Prevention or treatment of age-associated decline in motor function and internal organ diseases, and composition for use in suppression of age-associated decline in α-klotho level in blood

Abstract

The present invention relates to: prevention of age-associated decline in motor function and / or prevention or treatment of diseases induced by age-associated decline in motor function; prevention of age-associated decline in internal organ functions such as liver, heart, kidney, and pancreas functions and / or prevention or treatment of diseases induced by age-associated decline in internal organ functions; and a composition containing a glycyrrhizic acid analog that suppresses age-associated decline in the blood concentration of α-Klotho, a longevity protein that attenuates decline in various organ functions.

Description

A composition for preventing decline in motor function associated with aging, preventing or treating visceral diseases, and suppressing the decrease in blood concentration of α - klotho associated with aging

[0001] The present invention relates to a composition containing a glycyrrhizin analog that suppresses the decline in motor function associated with aging and / or prevents or treats diseases induced by the decline in motor function associated with aging, suppresses the decline in visceral functions such as the liver, heart, kidneys, and spleen associated with aging, and / or prevents or treats diseases induced by the decline in visceral functions associated with aging, and suppresses the decrease in blood concentration of the longevity protein α - klotho associated with aging of various organ functions.

[0002] In recent years, aging has been progressing in developed countries including Japan. As a result, the number of elderly people who require medical treatment for diseases such as lifestyle-related diseases and cancer, or support and care for the decline in physical function and cognitive function, has been increasing, which has become a heavy burden on society. Therefore, for the elderly, maintaining a healthy state without seeking medical treatment or support and care is required.

[0003] Originally, aging is an inevitable phenomenon in all living organisms. However, various countermeasures have been considered to delay its progression. Preventing the decline in motor function by utilizing physical functions through exercise, or improving nutrition to avoid lifestyle-related diseases such as diabetes and hypertension are part of this, but the effects of these are limited.

[0004] In the future, with the advent of a super-aged society, there is concern that the significant increase in the number of elderly people and the resulting strain on medical care and nursing will become a social problem. Therefore, more effective means are required to extend the healthy life expectancy of the elderly.

[0005] Incidentally, the progression of aging is inherently subject to individual differences based on genetic traits. Among these traits, one important factor involved in aging control has been identified as an aging-suppressing gene (Non-Patent Literature 1). This gene produces a protein called α-klotho, which has an aging-suppressing function. This protein is mainly produced in the kidneys, migrates to various organs via the bloodstream, and has been shown to suppress the aging of those organs as well (Non-Patent Literature 2). However, the amount of α-klotho produced by the aging-suppressing gene decreases with age, so the decline in organ function due to aging progresses more rapidly (Non-Patent Literature 3). Furthermore, there are individual differences in the level of α-klotho production. In elderly people, frailty can develop, which causes a decline in various organs, motor function, and cognitive function. It is also known that there is an inverse correlation between the incidence of frailty and α-klotho blood concentration (Non-Patent Literature 4). Therefore, increasing α-klotho blood concentration can also contribute to maintaining physical function in the elderly. In fact, there are studies showing that increasing α-klotho blood levels leads to improvements in the nervous system, circulatory system, and renal function (Non-Patent Documents 5, 6, 7). Focusing on this function of α-klotho, the search for compounds that promote the biosynthesis of this protein has been underway. As a result, novel compounds suitable for the purpose have been synthesized, and these compounds have resulted in increased α-klotho blood levels and improvements in renal function, etc. (Patent Document 1). Furthermore, searches from natural resources have also been conducted, and an active ingredient was found in the herbal medicine Gastrodia elata, and the aging of the circulatory system was mitigated along with an increase in α-klotho blood levels (Non-Patent Document 8). In response to these cases, searches from various natural resources are continuing (Non-Patent Document 2).

[0006] Japanese Patent Publication No. 2023-503217, Japanese Patent Publication No. 2023-122155

[0007] M.Kuro-o et al.“Mutation of the mouse klotho gene leads to a syndrome resembling ageing” Nature, (1997),390,45-51G. J. Prud’homme et al. “Pathobiology of the klotho antiaging protein and therapeutic considerations” Frontiers in Aging, (2022), Article931331S. Michelle et al. “Plasma klotho and frailty in older adults: findings from the inchianti study” Journal of Gerontology, (2019), 74, 1052S. Begona et al. “Low serum klotho concentration is associated with worse cognition,psychological components of frailty, dependence, and falls in nursing home residents” Scientific Reports,(2021), 11, 9098H. Kelsey et al. “Exploiting the neuroprotective effects of α-klotho to tackle aging- and neurodegeneration-related cognitive “dysfunction” Neuronal Signaling, (2021), 5, NS20200101Z. Lili et al. “Klotho ameliorates kidney injury and fibrosis and normalizes blood pressure by targeting therein-angiotensin system” The American Journal of Pathology, (2015), 74, 1052M. Ali et al. “Effects of klotho supplementation on hyperoxia-induced renal injury in a rodent model of postnatal nephrogenesis” Pediatric Research, (2020), 88, 565-570X. Zhau, et al. “Parishin alleviates vascular aging in mice by upregulation of Klotho” J Cell Mol Med. (2015), 27, 1398-1409Kevin Flurkey et al. The Mouse in Biomedical Research (Second Edition) Chapter 20 - Mouse Models in Aging Research,

