Compositions and methods using a combination of nicotinamide and glycine

Abstract

The present disclosure generally relates to compositions and methods to increase resistance to age-related pathologies, improve physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; improve mobility and / or healthspan and / or lifespan of an adult animal in need. More specifically, the present disclosure relates to administering a formulation comprising an effective amount of nicotinamide (NAM) and Glycine, optionally an autophagy inducer.

Description

TITLECOMPOSITIONS AND METHODS USING A COMBINATION OF NICOTINAMIDE AND GLYCINETECHNICAL FIELD

[0001] The present disclosure generally relates to compositions and methods to increase resistance to age-related pathologies, improve physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; improve mobility and / or healthspan and / or lifespan of an adult animal in need. More specifically, the present disclosure relates to administering a formulation comprising an effective amount of nicotinamide (NAM), Glycine and optionally an autophagy inducer.BACKGROUND

[0001] Population aging has been a remarkable demographic event. As the growth of the older population has outpaced the total population due to increased longevity, the proportion of older persons relative to the rest of the population has increased considerably due to decreased fertility rates. For example, one in every twelve individuals was at least 60 years of age in 1950, and one in every ten was aged 60 years or older by the end of 2000. By the end of 2050, the number of persons worldwide that is 60 years or over is projected to be one in every five.

[0002] Mitochondrial dysfunction, oxidative stress, altered intercellular communication (including chronic low-grade inflammation), genomic instability, telomere attrition, loss of proteostasis, altered nutrient sensing, epigenetic alterations, and stem cell exhaustion have been proposed as hallmarks of aging. Moreover, free radicals - reactive oxygen species (ROS)- are the main origin of aging by causing oxidative cellular injuries. Free radicals are necessary for many biochemical processes and they are produced as by-products during some biochemical reactions or as substrates for other biochemical reactions in each cell.

[0003] As mitochondria are the principal source of intracellular reactive oxygen species (ROS), this hypothesis suggested a central role for the mitochondrion in normal mammalian aging. In recent years, however, much work has questioned the importance of mitochondrial ROS in driving aging. Conversely new evidence points to other facets of mitochondrial dysfunction whichmay nevertheless suggest the mitochondrion retains a critical role at the center of a complex web of processes leading to cellular and organismal aging.

[0004] Aged or aging individuals frequently suffer some degree of physical decline and / or cognitive impairment, including decline in cognitive function, that progresses with age, and age-related changes in brain morphology and cerebrovascular function are commonly observed. Cognitive decline has been consistently reported with aging across a range of cognitive domains including processing speed, attention, episodic memory, spatial ability and executive function.

[0005] Currently, there are a range of methods that have been demonstrated to improve healthy ageing including dietary restriction (DR), the reduction of food intake without malnutrition. However, these dietary regimes are very difficult to maintain and follow, so research has focused on targeting the mechanisms underpinning this response.

[0006] Thus, there is a strong need to provide compositions and methods that can boost mitochondrial function and increase bioenergetics, and thereby treat or prevent conditions and diseases linked to mitochondrial dysfunction.SUMMARY

[0007] As detailed in the experimental data set forth later herein, the inventors surprisingly found that the combination of compounds disclosed herein is able to promote cellular activation by boosting mitochondrial function and increasing bioenergetics and improve lifespan and / or mobility.

[0008] It could support mitochondrial function in oxidative stress conditions, such as aging, exercise, musculo-skeletal diseases, respiratory diseases, pain syndromes, neurodegenerative diseases and metabolic diseases. In particular by reducing oxidative stress and improving mitochondrial function, the present composition can treat, reduce incidence of, or reduce severity of metabolic and degenerative diseases at least with a combined effect, possibly potentiating each other or providing synergy.

[0009] Reduction of oxidative stress and improvement of mitochondrial function are mechanistically linked. Mitochondrial dysfunction contributes to cellular damage, partially through reactive oxygen species (ROS) and metabolic derangements, by not being able to metabolize nutrients, in turn leading to metabolic and degenerative diseases.

[0010] Without being bound by theory, the present inventors believe that a composition according to the present invention improves mitochondrial function, which in turn can increase muscle energy metabolism to thereby improve cellular energy and mobility.

[0011] Accordingly, in a general embodiment, the present disclosure provides a composition comprising a combination of Nicotinamide and glycine, optionally further comprising an autophagy inducer, such as spermidine in an effective amount for use in achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

[0012] The present invention also relates to a composition comprising a combination of Nicotinamide (NAM) and Glycine, optionally an autophagy inducer, such as spermidine in an effective amount for use in delaying off-set of metabolic decline, decreasing oxidative stress, mitigating deleterious effects of aging, improving or maintaining muscle mass and function, improving at least one of muscle performance or muscle recovery from exercise, exercise capacity and / or physical function, improving immune function and / or cognitive function in an individual and / or reducing severity of metabolic and / or degenerative diseases in an individual.

[0013] The present invention also relates to a method of manufacturing a nutritional composition for use according to any of the preceding claims, comprising the steps of providing one or more ingredients for a nutritional composition, Nicotinamide (NAM) and Glycine, optionally an autophagy inducer and mixing.

[0014] The present invention also relates to a method of achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iv) increasing resistance to age-related pathologies; (v) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (vi) improving mobility and / or (vii) improving healthspan and / or lifespan in an individual, the method comprising orally administering to the individual in need thereof or at risk thereof an effectiveamount of the combination of Nicotinamide (NAM) and Glycine, optionally an autophagy inducer.

[0015] The present invention also relates to a method of treating in an individual in need thereof or preventing in an individual at risk thereof at least one condition selected from the group consisting of (i) impairment in at least one of muscle functionality, muscle performance, lean muscle mass or muscle strength, (ii) muscle fatigue or muscle weakness, (iii) pre-frailty, frailty, sarcopenia or impaired mobility, and (iv) a muscle disorder linked to calcium depletion or deficiency, the method comprising orally administering to the individual in need thereof or at risk thereof an effective amount of the combination of Nicotinamide (NAM) and Glycine, optionally an autophagy inducer.

[0016] The present invention also relates to a unit dosage form comprising a combination of Nicotinamide (NAM) and Glycine, optionally an autophagy inducer, the unit dosage form comprises an amount of the combination effective for at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

[0017] In an embodiment, the unit dosage form of consists essentially of the combination of Nicotinamide (NAM) and Glycine, optionally an autophagy inducer, such as spermidine.

[0018] Additional features and advantages are described herein and will be apparent from the following Detailed Description.BRIEF DESCRIPTION OF THE FIGURES

[0019] FIG. 1 is a graph showing the effect of glycine, NAM and their combination on mitochondrial respiration in the EDL (A) and the Soleus (B) muscles of old mice. Mitochondrial respiration analysis of mouse soleus and EDL muscle biopsies, respectively (A, B). Leak refers to the oxygen consumption measured after the addition of pyruvate, malate, and glutamate, which initiate the electron transport chain by fueling Complex I. CI indicates the addition of ADP to stimulate ATP production, reflecting active respiration linked to Complex I. CII refers tothe addition of succinate, which fuels Complex II. ETS indicates the addition of FCCP, which dissipates the proton gradient across the inner mitochondrial membrane, allowing for the measurement of the maximum electron transport system (ETS) capacity. ETS+CII involves the addition of rotenone to inhibit Complex I, thereby isolating Complex II activity. Statistical analysis was performed using two-way ANOVA followed by Dunnet’s post-hoc test for all panels, only significant p-values (<0.05) are reported in the graphs.

[0020] FIG. 2 is a graph showing the effect of each single ingredient glycine, spermidine and NAM (A-C) and in combination (Combo) (D) on lifespan in C. elegans measured in the NAGI microfluidics system. Data is a pool of three independent experiments, including on average 50-100 worms per condition. Data were analyzed using log-rank (Mantel Cox) test.

