotation and was dropped onto a protective pad placed under the rod. Activity time on the rotarod is measured in seconds. Each animal was recorded three times with a rest period of at least 10 min between each trial.

[0118]

[0135] result

[0136] As shown in Figures 5-1 and 5-2, adenosyl B12 specifically displays a gene expression signature of oxidative phosphorylation genes in skeletal muscle. Enrichment plot analysis of the oxidative phosphorylation gene set (Panel 5A) comparing aged rats treated in vivo with adenosyl B12 versus aged rats treated with methyl B12 indicates that mitochondrial energy production is a strong transcriptional signature in adenosyl B12-treated skeletal muscle. Furthermore, network analysis of regulated genes (Panel 5B) distinguished several clusters associated with mitochondrial respiratory chain complexes, oxidative phosphorylation, and mitochondrial function only in rats treated specifically with adenosyl B12.

[0119]

[0137] Furthermore, specific in vivo treatment of aged rats with adenosyl B12 improves skeletal muscle performance, as evidenced by a significant increase in active time on the rod.

[0120]

[0138] It should be understood that various changes and modifications to the presently preferred embodiments described herein will be apparent to those skilled in the art. Such changes and modifications can be made without departing from the spirit and scope of the subject matter of the present invention and without diminishing its intended advantages. It is therefore intended that such changes and modifications be encompassed within the scope of the appended claims.

Claims

1. 1. A composition for use in a method of increasing mitochondrial energy in one or more cells in an individual, the one or more cells being part of a skeletal muscle, A composition comprising adenosylcobalamin.

2. 2. The composition of claim 1, wherein the adenosylcobalamin is administered in an amount of 1-10 μg up to 100 μg-2000 μg per day.

3. 3. The composition of claim 1 or 2, for oral administration.

4. The composition of any one of claims 1 to 3, wherein the individual has an elevated blood concentration of methylmalonic acid.

5. The composition of any one of claims 1 to 4, wherein the individual is selected from the group consisting of middle-aged and elderly people, elderly people, critically ill patients, and ICU patients.

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