[0008] The object of the present invention is to provide a novel means that is effective in preventing or treating age-related declines in motor function, declines in the function of internal organs such as the liver, heart, kidneys, and spleen, and diseases induced by declines in internal organ function, as well as in suppressing the age-related decline in the blood concentration of the longevity protein α-Klotho, which suppresses the decline in the function of various organs.

[0009] In this context, the inventors diligently conducted research to find an effective composition to solve the above problems, using changes in motor function and visceral functions such as the liver, heart, kidneys, and spleen, as well as the suppression of age-related declines in α-Klotho blood concentration, as indicators. As a result, in animal studies, they found that glycyrrhizic acid derivatives also suppress age-related declines in motor function and visceral functions such as the liver, heart, kidneys, and spleen. Furthermore, they discovered for the first time that the decline in α-Klotho blood concentration is also suppressed. These findings are consistent with the results of prior art research on the aging-inhibiting properties of α-Klotho inherently present in various organs. With these findings obtained, the present invention is complete. That is, the composition of the present invention contains a glycyrrhizic acid derivative as an active ingredient, which has the function of preventing age-related declines in motor function and visceral functions such as the liver, heart, kidneys, and spleen, as well as preventing diseases that follow these declines, and suppressing the decline in α-Klotho blood concentration in the blood. The composition of the present invention may be licorice extract itself, a glycyrrhizic acid derivative extracted and purified from licorice, or a pharmaceutically acceptable salt or solvate thereof.

[0010] Furthermore, glycyrrhizic acid has isomers based on the arrangement of the hydrogen atom at position 18 in its molecule. Typically, more than 90% of the glycyrrhizic acid produced by licorice plants is the 18β form, but a small portion is the 18α form. While much of the physiological activity of the 18α form is basically the same as that of the 18β form, the specific activity of the 18α form in anti-inflammatory and anti-allergic effects is higher than that of the 18β form, leading to the creation of pharmaceuticals with the 18α form as the main component. In addition, the 18α form differs from the 18β form in terms of physical properties; specifically, the 18α form does not gel in acidic solutions like the 18β form, giving it the advantage of being able to be incorporated into relatively acidic cosmetics. Furthermore, it has been confirmed that the 18α compound exhibits weaker inhibitory activity against 11β-hydroxysteroid dehydrogenase type 2 (hereinafter referred to as "11β-HSD2"), which causes pseudoaldosteronism, due to its metabolite (glycyrrhetinic acid, or its sulfate conjugate, glycyrrhetinic acid-3-sulfate), compared to the 18β compound. Therefore, a safer pharmaceutical composition can be provided.

[0011] In other words, the present invention is not limited to the following inventions, but encompasses the following inventions.

[0012] 1-a. A pharmaceutical composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in motor function in a subject, and / or preventing or treating diseases induced by age-related decline in motor function. 1-b. A method for preventing age-related decline in motor function in a subject, and / or preventing or treating diseases induced by age-related decline in motor function, comprising the step of administering at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, to the subject.

[0013] 2-a. A pharmaceutical composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in visceral function and / or preventing or treating diseases induced by age-related decline in visceral function in a subject. 2-b. A method for preventing age-related decline in visceral function and / or preventing or treating diseases induced by age-related decline in visceral function in a subject, comprising the step of administering at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, to the subject.

[0014] 3-a. A pharmaceutical composition comprising glycyrrhizic acid and its derivatives, and at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in α-Klotho blood concentration in a subject, and / or preventing or treating diseases induced by age-related decline in α-Klotho blood concentration. 3-b. A method for preventing age-related decline in α-Klotho blood concentration in a subject, and / or preventing or treating diseases induced by age-related decline in α-Klotho blood concentration, comprising the step of administering at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, to the subject.