[0021] FIG. 3 is a graph showing the effect of each single ingredient glycine, spermidine and NAM and in combination on mobility in C. elegans measured in the NAGI microfluidics system. Data is representative of three independent experiments for velocity of worm movement (A-C) and the area under de curve (AUC) over the first 10 days of life was pooled for the three experiments (D) including on average 50-100 worms per condition. Data were analyzed using two-way ANOVA followed by Bonferroni’s post-test (A-C) and one-way ANOVA followed by Dunnet’s post-test (D). * P<0.05.DETAILED DESCRIPTION

[0022] Definitions

[0023] Some definitions are provided hereafter. Nevertheless, definitions may be located in the “Embodiments” section below, and the above header “Definitions” does not mean that such disclosures in the “Embodiments” section are not definitions.

[0024] All percentages expressed herein are by weight of the total weight of the composition unless expressed otherwise. As used herein, “about,” “approximately” and “substantially” are understood to refer to numbers in a range of numerals, for example the range of -10% to +10% of the referenced number, preferably -5% to +5% of the referenced number, more preferably -1% to +1% of the referenced number, most preferably -0.1% to +0.1% of the referenced number. All numerical ranges herein should be understood to include all integers, whole or fractions, within the range. Moreover, these numerical ranges should be construed as providing support for a claim directed to any number or subset of numbers in that range. For example, a disclosure of from 1 to10 should be construed as supporting a range of from 1 to 8, from 3 to 7, from 1 to 9, from 3.6 to 4.6, from 3.5 to 9.9, and so forth.

[0025] 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 metabolite” or “the metabolite” includes one metabolite but also two or more metabolites.

[0026] The words “comprise,” “comprises” and “comprising” are to be interpreted inclusively rather than exclusively. Likewise, the terms “include,” “including” and “or” should all be construed to be inclusive, unless such a construction is clearly prohibited from the context. Nevertheless, the compositions disclosed herein may lack any element that is not specifically disclosed herein. Thus, a disclosure of an embodiment using the term “comprising” includes a disclosure of embodiments “consisting essentially of’ and “consisting of’ the components identified.

[0027] 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 “both X and Y.”

[0028] Where used herein, the terms “example” and “such as,” particularly when followed by a listing of terms, are merely exemplary and illustrative and should not be deemed to be exclusive or comprehensive. As used herein, “associated with” and “linked with” mean occurring concurrently, preferably means caused by the same underlying condition, and most preferably means that one of the identified conditions is caused by the other identified condition.

[0029] The terms “food,” “food product” and “food composition” mean a product or composition that is intended for ingestion by an individual such as a human and provides at least one nutrient to the individual. 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 otherwise useful in a diet.

[0030] As used herein, the terms “treat” and "treatment" mean to administer a composition as disclosed herein to a subject having a condition in order to lessen, reduce or improve at least one symptom associated with the condition and / or to slow down, reduce or block the progression of the condition. The terms “treatment” and “treat” include both prophylactic or preventivetreatment (that prevent and / or slow the development or progression of a targeted pathologic condition or disorder) and curative, therapeutic or disease-modifying treatment, including therapeutic measures that cure, slow down, lessen symptoms of, and / or halt progression of a diagnosed pathologic condition or disorder; and treatment of patients at risk of contracting a disease or suspected to have contracted a disease, as well as patients who are ill or have been diagnosed as suffering from a disease or medical condition. The terms “treatment” and “treat” do not necessarily imply that a subject is treated until total recovery. The terms “treatment” and “treat” also refer to the maintenance and / or promotion of health in an individual not suffering from a disease but who may be susceptible to the development of an unhealthy condition. The terms “treatment” and “treat” are also intended to include the potentiation or otherwise enhancement of one or more primary prophylactic or therapeutic measures. As non-limiting examples, a treatment can be performed by a patient, a caregiver, a doctor, a nurse, or another healthcare professional.

[0031] Both human and veterinary treatments are within the scope of the present disclosure. Preferably the combination according to the present invention is administered in a serving or unit dosage form that provides a therapeutically effective or prophylactically effective amount of the combination.

[0032] The terms "prevent" and “prevention” mean to administer a composition as disclosed herein to a subject is not showing any symptoms of the condition to reduce or prevent development of at least one symptom associated with the condition. Furthermore, “prevention” includes reduction of risk, incidence and / or severity of a condition or disorder.

[0033] As used herein, an “effective amount” is an amount that treats or prevents a deficiency, treats or prevents a disease or medical condition in an individual, or, more generally, reduces symptoms, manages progression of the disease, or provides a nutritional, physiological, or medical benefit to the individual.

[0034] ‘Animal” includes, but is not limited to, mammals, which includes but is not limited to rodents; aquatic mammals; domestic animals such as dogs, cats and other pets; farm animals such as sheep, pigs, cows and horses; and humans. Where “animal,” “mammal” or a plural thereof is used, these terms also apply to any animal that is capable of the effect exhibited or intended to be exhibited by the context of the passage, e.g., an animal benefitting from improved mitochondrial calcium import. While the term “individual” or “subject” is often used herein to refer to a human,the present disclosure is not so limited. Accordingly, the term “individual” or “subject” refers to any animal, mammal or human that can benefit from the methods and compositions disclosed herein.

[0035] The term “pet” means any animal which could benefit from or enjoy the compositions provided by the present disclosure. For example, the pet can be an avian, bovine, canine, equine, feline, hircine, lupine, murine, ovine, or porcine animal, but the pet can be any suitable animal. The term “companion animal” means a dog or a cat.

[0036] The term “elderly” in the context of a human means an age from birth of at least 60 years, preferably above 63 years, more preferably above 65 years, and most preferably above 70 years. In the context of non-human animals, “elderly” means a non-human subject that has reached 60% of its likely lifespan, in some embodiments at least 70%, at least 80% or at least 90% of its likely lifespan. A determination of lifespan may be based on actuarial tables, calculations, or estimates, and may consider past, present, and future influences or factors that are known to positively or negatively affect lifespan. Consideration of species, gender, size, genetic factors, environmental factors and stressors, present and past health status, past and present nutritional status, and stressors may be taken into consideration when determining lifespan.

[0037] The term “older adult” in the context of a human means an age from birth of at least 45 years, preferably above 50 years, more preferably above 55 years, and includes elderly individuals.

[0038] “Mobility” is the ability to move independently and safely from one place to another.

[0039] As used herein, “frailty” is defined as a clinically recognizable state of increased vulnerability resulting from aging-associated decline in reserve and function across multiple physiologic systems such that the ability to cope with everyday or acute stressors is compromised. A pre-frail stage, in which one or two of these criteria are present, identifies a high risk of progressing to frailty.

[0040] ‘Muscle fatigue” means a reduced contractile force in one or more muscles due to a shortage of substrates within the muscle fiber and / or an accumulation of metabolites within the muscle fiber which interfere either with the release of calcium or with the ability of calcium to stimulate muscle contraction.

[0041] ‘Muscle weakness” is a condition where the force exerted by the muscles is less than would be expected. The U.S. Medical Research Council’s grading system for muscle strength iswidely used to identify muscle weakness and the severity thereof. Specifically, the examiner assesses the patient’s ability to move the muscle against resistance provided by the examiner who, through experience, has developed a sense of the expected range of normal. This will vary from patient-to-patient depending upon the underlying size and conditioning of the subject; the fully trained athlete can be expected to perform differently from a small, sedentary, or deconditioned individual. The expected strength should also be adjusted for degree of atrophy in patients with wasting illnesses.

[0042] The patient’s effort is graded on a scale of 0 to 5. As used herein, “muscle weakness” refers to any of grades 0-4.Grade 5: Muscle contracts normally against full resistance.Grade 4: Muscle strength is reduced, but muscle contraction can still move joint against resistance. Grade 3: Muscle strength is further reduced, such that the joint can be moved only against gravity with the examiner’s resistance completely removed. As an example, the elbow can be moved from full extension to full flexion starting with the arm hanging down at the side.Grade 2: Muscle can move only if the resistance of gravity is removed. As an example, the elbow can be fully flexed only if the arm is maintained in a horizontal plane.Grade 1 : Only a trace or flicker of movement is seen or felt in the muscle, or fasciculations are observed in the muscle.Grade 0: No movement is observed.

[0043] As used herein, a “sportsman” is an individual who participates in at least one of 1) resistance exercise, 2) anaerobic or repeated sprint-type exercise, or 3) endurance exercise.

[0044] “Overweight” is defined for a human as a body mass index (BMI) between 25 and 30 kg / m2. “Obese” is defined for a human as a BMI of at least 30 kg / m2, for example 30-39.9 kg / m2“Weight loss” is a reduction of the total body weight. Weight loss may, for example, refer to the loss of total body mass in an effort to improve one or more of health, fitness or appearance.

[0045] Diabetes" encompasses both the type I and type II forms of the disease. Non-limiting examples of risk factors for diabetes include: waistline of more than 40 inches for men or 35 inches for women, blood pressure of 130 / 85 mmHg or higher, triglycerides above 150 mg / dl, fasting blood glucose greater than 100 mg / dl or high-density lipoprotein of less than 40 mg / dl in men or 50 mg / dl in women.

[0046] As used herein, the term "metabolic syndrome" refers to a combination of medicaldisorders that, when occurring together, increase the risk of developing cardiovascular disease and diabetes. It affects one in five people in the United States and prevalence increases with age. Some studies have shown the prevalence in the United States to be an estimated 25% of the population. In accordance with the International Diabetes Foundation consensus worldwide definition (2006), metabolic syndrome is central obesity plus any two of the following:

[0047] Raised triglycerides: > 150 mg / dL (1.7 mmol / L), or specific treatment for this lipid abnormality;

[0048] Reduced HDL cholesterol: < 40 mg / dL (1.03 mmol / L) in males, < 50 mg / dL (1.29 mmol / L) in females, or specific treatment for this lipid abnormality;

[0049] Raised blood pressure: systolic BP > 130 or diastolic BP >85 mm Hg, or treatment of previously diagnosed hypertension; and

[0050] Raised fasting plasma glucose: (FPG) > 100 mg / dL (5.6 mmol / L), or previously diagnosed type 2 diabetes.

[0051] As used herein, a “cognitive disorder” refers to any condition that impairs cognitive function. Non-limiting examples of a cognitive disorder include delirium, dementia, learning disorder, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD).

[0052] The terms “serving” or "unit dosage form," as used herein, are interchangeable and refer to physically discrete units suitable as unitary dosages for human and animal subjects, each unit containing a predetermined quantity of the composition comprising a combination according to the present invention, as disclosed herein, in an amount sufficient to produce the desired effect, preferably in association with a pharmaceutically acceptable diluent, carrier or vehicle. The specifications for the unit dosage form depend on the particular compounds employed, the effect to be achieved, and the pharmacodynamics associated with each compound in the host. In an embodiment, the unit dosage form can be a predetermined amount of liquid housed within a container such as a bottle.

[0053] An “oral nutrition supplement” or “ONS” is a composition comprising at least one macronutrient and / or at least one micronutrient, for example in a form of sterile liquids, semi-solids or powders, and intended to supplement other nutritional intake such as that from food. Non-limiting examples of commercially available ONS products include MERITENE®, BOOST®, NUTREN® and SUSTAGEN®. In some embodiments, an ONS can be a beverage inliquid form that can be consumed without further addition of liquid, for example an amount of the liquid that is one serving of the composition.

[0054] As used herein, “incomplete nutrition” refers to preferably nutritional products that do not contain sufficient levels of macronutrients (protein, fats and carbohydrates) or micronutrients to be sufficient to be a sole source of nutrition for the animal to which the nutritional product is being administered. The term "complete nutrition" refers to a product which is capable of being the sole source of nutrition for the subject. An individual can receive 100% of their nutritional requirements from a complete nutrition composition.

[0055] A “kit” means that the components of the kit are physically associated in or with one or more containers and considered a unit for manufacture, distribution, sale, or use. Containers include, but are not limited to, bags, boxes, cartons, bottles, packages of any type or design or material, over- wrap, shrink-wrap, affixed components (e.g., stapled, adhered, or the like), or combinations thereof.

[0056] Embodiments

[0057] An aspect of the present disclosure is a composition comprising an effective amount of a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer for use in achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

[0058] In an embodiment the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP-activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxy butyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

[0059] In a preferred embodiment, the autophagy inducer is spermidine.

[0060] In another embodiment, the autophagy inducer is a combination of spermidine and thymol.

[0061] In an embodiment, the mitochondria-related disease or condition is selected from the group consisting of stress, obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, complications from diabetes, cardiovascular disease, hyperlipidemia, respiratory diseases, pain syndromes, neurodegenerative disease, cognitive disorder, stress-induced or stress-related cognitive dysfunction, mood disorder, anxiety disorder, age-related neuronal death or dysfunction, musculo-skeletal disorder, muscle aging, sarcopenia, frailty, pre-frailty, chronic kidney disease, macular degeneration, and combinations thereof.

[0062] In an embodiment, the at least one physical state is selected from the group consisting of deleterious effects of aging, muscle loss, pre-diabetes, gestational diabetes, type I diabetes, type II diabetes, complications from diabetes, insulin resistance, metabolic syndrome, dyslipidemia, overweight, obesity, raised cholesterol levels, raised triglyceride levels, elevated fatty acid levels, fatty liver disease, renal disease, cardiovascular disease, musculo-skeletal diseases, respiratory diseases, pain syndromes, neurodegenerative disease, impaired cognitive function, myopathy such as statin-induced myopathy, non-alcoholic steatohepatitis, tinnitus, dizziness, alcohol hangover, hearing impairment, osteoporosis, hypertension, atherosclerosis / coronary artery disease, myocardial damage after stress, traumatic brain injury, cystic fibrosis, inflammation, cancer, and HIV infection.

[0063] In an embodiment, the physiological state or disorder related to cell ageing or metabolic fatigue comprises muscle fatigue or weakness, lack of energy, physical energy, lack of vitality or weakness.

[0064] In an embodiment, the composition comprising an effective amount of nicotinamide, glycine and optionally an autophagy inducer, can be administered to an animal such as a human, e.g., an ageing individual or a pet animal, in a therapeutically effective dose. The therapeutically effective dose can be determined by the person skilled in the art and will depend on a number of factors known to those of skill in the art, such as the severity of the condition and the weight and general state of the individual.

[0065] Glycine

[0066] The glycine is preferably L-glycine and / or L-glycine ethyl ester. In another embodiment it can be suitable functional derivatives of glycine.

[0067] Non-limiting examples of suitable glycine functional derivatives include D- Allylglycine; N-[Bis(methylthio)methylene]glycine methyl ester; Boc-allyl-Gly-OH (dicyclohexylammonium) salt; Boc-D-Chg-OH; Boc-Chg-OH; (R)-N-Boc-(2'-chlorophenyl)glycine; Boc-L-cyclopropylglycine; Boc-L-cyclopropylglycine; (R)-N-Boc-4-fluorophenylglycine; Boc-D-propargylglycine; Boc-(S)-3 -thienylglycine; Boc-(R)-3 -thienylglycine;D-a-Cyclohexylglycine; L-a-Cyclopropylglycine;N-(2-fluorophenyl)-N-(methylsulfonyl)glycine; N-(4-fluorophenyl)-N-(methylsulfonyl)glycine; Fmoc-N-(2,4-dimethoxybenzyl)-Gly-OH; N-(2-Furoyl)glycine; L-a-Neopentylglycine; D-Propargylglycine; sarcosine; Z-a-Phosphonoglycine trimethyl ester, and a mixture thereof.