[0015] 4-a. The pharmaceutical composition according to any one of paragraphs 1, 2, and 3-a, wherein the glycyrrhizic acid analog of paragraphs 1, 2, and 3-a is at least one selected from the group consisting of 18β-glycyrrhizic acid, 18β-glycyrrhetinic acid, and 18β-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof. 4-b. The method according to any one of paragraphs 1, 2, and 3-b, wherein the glycyrrhizic acid analog of paragraphs 1, 2, and 3-b is at least one selected from the group consisting of 18β-glycyrrhizic acid, 18β-glycyrrhetinic acid, and 18β-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

[0016] 5-a. The pharmaceutical composition according to any one of the preceding paragraphs 1, 2, and 3-a, wherein the inhibitory activity of the glycyrrhizic acid analogs of the preceding paragraphs 1, 2, and 3-a against 11β-hydroxysteroid dehydrogenase type 2 is lower than that of 18β-glycyrrhetinic acid. 5-b. The method according to any one of the preceding paragraphs 1, 2, and 3-b, wherein the inhibitory activity of the glycyrrhizic acid analogs of the preceding paragraphs 1, 2, and 3-b against 11β-hydroxysteroid dehydrogenase type 2 is lower than that of 18β-glycyrrhetinic acid.

[0017] 6-a. The pharmaceutical composition according to any one of paragraphs 1, 2, and 3-a, wherein the glycyrrhizic acid analog of paragraphs 1, 2, and 3-a is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof. 6-b. The method according to any one of paragraphs 1, 2, and 3-b, wherein the glycyrrhizic acid analog of paragraphs 1, 2, and 3-b is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

[0018] 7-a. A pharmaceutical composition according to any one of paragraphs 1 to 6-a, wherein the subject is a human or a non-human animal. 7-b. A method comprising the step of administering a pharmaceutical composition according to any one of paragraphs 1 to 6-b, wherein the subject is a human or a non-human animal.

[0019] 8. A food composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in motor function and / or preventing or improving diseases induced by age-related decline in motor function in a subject.

[0020] 9. A food composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in organ function and / or preventing or improving diseases induced by age-related decline in organ function in a subject.

[0021] 10. A food composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in α-Klotho blood concentration and / or preventing or improving diseases associated with age-related decline in α-Klotho blood concentration in a subject.

[0022] 11. The food composition according to any one of paragraphs 8, 9, and 10, wherein the glycyrrhizic acid analog of paragraphs 8, 9, and 10 is at least one selected from the group consisting of 18β-glycyrrhizic acid, 18β-glycyrrhetinic acid, and 18β-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

[0023] 12. The food composition according to any one of paragraphs 8, 9, and 10, wherein the glycyrrhizic acid analog of paragraphs 8, 9, and 10 is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

[0024] 13. The food composition according to any one of items 8, 9, and 10, wherein the inhibitory activity of the glycyrrhizic acid analogs of items 8, 9, and 10 against 11β-hydroxysteroid dehydrogenase type 2 is lower than that of 18β-glycyrrhetinic acid.

[0025] 14. The food composition according to any one of paragraphs 8, 9, and 10, wherein the glycyrrhizic acid analog of paragraphs 8, 9, and 10 is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

[0026] 15. A food composition according to any one of items 8 to 14 above, which is ingested by humans or non-human animals.

[0027] The present invention provides a composition for preventing age-related decline in motor function and internal organ functions such as the liver, heart, kidneys, and spleen, and / or for preventing, treating, or improving diseases induced by these declines. It also provides a composition for suppressing age-related decline in α-Klotho blood concentration and / or for preventing, treating, or improving diseases induced by this decline in α-Klotho blood concentration. The composition of the present invention contains glycyrrhizic acid, which has been used for a long time, as an active ingredient, and is highly safe even when taken over a long period at normal doses. Furthermore, if the active ingredient is 18α-glycyrrhizic acid, an even safer composition can be provided.

[0028] This shows the appearance of the mice at the start of the experiment and at the end of the experiment after 12 months. This shows the results of measuring the weight of the mice over time. This shows the results of measuring the weight of the mice at the start of the experiment and at the end of the experiment after 12 months. This shows the results of measuring the amount of mouse motility using a spontaneous motility measuring device. This shows the results of measurements using a rotordo measuring device. This shows the results of measurements using a grip strength measuring device. This shows the blood cholesterol concentration of the mice. This shows the blood triglyceride concentration of the mice. This shows the weight measurement results of the mouse liver. This shows the appearance of the mouse liver. This shows the histological image of the mouse liver. This shows the weight measurement results of the mouse kidney. This shows the appearance of the mouse kidney. This shows the histological image of the mouse kidney. This shows the weight measurement results of the mouse heart. This shows the appearance of the mouse heart. This shows the weight measurement results of the mouse spleen. This shows the appearance of the mouse spleen. This shows the blood concentration of mouse α-Klotho. This shows the blood concentration of cystatin C in aging-accelerated mice, etc. This shows the blood concentration of α-Klotho in aging-accelerated mice, etc.