[0068] In an embodiment, the glycine may be provided in a dipeptide, such as N-acetylcysteinylglycine(GlyNAC) or cysteinylglycine.

[0069] The glycine (GLY) or functional derivative thereof can be administered in an amount of about 0.1 - 100 milligram (mg) of glycine or functional derivative thereof per kilogram (kg) of body weight of the subject. In some embodiments, these amounts are provided at least partially by a dipeptide comprising both the N-acetylcysteine and the glycine or functional derivative thereof.

[0070] In another embodiment, collagen and collagen peptides may be used as source of glycine. Additionally, whey protein is rich in glycine and cysteine, so that in another embodiment, the glycine and N-acetylcysteine are provided by whey protein. Some plant-based protein source may also provide source of glycine.

[0071] In a particular non-limiting example, the daily doses of glycine for a 60 kg subject can be from 6 to 6,000 mg / day.

[0072] Nicotinamide

[0073] In an embodiment, the Nicotinamide, also known as niacinamide or nicotinic acid amide, is the water-soluble, active form of vitamin B3.

[0074] The method can comprise administering, in particular to pet animal, a daily amount of nicotinamide (NAM) in the weight range of 0.05 mg - 1 g per kg body weight, preferably 1 mg -500 mg per kg body weight, more preferably 5 mg - 250 mg per kg body weight, even more preferably 4 mg - 125 mg per kg body weight, or most preferably 2 mg - 80 mg per kg body weight.

[0075] In some embodiments, the method can comprise administering to small breed dogs, an amount of about 100 mg / day to 3600 mg / day, preferably about 50 mg / day to 1800 mg / day, morepreferably about 20 mg / day to 840 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0076] In some embodiments, the method can comprise administering to medium breed dogs, an amount of about 800 mg / day to 8000 mg / day, preferably about 400 mg / day to 5000 mg / day, more preferably about 100 mg / day to 2400 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0077] In some embodiments, the method can comprise administering to large breed dogs, an amount of about 1800 mg / day to 24000 mg / day, preferably about 1200 mg / day to 16000 mg / day, more preferably about 600 mg / day to 8000 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0078] In some embodiments, the method can comprise administering to small breed cats, an amount of about 150 mg / day to 1200 mg / day, preferably about 100 mg / day to 750 mg / day, more preferably about 50 mg / day to 360 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0079] In some embodiments, the method can comprise administering to medium breed cats, an amount of about 250 mg / day to 1500 mg / day, preferably about 150 mg / day to 900 mg / day, more preferably about 75 mg / day to 450 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0080] In some embodiments, the method can comprise administering to large breed cats, an amount of about 300 mg / day to 3000 mg / day, preferably about 200 mg / day to 2000 mg / day, more preferably about 90 mg / day to 1000 mg / day of nicotinamide (NAM) per serving of a food item or supplement or treat.

[0081] The nicotinamide can be administered in particular to a human, in an amount of about 0.001 mg / day to about 3000 mg / day, for example 1 mg / day to about 3000 mg / day, preferably about 10 mg / day to about 2000 mg / day, more preferably from 500 mg / day to about 1000 mg / day. Of course, the daily dose can be administered in portions at various hours of the day. However, in any given case, the amount of compound administered will depend on such factors as the solubility of the active component, the formulation used, subject condition (such as weight), and / or the route of administration. For example, the daily doses of nicotinamide disclosed above are non-limiting and, in some embodiments, may be different; in particular, the compositions disclosed herein canbe utilized as an acute care food for special medical purposes (FSMP) and contain up to about 3.0 g nicotinamide / day.

[0082] Autophagy inducer

[0083] In an embodiment the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP-activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxybutyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

[0084] In a preferred embodiment, the autophagy inducer is spermidine.

[0085] In another embodiment, the autophagy inducer is a combination of spermidine and thymol.

[0086] In an embodiment, spermidine is added to the combination of Nicotinamide and Glycine. Non-limiting examples of suitable forms of spermidine include spermidine trihydrochloride, spermidine phosphate hexahydrate, spermidine phosphate hexahydrate, and L-arginyl-3,4-spermidine. Wheat germ is rich in spermidine. Therefore, some embodiments of the composition comprise wheat germ and / or enriched wheat germ extracts that provide at least a portion of the spermidine in the composition.

[0087] In particular embodiments, the method can comprise administering daily the autophagy inducer, such as spermidine in the weight range of 0.05 mg - 1 g per kg body weight, preferably 1 mg -200 mg per kg body weight, more preferably 5 mg - 150 mg per kg body weight, even more preferably 10 mg - 120 mg per kg body weight, or most preferably 40 mg - 80 mg per kg body weight. In some embodiments, the method can comprise administering 10 mg - 100 mg of spermidine per serving, for example about 16 mg per serving of a food item or about 80 mg per serving of a supplement or treat.

[0088] In a further embodiment, the method can comprise administering daily thymol and / or carvacrol in the weight range of 0.05 mg - 1 g per kg body weight, preferably 1 mg -200 mg per kg body weight, more preferably 5 mg - 150 mg per kg body weight, even more preferably 10 mg - 120 mg per kg body weight, or most preferably 40 mg - 80 mg per kg body weight.

[0089] Typically between 50 pg to 10 g of thymol and / or carvacrol, per daily serving in one or more portions is administered to an individual.

[0090] Thymol (10-64%) is one of the major constituent of essential oils of thyme (Thymus vulgaris L., Lamiaceae). Carvacrol is present in the essential oil of Origanum vulgare (oregano), oil of thyme, oil obtained from pepperwort, and wild bergamot. The essential oil of thyme subspecies containsbetween 5% and 75% of carvacrol, while Satureja (savory) subspecies have a content between 1% and 45%. Origanum majorana (marjoram) and Dittany of Crete are rich in carvacrol, 50% and 60-80% respectively. Therefore, some embodiments of the composition comprise such plant and / or enriched plant extracts, essential oils or fractions that provide at least a portion of thymol and / carvacrol in the composition, in particular from thyme and oregano.

[0091] Subject

[0092] In an embodiment, the individual is selected from the group consisting of an adult, an aging subject; an elderly subject; a subject with muscle fatigue or muscle weakness; a subject with impaired mobility; a frail subject; a pre-frail subject; a sarcopenic subject; a subject recovering from pre-frailty, frailty, sarcopenia or impaired mobility; a subject undergoing physical rehabilitation; a sportsman; and a pet.

[0093] In some embodiments, the individual is healthy. In some embodiments, the individual has sarcopenia, frailty, muscle fatigue or muscle weakness, or impairment in one or more of muscle functionality, muscle performance, lean muscle mass or muscle strength, but optionally is otherwise healthy.

[0094] For example, the combination can be administered to a sportsman before, during and / or after exercise, for example less than two hours before the exercise or less than one hour before the exercise and less than two hours after the exercise or less than one hour after the exercise.

[0095] In an embodiment, the combination is administered in a composition selected from the group consisting of a food product, a petfood, a food for special medical purposes (FSMP), a nutritional supplement, a dairy-based drink, a low-volume liquid supplement, a meal replacement beverage, and combinations thereof.

[0096] Each of the compounds can be administered at the same time as the other compounds (i.e., as a single unit) or separated by a time interval (i.e., in separate units).

[0097] The compositions for use according to the invention may also comprise at least one further bioactive compound selected from the group consisting of antioxidants, anti-inflammatory compounds, glycosaminoglycans, prebiotics, fibers, probiotics, fatty acids, enzymes, minerals, trace elements and / or vitamins.

[0098] Nutritional compositions

[0099] The compositions for use according to the invention may be nutritional compositions or pharmaceutical compositions, and may be for human or veterinary use. In an embodiment, the combination is administered orally.

[0100] Thus, in preferred embodiments, the composition for use according to the invention is a nutritional composition.

[0101] By "nutritional composition" is meant in the context of the present application a composition which is a source of nutrition to an individual.

[0102] The nutritional products or compositions of the invention may be a source of complete nutrition or may be a source of incomplete nutrition. As used herein, “complete nutrition” includes nutritional products and compositions that contain sufficient types and levels of macronutrients (protein, fats and carbohydrates) and micronutrients to be sufficient to be a sole source of nutrition for the animal to which it is being administered to. Patients can receive 100% of their nutritional requirements from such complete nutritional compositions. As used herein, “incomplete nutrition” includes nutritional products or compositions that do not contain sufficient levels of macronutrients (protein, fats and carbohydrates) or micronutrients to be sufficient to be a sole source of nutrition for the animal to which it is being administered to. Partial or incomplete nutritional compositions can be used as a nutritional supplement.

[0103] Non-limiting examples of suitable compositions for the include food compositions, dietary supplements, dietary supplements (e.g., liquid ONS), complete nutritional compositions, beverages, pharmaceuticals, oral nutritional supplement, medical food, nutraceuticals, food for special medical purpose (FSMP), powdered nutritional products to be reconstituted in water or milk before consumption, food additives, medicaments, drinks, petfood, and combinations thereof.

[0104] In an embodiment, the compositions for use according to the invention include a source of protein. The protein source may be dietary protein including, but not limited to animal protein (such as milk protein, meat protein or egg protein), vegetable protein (such as soy protein, wheat protein, rice protein, and pea protein), or combinations thereof. In an embodiment, the protein is selected from the group consisting of whey, chicken, corn, caseinate, wheat, flax, soy, carob, pea or combinations thereof.

[0105] In an embodiment, the compositions include a source of carbohydrates. Any suitable carbohydrate may be used in the present compositions including, but not limited to, starch,sucrose, lactose, glucose, fructose, corn syrup solids, maltodextrin, modified starch, amylose starch, tapioca starch, corn starch, xylitol, sorbitol or combinations thereof.

[0106] In an embodiment, the compositions include a source of fat. The source of fat may include any suitable fat or fat mixture. For example, the fat source may include, but is not limited to, vegetable fat (such as olive oil, corn oil, sunflower oil, high-oleic sunflower, rapeseed oil, canola oil, hazelnut oil, soy oil, palm oil, coconut oil, blackcurrant seed oil, borage oil, lecithins, and the like), animal fats (such as milk fat), or combinations thereof. The source of fat may also be less refined versions of the fats listed above (e.g., olive oil for polyphenol content).

[0107] In addition, compositions for use according to the invention may also comprise natural or artificial flavours, for example fruit flavours like banana, orange, peach, pineapple or raspberry or other plant flavours like vanilla, cocoa, coffee, etc.

[0108] Nutritional composition formats

[0109] The nutritional compositions may include, besides the main bioactive components and any further bioactive components, and optionally one or more of a protein, carbohydrate and fat source, any number of optional additional food ingredients, including conventional food additives (synthetic or natural), for example one or more acidulants, additional thickeners, buffers or agents for pH adjustment, chelating agents, colorants, emulsifiers, excipient, flavor agent, mineral, osmotic agents, a pharmaceutically acceptable carrier, preservatives, stabilizers, sugar, sweeteners, texturizers, and / or vitamins. The optional ingredients can be added in any suitable amount.

[0110] The nutritional composition may be provided in any suitable format. Examples of nutritional composition formats in which the composition for use according to the invention may be provided include solutions, ready -for-consumption compositions (e.g. ready -to-drink compositions or instant drinks), liquid comestibles, soft drinks, juice, sports drinks, milk drinks, milk-shakes, yogurt drinks, soup, etc.

[0111] In another embodiment, the nutritional compositions may be provided in the form of a concentrate, a powder, or granules (e.g. effervescent granules), which are diluted with water or other liquid, such as milk or fruit juice, to yield the ready -for-consumption composition.

[0112] Further nutritional composition formats include, baked products, dairy products, desserts, confectionery products, cereal bars, and breakfast cereals. Examples of dairy productsinclude milk and milk drinks, yoghurts and other cultured milk products, ice creams and cheeses. Examples of baked products include bread, biscuits and cakes.

[0113] In one embodiment, the composition for use according to the invention may also be available in a great variety of formats designed as animal foods, in particular for the dog or the cat, whether in a wet form, semi-wet form or dry form, in particular in the form of biscuits.

[0114] The compositions disclosed herein can use any of a variety of formulations for therapeutic administration. More particularly, pharmaceutical compositions can comprise appropriate pharmaceutically acceptable carriers or diluents and may be formulated into preparations in solid, semi-solid, liquid or gaseous forms, such as tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres, and aerosols. As such, administration of the composition can be achieved in various 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 regional administration, intramural administration, or use of an implant that acts to retain the active dose at the site of implantation.

[0115] In pharmaceutical dosage forms, the compounds may be administered as their pharmaceutically acceptable salts. They may also be used in appropriate association with other pharmaceutically active compounds. The following methods and excipients are merely exemplary and are in no way limiting.

[0116] For oral preparations, the compounds can be used alone or in combination with appropriate additives to make tablets, powders, granules or capsules, for example, with conventional additives, such as lactose, mannitol, corn starch or potato starch; with binders, such as crystalline cellulose, cellulose functional derivatives, acacia, corn starch or gelatins; with disintegrators, such as corn starch, potato starch or sodium carboxymethylcellulose; with lubricants, such as talc or magnesium stearate; and if desired, with diluents, buffering agents, moistening agents, preservatives and flavoring agents.

[0117] Routes of administration

[0118] The nutritional compositions of the present disclosure may be administered by any means suitable for human administration, and in particular for administration in any part of the gastrointestinal tract. Enteral administration, oral administration, and administration through atube or catheter are all covered by the present disclosure. The nutritional compositions may also be administered by means selected from oral, rectal, sublingual, sublabial, buccal, topical, etc.

[0119] In an embodiment, the composition according to this invention is administered orally.

[0120] In an embodiment, the combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, is administered in a composition selected from the group consisting of food compositions, dietary supplements, nutritional compositions, beverages, nutraceuticals, powdered nutritional products to be reconstituted in water or milk before consumption, food additives, medicaments, drinks, petfood, and combinations thereof.

[0121] The nutritional compositions may be administered in any known form including, for example, tablets, capsules, liquids, chewables, soft gels, sachets, powders, syrups, liquid suspensions, emulsions and solutions in convenient dosage forms. In soft capsules, the active ingredients are preferably dissolved or suspended in suitable liquids, such as fatty oils, paraffin oil or liquid polyethylene glycols. Optionally, stabilizers may be added.

[0122] If the nutritional compositions are administered by tube feeding, the nutritional compositions may be used for short term or long term tube feeding.

[0123] The composition can be administered to an individual such as a human, e.g., an ageing individual or a critically ill individual, in a therapeutically effective dose. The therapeutically effective dose can be determined by the person skilled in the art and will depend on a number of factors known to those of skill in the art, such as the severity of the condition and the weight and general state of the individual.

[0124] The composition is preferably administered to the individual at least one day per week, preferably at least two days per week, more preferably at least three days per week, most preferably all seven days of the week; for at least one week, at least one month, at least two months, at least three months, at least six months, or even longer. In some embodiments, the composition is administered to the individual consecutively for a number of days, for example at least until a therapeutic effect is achieved. In an embodiment, the composition can be administered to the individual daily for at least 30, 60 or 90 consecutive days.