[0029] Embodiments of the present invention will be described in detail below. One embodiment of the present invention is a composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof. In one embodiment, the composition of the present invention is a pharmaceutical composition. In one embodiment, the composition of the present invention is a pharmaceutical composition for use in preventing age-related decline in motor function and internal organ function such as the liver, heart, kidneys, and spleen, and / or for the prevention or treatment of diseases induced by age-related decline in internal organ function. In one embodiment, the composition of the present invention is a food composition. In one embodiment, the composition of the present invention is a food composition for use in preventing age-related decline in motor function and internal organ function such as the liver, heart, and kidneys, and / or for the prevention or improvement of diseases induced by age-related decline in function.

[0030] The terms "analogs" and "derivatives" of a compound usually refer to other compounds having a structure similar to that compound. However, in this invention, the "analogs" of a compound include the compound itself, while the "derivatives" of a compound do not include the compound itself. Examples of derivatization to give glycyrrhizic acid derivatives include, but are not limited to, the introduction of substituents (hydrocarbon groups, halogen atoms, oxygen-containing functional groups (hydroxyl groups, alkoxy groups, epoxy groups, oxo groups, carboxyl groups, etc.), nitrogen-containing functional groups (unsubstituted or substituted amino groups, cyano groups, etc.), sulfur-containing functional groups (thiol groups, sulfo groups, etc.)), oxidation, reduction, esterification, amidation, isomerization (including stereoisomerization), and modification of sugar chains (including removal of sugar chains). Salts and solvates are also included as derivatives and analogs. In this invention, these glycyrrhizic acid analogs may be used individually or in combination of two or more.

[0031] In the present invention, a "pharmaceutically acceptable salt" of the glycyrrhizic acid analog refers to a salt of the glycyrrhizic acid analog that retains the biological properties (including pharmacological activity) of the glycyrrhizic acid analog and is free from any undesirable biological or other aspects. When the glycyrrhizic acid analog is a basic compound, its pharmaceutically acceptable salt can be prepared by the addition of an inorganic or organic acid. Salts derived from inorganic acids include, but are not limited to, salts with hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc. Salts derived from organic acids include, but are not limited to, salts with acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, N-acetylcysteine, etc. If the above-mentioned glycyrrhizic acid analogs are acidic compounds, their pharmaceutically acceptable salts can be prepared by addition of an inorganic or organic base. Salts derived from inorganic bases include, but are not limited to, salts with sodium, potassium, lithium, ammonium, calcium, magnesium, etc. Salts derived from organic bases include, but are not limited to, salts with primary, secondary, and tertiary amines (e.g., isopropylamine, trimethylamine, diethylamine, triethylamine, and tripropylamine), substituted amines (e.g., ethanolamine, lysine, and arginine), cyclic amines (e.g., N-ethylpiperidine and piperidine), etc.

[0032] In the present invention, the glycyrrhizic acid analog may be in the form of a pharmaceutically acceptable solvate. A "solvate" of a compound means a complex having a structure in which a stoichiometric or non-stoichiometric amount of some solvent molecule is attached to the molecule of the compound. Examples of solvents that form a solvate include, but are not limited to, water, alcohol (methanol, ethanol, n-propanol, isopropanol, etc.), DMSO, ethyl acetate, etc. A solvate in which the solvent is water is called a "hydrate." A "pharmaceutically acceptable solvate" of a compound means a solvate of the compound in which the biological properties (including pharmacological activity) of the compound are retained and which is not undesirable in any biological or other respect.

[0033] In the present invention, it is preferable that the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, 18α-glycyrrhetinic acid-3-O-monoglucuronide, and 18β-glycyrrhizic acid, as well as pharmaceutically acceptable salts and solvates thereof. Among these glycyrrhizic acid analogs, 18β-glycyrrhizic acid is the main saponin contained in the underground parts of the genus Glycyrrhiza (Glycyrrhiza uralensis, G. glabra, or G. inflata, etc.), which has been used as a medicine for a long time, and has the following formula: It has the structure represented by . The compound that has traditionally been simply called "glycyrrhizic acid" is usually this 18β-glycyrrhizic acid. Currently, (18β-)glycyrrhizic acid extracted and purified from the underground parts of licorice plants is used in pharmaceuticals indicated for anti-inflammatory, anti-allergic, or anti-chronic hepatitis, as well as in quasi-drugs and cosmetics that utilize its anti-inflammatory and anti-allergic effects. Furthermore, it is also used as a food additive that takes advantage of its sweetness.

[0034] 18α-glycyrrhizic acid is one of the stereoisomers of 18β-glycyrrhizic acid (in which the stereochemistry of the 18th carbon is reversed), and is given by the following formula: It has the structure represented by [the formula shown]. Saponins obtained from the genus Glycyrrhiza as described above may contain a large amount of 18β-glycyrrhizic acid, as well as a very small amount of 18α-glycyrrhizic acid. As described later, 18α-glycyrrhizic acid can be synthesized from 18β-glycyrrhizic acid.

[0035] 18α-Glycyrrhetinic acid is given by the following formula: It has a structure corresponding to the aglycone of 18α-glycyrrhizic acid, represented by the formula: 18α-glycyrrhizic acid can be prepared by known methods, for example, by hydrolysis of 18α-glycyrrhizic acid by heating it in a strong acid aqueous solution (e.g., hydrochloric acid). 18-α-glycyrrhetinic acid-3-O-monoglucuronide has the following formula: It has a structure represented by 18α-glycyrrhizic acid, in which one of the two glucuronic acid residues has been removed. 18α-glycyrrhetinic acid-3-O-monoglucuronide can be prepared by known methods, for example, by enzymatic hydrolysis of 18α-glycyrrhizic acid (for example, by treating 18α-glycyrrhizic acid in the culture medium of the fungus Cryptococcus magnus MG-27 strain) (T. Kuramoto et al., Microbial Production of Glycyrrhetic Acid 3-O-Mono-β-D-Glucuronide from Glycyrrhizin by Cryptococcus magnus MG-27, Bioscience, Biotechnology, and See Biochemistry (1994), 58:3, 455–458).

[0036] When glycyrrhizic acid was administered to mice for one year, compared to mice that did not receive the treatment, it was found to prevent declines in motor function and internal organ function such as the liver, heart, and kidneys, and to have a preventive effect against disease. Glycyrrhizic acid analogs are known to exhibit various useful pharmacological effects, such as anti-allergic and anti-inflammatory effects. However, the fact that glycyrrhizic acid analogs are useful in preventing age-related declines in internal organ function such as the liver, heart, kidneys, and spleen, and in preventing or treating diseases induced by such declines, was discovered for the first time by the present inventors. Therefore, in one embodiment of the present invention, the effect of glycyrrhizic acid analogs may be confirmed by measuring blood cholesterol and triglyceride levels in subjects before, after, or both administration of glycyrrhizic acid analogs. Furthermore, regarding blood components, in addition to a decrease in triglyceride and cholesterol levels, suppression of the decrease in blood concentration of α-Klotho, known as a longevity protein, was also confirmed. The α-Klotho gene (α klotho; sometimes simply referred to as klotho; HGNC: 6344 NCBI Gene: 9365 Ensemble: ENSG00000133116 OMIM(R): 604824 UniProtKB / Swiss-Prot: Q9UEF7) encodes a type I membrane protein associated with β-glucosidase. Secreted Klotho peptides, produced by cleavage of the membrane binding site by peptidase, have been reported to extend lifespan by inhibiting insulin / IGF1 signaling. Hereinafter, α-Klotho includes type I membrane proteins, secreted peptides, or both, without particular limitation. Blood levels of α-Klotho decrease with age. Originally, α-Klotho suppresses the decline in function of various organs, skeletal muscles, and bone tissue. Therefore, the age-related decrease in α-Klotho leads to an accelerated decline in physical function. Thus, drugs that maintain α-Klotho blood levels are considered to be very useful in preventing age-related problems. This effect of glycyrrhizic acid analogs was also discovered for the first time by the inventors.Therefore, in one embodiment of the present invention, the effect of the glycyrrhizic acid analog may be confirmed by measuring the α-Klotho blood concentration in the subject before, after, or both administration of the glycyrrhizic acid analog. Furthermore, a decrease in cystatin C, whose blood concentration increases with declining renal function, was observed after administration of the glycyrrhizic acid analog, thus further confirming the effect of the glycyrrhizic acid analog on maintaining renal function.

[0037] The maintenance effect of α-Klotho blood concentration by 18α-glycyrrhizic acid, a glycyrrhizic acid analog, was confirmed in administration studies to aging-accelerated mice. Specifically, when administered to aging-accelerated mice (SAMP8) for 12 weeks, it showed a maintenance effect of α-Klotho blood concentration compared to untreated mice, suggesting that it can be used in a similar way to 18β-glycyrrhizic acid.