[0125] In some embodiments, the administration continues for the remaining life of the individual. In other embodiments, the administration occurs until no detectable symptoms of the medical condition remain. In specific embodiments, the administration occurs until a detectableimprovement of at least one symptom occurs and, in further cases, continues to remain ameliorated.

[0126] Method of treatment

[0127] Mitochondrial diseases are the result of either inherited or spontaneous mutations in mitochondrial DNA or nuclear DNA which lead to altered functions of the proteins or RNA molecules that normally reside in mitochondria. Problems with mitochondrial function, however, may only affect certain tissues as a result of factors occurring during development and growth that are not yet fully understood. Even when tissue-specific isoforms of mitochondrial proteins are considered, it is difficult to explain the variable patterns of affected organ systems in the mitochondrial disease syndromes seen clinically.

[0128] Mitochondrial diseases result from failures of the mitochondria, specialized compartments present in every cell of the body except red blood cells. Mitochondria are responsible for creating more than 90% of the energy needed by the body to sustain life and support growth. When they fail, less and less energy is generated within the cell. Cell injury and even cell death follow. If this process is repeated throughout the body, whole systems begin to fail, and the life of the person in whom this is happening is severely compromised. Mitochondrial diseases primarily affect children, but adult onset is becoming more recognized. Diseases of the mitochondria appear to cause the most damage to cells of the brain, heart, liver, skeletal muscles, kidney, and the endocrine and respiratory systems.

[0129] Many symptoms in mitochondrial disorders are non-specific. The symptoms may also show an episodic course, with periodic exacerbations. The episodic condition of migraine, as well as myalgia, gastrointestinal symptoms, tinnitus, depression, chronic fatigue, and diabetes, have been mentioned among the various manifestations of mitochondrial disorders in review papers on mitochondrial medicine. In patients with mitochondrial disorders, clinical symptomatology typically occurs at times of higher energy demand associated with physiological stressors, such as illness, fasting, over-exercise, and environmental temperature extremes. Furthermore, psychological stressors also frequently trigger symptomatology, presumably due to higher brain energy demands for which the patient is unable to match with sufficient ATP production.

[0130] Depending on which cells are affected, symptoms may include loss of motor control, muscle weakness and pain, gastro-intestinal disorders and swallowing difficulties, poor growth,cardiac disease, liver disease, diabetes, respiratory complications, seizures, visual / hearing problems, lactic acidosis, developmental delays and susceptibility to infection.

[0131] Mitochondrial diseases include, without limitation, Alper's disease; Barth syndrome; beta-oxidation defects; carnitine deficiency; carnitine-acyl- carnitine deficiency; chronic progressive external ophthalmoplegia syndrome; co-enzyme Q10 deficiency; Complex I deficiency; Complex II deficiency; Complex III deficiency; Complex IV deficiency; Complex V deficiency; CPT I deficiency; CPT II deficiency; adenosylcobalamin deficiency syndrome; cytochrome c oxidase deficiency; glutaric aciduria type II; Kearns-Sayre syndrome; lactic acidosis; LCHAD (long-chain acyl-CoA dehydrogenase deficiency); Leber's hereditary optic neuropathy; Leigh disease; lethal infantile cardiomyopathy; Luft 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 syndrome; POLG mutations; pyruvate carboxylase deficiency; pyruvate dehydrogenase deficiency; SCHAD (short-chain acyl-CoA dehydrogenase deficiency); and very long-chain acyl-CoA dehydrogenase deficiency.

[0132] Accordingly, an aspect of the present disclosure is a method of achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (v) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (vi) improving mobility and / or (vii) improving healthspan and / or lifespan in an individual, the method comprising orally administering to the individual in need thereof or at risk thereof an effective amount of the combination of nicotinamide (NAM) and glycine, optionally an autophagy inducer.

[0133] The present invention also relates to a method of treating in an individual in need thereof or preventing in an individual at risk thereof at least one condition selected from the group consisting of (i) impairment in at least one of muscle functionality, muscle performance, lean muscle mass or muscle strength, (ii) muscle fatigue or muscle weakness, (iii) pre-frailty, frailty, 1sarcopenia or impaired mobility, and (iv) a muscle disorder linked to calcium depletion or deficiency, the method comprising orally administering to the individual in need thereof or at risk thereof an effective amount of the combination according to the present invention.

[0134] The present invention also relates to a composition comprising a combination of nicotinamide (NAM) and glycine, optionally an autophagy inducer, such as spermidine in an effective amount for use in delaying off-set of metabolic decline, decreasing oxidative stress, mitigating deleterious effects of aging, improving or maintaining muscle function, improving at least one of muscle performance or muscle recovery from exercise, exercise capacity and / or physical function, improving immune function and / or cognitive function in an individual, and / or reducing severity of metabolic and / or degenerative diseases in an individual.

[0135] Preferably, the muscle functionality that can be improved by the methods disclosed herein comprises a characteristic selected from the group consisting of muscle strength, gait speed, and combinations thereof. Muscle function is typically defined as strength per unit of appendicular skeletal muscle mass or per muscle volume.

[0136] Non-limiting examples of a muscle disorder linked to calcium depletion or deficiency that can be treated by the methods disclosed herein include muscular dystrophies, congenital core myopathies and mitochondrial myopathies. Particular non-limiting examples include Barth syndrome; chronic progressive external ophthalmoplegia (cPEO); Kearns-Sayre syndrome (KSS); Leigh syndrome; mitochondrial DNA depletion syndromes (MDDS); mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS); mitochondrial neurogastro intestinal encephalomyopathy (MNGIE); myoclonus epilepsy with ragged red fibers (MERRF); neuropathy, ataxia, and retinitis pigmentosa (NARP); and Pearson syndrome.

[0137] The individual can be at risk of a disorder or condition (e.g., sarcopenia, frailty, muscle fatigue or muscle weakness, or impairment in one or more of muscle functionality, muscle performance, lean muscle mass or muscle strength), in which case the effective amount of the composition is a prophylactically effective dose; or the individual can have a disorder or condition, in which case the effective amount of the composition is a therapeutically effective dose. In some embodiments, the methods comprise identifying the individual as having the condition or being at risk of the condition before the administration.

[0138] Further regarding muscle performance, the increased muscle performance may be one or more of improved muscle function, reduced decline in muscle function, improved musclestrength, improved muscle endurance and improved muscle recovery. The composition can improve physical endurance (e.g., ability to perform a physical task such as exercise, physical labor, sports activities), inhibit or retard physical fatigue, enhance blood oxygen levels, enhance energy in healthy individuals, enhance working capacity and endurance, reduce muscle fatigue, reduce stress, enhance cardiac and cardiovascular function, improve sexual ability, increase muscle ATP levels, and / or reduce lactic acid in blood. “Endurance capacity” refers to the time to fatigue when exercising at a constant workload, generally at an intensity <80% V02max. In some embodiments, the composition is administered in an amount that increases mitochondrial activity, increases mitochondrial biogenesis, and / or increases mitochondrial mass.

[0139] The compositions and the methods disclosed herein can also be effective in the treatment of muscle-related pathological conditions, including myopathies; neuromuscular diseases, such as Duchenne muscular dystrophy; acute sarcopenia, for example, muscle atrophy; and / or cachexia associated with burns, bed rest, limb immobilization, or major thoracic, abdominal, and / or orthopedic surgery.

[0140] In some embodiments, the composition is administered to an individual having impaired physical performance, impaired endurance capacity, and / or impaired muscle function. Improved muscle function can be particularly beneficial in elderly subjects with reduced muscle function as a result of an age-related condition. For example, a subject who may benefit from improved muscle function may experience a decline in muscle function which then leads to pre-frailty and frailty. Such subjects may not necessarily experience muscle wastage in addition to their decline in muscle function. Some subjects do experience both muscle wasting and a decline in muscle function, for example subjects with sarcopenia. The composition may enhance muscle performance in a subject who is frail or pre- frail.