[0038] One aspect of the present invention provides a glycyrrhizic acid analog for use in preventing age-related decline in motor function and visceral function. This glycyrrhizic acid analog also provides a method for simultaneously preventing a decrease in the blood concentration of the longevity protein α-Klotho. Another aspect of the present invention provides a pharmaceutical composition comprising the glycyrrhizic acid analog and a pharmaceutically acceptable carrier and / or a pharmaceutically acceptable excipient. Another aspect of the present invention provides the use of the glycyrrhizic acid analog for preventing age-related decline in motor function and visceral function. Another aspect of the present invention provides the use of the glycyrrhizic acid analog for the manufacture of a pharmaceutical for preventing age-related decline in motor function and visceral function such as the liver, heart, kidneys, and spleen, and for preventing or treating diseases induced by age-related decline in visceral function. Another aspect of the present invention provides a method for preventing age-related decline in motor function and visceral function such as the liver, heart, and kidneys, and for preventing or treating diseases induced by age-related decline in visceral function, comprising the step of administering the glycyrrhizic acid analog to a human or non-human animal.

[0039] In this invention, "decreased motor function" refers to, but is not limited to, a decrease in spontaneous movement, a decrease in motor coordination, fatigue tolerance, and motor learning function, and a decrease in grip strength.

[0040] In the present invention, "visceral function" refers to liver function, heart function, kidney function, and spleen function. "Decreased visceral function" refers to, but is not limited to, elevated blood cholesterol and triglyceride levels, fatty liver, cardiac hypertrophy, and nephrohypertrophy. In the present invention, "diseases induced by decreased visceral function" refers to, but is not limited to, liver fibrosis, renal fibrosis, cholangiofibrosis, non-alcoholic steatohepatitis, non-alcoholic fatty liver disease, chronic kidney disease, diabetic kidney disease, primary sclerosing cholangitis, primary biliary cirrhosis, or idiopathic fibrosis, metabolic disorders or metabolic syndrome, atherosclerosis, diabetes mellitus, hyperglycemic diabetes mellitus, type 2 diabetes mellitus, dyslipidemia, hypercholesterolemia, hyperlipidemia, hypertension, hypoglycemia, obesity, or Prader-Willi syndrome.

[0041] The carriers and excipients for preparing the pharmaceutical composition are not particularly limited, and conventionally known ones can be used. The pharmaceutical composition may optionally contain conventionally known additives such as binders, disintegrants, coating agents, lubricants, preservatives, stabilizers, buffers, surfactants, antioxidants, isotonic agents, and colorants.

[0042] In the present invention, from the viewpoint of safety, the glycyrrhizic acid analog preferably contains at least one selected from the group consisting of glycyrrhetinic acid, 18α-glycyrrhetinic acid, 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof, particularly preferably 18α-glycyrrhetinic acid. 18β-glycyrrhetinic acid is known to be absorbed by the living body after being metabolized to its aglycone, 18β-glycyrrhetinic acid. Similarly, 18α-glycyrrhetinic acid and 18α-glycyrrhetinic acid-3-O-monoglucuronide are presumed to be absorbed by the living body after being metabolized to their aglycone, 18α-glycyrrhetinic acid. Both 18α-glycyrrhetinic acid and 18β-glycyrrhetinic acid exhibit inhibitory activity against 11β-HSD2. Pseudohyperaldosteronism, known as a side effect of pharmaceuticals containing licorice components, is caused by this inhibitory activity. The inhibitory activity of 18α-glycyrrhetinic acid is lower than that of 18β-glycyrrhetinic acid. Therefore, the above three glycyrrhizic acid analogs are superior in safety to 18β-glycyrrhetinic acid in that they are less likely to cause pseudohyperaldosteronism (Patent Document 2).

[0043] On the other hand, from the viewpoints of availability and further cost, in the present invention, the glycyrrhizic acid analog preferably contains at least one selected from the group consisting of 18β-glycyrrhetinic acid and pharmaceutically acceptable salts and solvates thereof, particularly preferably 18β-glycyrrhetinic acid.

[0044] In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof. In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18β-glycyrrhizic acid, and pharmaceutically acceptable salts and solvates thereof. In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid and 18α-glycyrrhetinic acid, and pharmaceutically acceptable salts and solvates thereof. In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid and 18β-glycyrrhizic acid, and pharmaceutically acceptable salts and solvates thereof. In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid, and pharmaceutically acceptable salts and solvates thereof. In another embodiment, the glycyrrhizic acid analog is at least one selected from the group consisting of 18β-glycyrrhizic acid, and pharmaceutically acceptable salts and solvates thereof.

[0045] Glycyrrhizic acid analogs themselves, or licorice extract containing them, can also be used as part of the composition of the present invention. The glycyrrhizic acid analogs used in the present invention can be obtained by extracting, isolating, and purifying them from licorice extract using a general method. That is, after removing lipid-soluble coloring components from licorice extract by organic solvent extraction, a purified product with a purity of 70% or more can be obtained by purifying it using means such as crystallization. On the other hand, when using licorice extract as part of the composition of the present invention, it can be extracted from the roots and / or stems of the plant using a general method known in the art. In the present invention, commercially available licorice extract can also be used. The content of glycyrrhizic acid analogs in the licorice extract used in the present invention is 5% or more, preferably 10% or more, and more preferably 13% or more.

[0046] The extraction solvent used in the production of licorice extract is not particularly limited. For example, water, alcohol (such as methanol, ethanol, n-propanol, n-butanol, etc.), acetone, ethyl acetate, etc. can be used. When obtaining the dry powder of licorice extract, for example, by methods commonly used by those skilled in the art such as vacuum drying or spray drying, the solvent of the extract can be removed to obtain it.

[0047] Use of the composition of the present invention for application to a living body As uses of the composition of the present invention, pharmaceuticals, quasi-drugs, cosmetics, foods, etc. can be mentioned. The application target of the composition of the present invention is a human or a non-human animal, preferably a human. In one embodiment, the pharmaceutical composition of the present invention is administered to a human. In one embodiment, the food composition of the present invention is ingested by a human. Although not particularly limited, in the case of humans, it is preferably administered during the mature period (20 to 30 years old), the middle-aged period (38 to 47 years old), or in between. This is because if the decline in motor function and visceral function can be suppressed before reaching old age, a healthy old age can be spent.

[0048] The form of the composition of the present invention is not particularly limited, and this composition can take a commonly used form according to its use. The form of the pharmaceutical composition of the present invention used as a pharmaceutical is roughly classified into a form suitable for oral administration and a form suitable for parenteral administration. Examples of the former include tablets, troches, pills, suspensions (aqueous or oily), powders, granules, emulsions, hard or soft capsules (for example, gelatin capsules), syrups, elixirs, etc. Examples of the latter include injections, creams, ointments, inhalants, eye drops, nasal drops, suppositories, etc. The pharmaceutical composition of the present invention used as a quasi-drug (including medicinal cosmetics) may be in various forms of conventional cosmetics in addition to dentifrices, etc. The food composition of the present invention may be in various forms of conventional use.

[0049] The dosage of glycyrrhizic acid analogs contained in the composition of the present invention is not particularly limited, but may be 5 to 120 mg / day / person, preferably 10 to 60 mg / day / person. In this case, the dosage may be divided and administered two to three times a day. Furthermore, when licorice extract is used as the composition of the present invention, the dosage of glycyrrhizic acid analogs contained therein may be adjusted to be the same as above, taking into consideration its glycyrrhizic acid analog content.

[0050] The present invention will be described below with reference to examples, but the present invention is not limited to these examples.

[0051] The outline of the method for measuring the anti-aging effect of the test substance is as follows: Ten-week-old ICR mice were used as test animals and raised for one year. These mice were divided into an experimental group and a control group, and the mice in the experimental group were orally administered 18β-glycyrrhizic acid at a dose of 15 mg / kg three times a week. Water and food were provided as usual. The following measurements and tests were performed on these animals at the start of the study (10 weeks old) and at the end of the study (12 months old). It should be noted that 10 weeks old in experimental mice corresponds to pre-maturity in humans (i.e., under 20 years old); and 12 months old in mice corresponds to middle age in humans (36-47 years old, preferably 42.5 years old) (Non-patent Literature 9).

[0052] Appearance and Weight Observation of Appearance at the Start (10 weeks of age) and End (12 months of age) of the Study showed that obesity was suppressed in mice treated with glycyrrhizic acid compared to mice not treated with glycyrrhizic acid (Figure 1). Furthermore, when the body weight of the above ICR mice was measured every four weeks from 10 weeks of age to 52 weeks of age, the body weight of the mice treated with glycyrrhizic acid was consistently lower than that of the mice not treated with glycyrrhizic acid (Figure 2), and ultimately, there was a difference of 20g per individual, or about 25%, between the glycyrrhizic acid-treated group and the non-treated group (Figure 3). Considering that triglyceride and cholesterol levels in the blood were also lower in the glycyrrhizic acid-treated group, and that the non-treated group showed symptoms of fatty liver, these results indicate that obesity is suppressed by the administration of glycyrrhizic acid.

[0053] Spontaneous motility was measured using a spontaneous motility measurement device, where an infrared sensor attached to the bottom of the cage quantified the amount of activity. Each test mouse was placed in a new environment individually (without acclimatization), and their spontaneous motility was measured for 20 minutes. As shown in Figure 4, the group administered glycyrrhizic acid had approximately 55% more spontaneous motility. This result indicates that the age-related decline in motility was suppressed.