[0141] In another embodiment, the present disclosure provides a method of treating or preventing impaired mobility in an older adult. The method comprises orally administering to the older adult an effective amount of a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer. The older adult can be an elderly individual. In some embodiments, the older adult has a condition selected from the group consisting of frailty, pre-frailty, sarcopenia, recovering from sarcopenia, osteoporosis, osteoarthritis, malnutrition, at risk of malnutrition, undergoing rehabilitation, scheduled to undergo rehabilitation within the next year, and combinations thereof.

[0142] The composition may be administered to the older adult in an amount sufficient to prevent, at least partially reduce the risk of developing frailty or sarcopenia, and / or at least partially reduce the severity of pre-frailty, frailty, sarcopenia or impaired mobility in instances where the condition has yet not been developed in the individual. Such an amount is defined to be “a prophylactically effective dose.” Again, the precise amounts depend on a number of factors relating to the individual, such as their weight, health and how much muscle functionality (e.g., muscle strength, gait speed, etc.) is being lost.

[0143] The present invention also relates to a method of manufacturing a nutritional composition for use according to any of the preceding claims, comprising the steps of providing one or more ingredients for a nutritional composition, nicotinamide (NAM) and glycine, optionally an autophagy inducer, such as spermidine and mixing.

[0144] The present invention also relates to a unit dosage form comprising a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, the unit dosage form comprises an amount of the combination effective for at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

[0145] In an embodiment, the unit dosage form of consists essentially of the combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer.

[0146] In an embodiment the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP-activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxy butyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

[0147] In a preferred embodiment, the autophagy inducer is spermidine.

[0148] In another embodiment, the autophagy inducer is a combination of spermidine and thymol.

[0149] The compositions disclosed herein can also be used in the treatment of any of a variety of additional diseases and conditions in which defective or diminished mitochondrial activity participates in the pathophysiology of the disease or condition, or in which increased mitochondrial function will yield a desired beneficial effect.

[0150] It should be noted that embodiments and features described in the context of one of the aspects of the present invention also apply to the other aspects of the invention.

[0151] The compositions for use according to the invention are herein described in different parameters, such as the ingredients, nutritional composition formats, uses, target groups etc. It should be noted that embodiments and features described in the context of one of the parameters of the composition for use according to the invention, may also be combined with other embodiments and features described in the context of another parameter, unless expressly stated otherwise.

[0152] All patent and non-patent references cited in the present application, are hereby incorporated by reference in their entirety.

[0153] The invention will now be described in further details in the following non-limiting examples.EXAMPLE

[0154] The following non-limiting examples present experimental data supporting the compositions and methods disclosed herein.

[0155] Example 1

[0156] Material and methods

[0157] Mice were housed in IVC GM500 cages (500 cm2, Tecniplast) at the conventional room of the EPFL-SV animal facility. Animals were kept in a temperature -and humidity. Controlled environment with a 12:12-h light-dark cycle. They had access to nesting and enrichment materials and were provided with ad libitum access to water and food. They were housed 2 mice per cage, with the exception to be in single cage in case of fights. Mice were identified by standard ear clipping. Routine check of animal house staff was performed regularly as per standard method of the EPFL facility. Upon arrival animals had an acclimatization period of 1 week.

[0158] All animal experiments were performed according to national Swiss and EU ethical guidelines and approved by the local animal experimentation committee under license VD3519.

[0159] Male WT C57BL / 6J mice underwent to a long-term dietary supplementation for 14-weeks with different compositions. The young control group (n=17) and old control group (n=24) received a commercial standard powdered diet (Safe 150, SafeDiets), with 20% humidity. The compound-treated groups received the same commercial 20 standard powdered diet, mixed homogeneously with 1.6g / kg of glycine (n=24), 1.6g / kg of NAM (n=24) and 1.6g / kg of their combination (n=24) in 20% humidity. Compounds were mixed to the food in an isogenic and isocaloric diet. At the end of the study, mice were tested through cognitive and mobility phenotyping, and then mice were euthanized with isoflurane after 2-hours fasting.

[0160] Muscles EDL and soleus were freshly dissected to perform respiratory state using high-resolution oxygraphy with the Oroboros system.

[0161] EDL and Soleus muscles were dissected from the mice, carefully cleaned of blood, tendons, and fat, and placed in a 12- well plate containing 2 ml of ice-cold BIOPS preservation buffer. Muscle fiber bundles were manually isolated from any remaining tendons and kept at 4°C. After saponin permeabilization, the muscle bundles were transferred to ice-cold MIR05 buffer (mitochondrial respiration medium). Approximately 3 to 5 mg of tissue was collected and placed in the previously washed and calibrated O2k chamber filled with MIR05 for respiratory measurements.

[0162] Buffer composition:

[0163] - BIOPS 2.77 mM CaK2EGTA, 7.23 mM K2EGTA, 5.77 mM Na2ATP, 6.56 mM MgC12, 20 mM Taurine, 15 mM Na2Phosphocreatine, 20 mM Imidazole and 0.5 mM Dithiothreitol (pH 7.1).

[0164] - MiROS: EGTA 0.5 mM; MgC12.6H2O 3 mM, Taurine 20 mM, KH2PO4 10 mM, HEPES 20 mM, d-sucrose 110 mM and BSA, fatty acid free lg / 1.

[0165] Substrates were sequentially added to the chambers using a Hamilton glass microsyringe, and oxygen consumption rates were calculated after a minimum of 2 minutes of stabilization for each substrate. The analysis was performed using the instrument’s software. Areas were delineated after each injection in the specific region where respiration reached a steady state, as depicted, and metabolic parameters were determined accordingly.

[0166] Results

[0167] To assess the impact of the ingredients on mitochondrial performance, we used high-resolution respirometry to monitor various respiration states in mice (soleus and EDLmuscles) skeletal muscle. In the soleus muscle of mice, we observed an age-related decline in Complex I activity (though not statistically significant), Complex II activity, and an overall reduction in OXPHOS respiration. However, these declines were effectively rescued by the combination of glycine and NAM (Figure 1A). The addition of rotenone (ETS+CII) further suggested that this increase was predominantly mediated by enhanced Complex II activity. Similar improvements were observed in the EDL muscle, where the combination of glycine and NAM increased the activity of both Complex I and Complex II, as well as overall OXPHOS respiration (Figure IB), with the effects presumably driven by Complex I in this case. Importantly, no changes in mitochondrial permeability were detected in either muscle tissue, indicating that the interventions specifically enhanced mitochondrial respiration without compromising membrane integrity. Overall, these data indicate that the combination of NAM and glycine improves mitochondrial fitness and respiration in aged animals, potentially offering an approach to counteract age-associated mitochondrial decline.

[0168] Example 2 - Lifespan and mobility analysis in C. elegans

[0169] Materials and Methods

[0170] The effect of glycine, NAM, spermidine and their combination on lifespan and mobility was tested with C. Elegans using a microfluidic platform (NAGI SydLab device). All strains were obtained from the Caenorhabditis Genetics Center, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Eggs of C. elegans (N2 wild-type strain) were synchronously hatched on solid nematode growth medium (NGM) agar plates at 20°C, that were seeded with Escherichia Coli OP50 strain, grown to L4 stage and harvested in S -medium, and injected into the microfluidic chip. Worms were fed with a bacterial medium on chip (freeze-dried bacteria) over their life course with media change every 2-3 days. The treatments were added at day 0 and at each media change (every 2-3 days). Video recordings and picture of each chamber were taken every 6 hours. Worm death events were detected automatically with an in-house developed machine learning algorithm that temporally processes each chamber’s videos over the experimental run. These death events are then aggregated with Kaplan-Meier survival analysis. To extract the motility features, worms were first detected and tracked on each frame of the video by a trained machine learning algorithm. Then, the midline of the worm was defined, and the wormskeleton was segmented in 100 points. The outer two of the 100 points are considered the worm end points in the head / tail region, while the middle two points are considered for the worm’s middle region. For the velocity analysis, the centroid of the segmentation mask of each worm was determined in each video frame assuming a constant density across the entire worm length. The evolution of the x and y centroid coordinates over time was independently approximated by fitting a fifth-degree polynomial. The swimming velocity was then calculated by differentiating the centroid position with respect to time.

[0171] Statistical Analysis

[0172] Data were plotted and analyzed using GraphPad Prism 10. Normality was tested to assess distribution using and non-parametric tests were conducted when data did not follow a normal distribution. Details of the statistical tests are included in the figure legends with associated p-values depicted. P<0.05 were considered significant.

[0173] Results

[0174] We measured lifespan in Wild-type N2 C. elegans worms using a high-content automated microfluidics analyser. We observed no changes in lifespan for worms treated with each single ingredients, glycine (100 pM), spermidine (50 pM) or NAM (10 pM) (Figure 2). When combined at these concentrations, glycine, NAM and spermidine showed a significant increase in lifespan, indicating a synergy for the three ingredients.

[0175] We then tested the effect of the single ingredients and their combination on mobility in C. elegans using the same analyser. This system allows to track velocity for the worms in mm2 / second. First, we observed a clear decrease in all motility parameters over the life-course of the worms towards almost complete immobility (data not shown) confirming findings in the literature. For this reason, we focused our analysis on the first 10 days of life, as no rescue was observed after day 11 with any of the treatments (data not shown). In these conditions, the combination of ingredients (with NAM at lOpM) was significantly superior to all single ingredients when considering velocity (Figure 3). In conclusion, in the worm, we observed a beneficial effect of the combination of ingredients not only on lifespan but also an impact on a mobility health benefit.

[0176] 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 present subjectmater and without diminishing its intended advantages. It is therefore intended that such changes and modifications be covered by the appended claims.

Claims

The invention is claimed as follows:

1. A composition comprising an effective amount of a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, for use in achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

2. The composition according to claim 1, wherein the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP -activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxybutyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

3. The composition according to claim 1, wherein the autophagy inducer is spermidine.

4. The composition according to claim 1, wherein the autophagy inducer is a combination of spermidine and thymol.

5. The composition for use according to any preceding claim, wherein the mitochondria-related disease or condition is selected from the group consisting of stress, obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, complications from diabetes, cardiovascular disease, hyperlipidemia, respiratory diseases, pain syndromes, neurodegenerative disease, cognitive disorder, stress-induced or stress-related cognitive dysfunction, mood disorder, anxiety disorder, age-related neuronal death or dysfunction, musculo-skeletal disorder, muscle aging, sarcopenia, frailty, pre-frailty, chronic kidney disease, macular degeneration, and combinations thereof.

6. The composition for use according to any one of the preceding claims, wherein the at least one physical state is selected from the group consisting of deleterious effects of aging, muscle loss, pre-diabetes, gestational diabetes, type I diabetes, type II diabetes,complications from diabetes, insulin resistance, metabolic syndrome, dyslipidemia, overweight, obesity, raised cholesterol levels, raised triglyceride levels, elevated fatty acid levels, fatty liver disease, renal disease, cardiovascular disease, musculo-skeletal diseases, respiratory diseases, pain syndromes, neurodegenerative disease, impaired cognitive function, myopathy such as statin-induced myopathy, non-alcoholic steatohepatitis, tinnitus, dizziness, alcohol hangover, hearing impairment, osteoporosis, hypertension, atherosclerosis / coronary artery disease, myocardial damage after stress, traumatic brain injury, cystic fibrosis, inflammation, cancer, and HIV infection.

7. The composition for use according to any one of the preceding claims, wherein the physiological state or disorder related to cell ageing or metabolic fatigue comprises muscle fatigue or weakness, lack of energy, physical energy, lack of vitality or weakness.

8. The composition for use according to any one of the preceding claims, wherein the individual is selected from the group consisting of an adult, an aging subject; an elderly subject; a subject with muscle fatigue or muscle weakness; a subject with impaired mobility; a frail subject; a pre-frail subject; a sarcopenic subject; a subject recovering from pre-frailty, frailty, sarcopenia or impaired mobility; a subject undergoing physical rehabilitation; a sportsman; and a pet.

9. The composition for use according to any one of the preceding claims, wherein it is administered orally.

10. The composition for use according to any one of the preceding claims, wherein the combination is administered in a composition selected from the group consisting of a food product, a petfood, a food for special medical purposes (FSMP), a nutritional supplement, a dairy-based drink, a low-volume liquid supplement, a meal replacement beverage, and combinations thereof.

11. A composition comprising a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, in an effective amount for use in delaying off-set of metabolic decline, decreasing oxidative stress, mitigating deleterious effects of aging, improving or maintaining muscle mass, improving at least one of muscle performance or muscle recovery from exercise, exercise capacity and / or physical function, improving immune function and / or cognitive function and / or reducing severity of metabolic and / or degenerative diseases in an individual.

12. A method of achieving at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual, the method comprising orally administering to the individual in need thereof or at risk thereof an effective amount of the combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer.

13. A method according to claim 12, wherein the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP-activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxybutyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

14. The method according to claim 12, wherein the autophagy inducer is spermidine.

15. The method according to claim 12, wherein the autophagy inducer is a combination of spermidine and thymol.

16. The method according to claim 10 to 15, wherein the combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, is administered in a composition selected from the group consisting of food compositions, dietary supplements, nutritional compositions, beverages, nutraceuticals, powdered nutritional products to be reconstituted in water or milk before consumption, food additives, medicaments, drinks, petfood, and combinations thereof.

17. A method of treating in an individual in need thereof or preventing in an individual at risk thereof at least one condition selected from the group consisting of (i) impairment in at least one of muscle functionality, muscle performance, lean muscle mass or muscle strength, (ii) muscle fatigue or muscle weakness, (iii) pre-frailty, frailty, sarcopenia or impaired mobility, and (iv) a muscle disorder linked to calcium depletion or deficiency, the method comprising orally administering to the individual in need thereof or at risk thereof aneffective amount of the combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer.

18. A unit dosage form comprising a combination of nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, the unit dosage form comprises an amount of the combination effective for at least one result selected from the group consisting of i) preventing and / or treating a mitochondria-related disease or condition associated with altered mitochondrial function and / or ii) at least one physical state selected from the group consisting of oxidative stress or a condition associated with oxidative stress in an individual; (iii) increasing resistance to age-related pathologies; (iv) improving a physiological state or disorder related to cell ageing or metabolic fatigue in one or more cells; (v) improving mobility and / or (vi) improving healthspan and / or lifespan in an individual.

19. The unit dosage form of Claim 18, consisting essentially of the combination of nicotinamide (NAM) and glycine, optionally further comprising the autophagy inducer.

20. A unit dosage according to claim 18 or 19, wherein the autophagy inducer is at least one of spermidine, thymol, carvacrol, urolithin (e.g., Urolithin A, B or D), rapamycin, Torinl, valproic acid, polyphenols (e.g., resveratrol), caffeine, metformin, 5' AMP-activated protein kinase (AMPK) activators, L-type calcium channel inhibitors, ketones (e.g., beta-hydroxybutyrate, ketone salts, or ketone ester derivatives), and mixtures thereof.

21. The unit dosage according to claim 18 or 19, wherein the autophagy inducer is spermidine.

22. The unit dosage according to claim 18 or 19, wherein the autophagy inducer is a combination of spermidine and thymol.

23. A method of manufacturing a nutritional composition for use according to any of the preceding claims, comprising the steps of providing one or more ingredients for a nutritional composition, nicotinamide (NAM) and glycine, optionally further comprising an autophagy inducer, and mixing.

Images