[0054] In the rotorod test, mice were placed on a rotating rod using a rotorod test apparatus, and the time it took for the mice to fall off the rod was measured. As shown in Figure 5, the glycyrrhizic acid-administered group remained on the rotorod for 12 seconds, which was approximately four times longer than the 3 seconds in the non-administered group. This result indicates that glycyrrhizic acid suppresses the age-related decline in balance, which is an important element of motor function.

[0055] Grip Strength Test: The grip strength of mice was measured using a grip strength measurement device. As shown in Figure 6, the group treated with glycyrrhizic acid maintained approximately 17% higher grip strength than the group not treated with glycyrrhizic acid.

[0056] Blood samples were collected from mice in the glycyrrhizic acid-administered and non-administered groups at 10 weeks and 12 months after the start of the blood cholesterol and triglyceride level study, and blood cholesterol and triglyceride levels were measured. In the glycyrrhizic acid-administered group, blood cholesterol levels were approximately 22% lower and triglyceride levels were approximately 27% lower than in the non-administered group (Figures 7 and 8).

[0057] Liver Weight and Histological Features The liver weight and histological features of mice in the glycyrrhizic acid-administered and unadministered groups were examined at 10 weeks and 12 months after the start of the study. As shown in Figure 9, the liver weight of the unadministered group was greater than that of the glycyrrhizic acid-administered group, and fat accumulation in the tissue was observed (Figure 10). Furthermore, observation of tissue sections revealed many cells with fat accumulation, and some cells had lost their nuclei due to significant accumulation. This reflects the high blood triglyceride and cholesterol levels in the unadministered group, which induced fatty liver and is also indicative of non-alcoholic steatohepatitis (NAFLD). In contrast, the livers of the glycyrrhizic acid-administered group were smaller in weight, and the histological features were healthy, similar to the state at the start of the study (Figure 11). This reflects the low blood cholesterol and triglyceride levels described in the previous section.

[0058] Kidney weight and histological features were measured and examined in mice administered glycyrrhizic acid and those not administered it at 10 weeks and 12 months after the start of the kidney weight and histological feature study. As shown in Figures 12 and 13, the kidneys in the glycyrrhizic acid-administered group were smaller in appearance and weighed approximately 17% less than those in the non-administered group. Furthermore, histological examination revealed that the non-glycyrrhizic acid-administered group showed significant aging changes, namely, abnormal tubules with low cytoplasmic staining and enlarged nuclei, deposition of unstructured material, lymphocyte infiltration, and scattered cells with accumulation of granules presumed to be stress granules. These are symptoms similar to chronic kidney disease. In contrast, the glycyrrhizic acid-administered group showed less accumulation of such waste products and only a small distribution of granules. Moreover, it was found that there were many cells exhibiting healthy staining. These results indicate a lower degree of aging in the glycyrrhizic acid-administered group (Figure 14).

[0059] The cardiac weights of mice were measured at 10 weeks and 12 months after the start of cardiac weight testing in both the glycyrrhizic acid-administered and non-administered groups. As shown in Figures 15 and 16, the non-glycyrrhizic acid-administered group had a cardiac weight approximately 25% greater than the administered group, showing signs of cardiac hypertrophy, whereas this was not the case in the administered group.

[0060] Spleen weight and histological features were measured and examined in mice administered glycyrrhizic acid and those not administered glycyrrhizic acid at 10 weeks and 12 months after the start of the study. As shown in Figures 17 and 18, the non-glycyrrhizic acid group showed an appearance suggestive of enlargement and an increase in weight (approximately 53%). On the other hand, these symptoms were not observed in the administered group.

[0061] Blood samples were collected from mice in the glycyrrhizic acid-administered and non-administered groups at the start of the α-Klotho blood concentration test and 12 months later. Blood α-Klotho levels were measured using the sandwich ELISA principle. The specific measurement method is as follows: Mouse blood was added to the wells of a micro-ELISA plate pre-coated with a capture antibody specific to mouse α-Klotho, and incubated at 37°C for 90 minutes to allow the antibody to bind. Next, a conjugate of a biotinylation detection antibody specific to mouse α-Klotho and avidin-horseradish peroxidase (HRP) was sequentially added to each microplate well and incubated at 37°C for 1 hour. Furthermore, when the peroxidase substrate solution was added to each well, only the wells containing mouse α-Klotho, biotinylation detection antibody, and avidin-HRP conjugate turned blue. The enzyme-substrate reaction was stopped by adding a stop solution, and the color turned yellow. The absorbance (OD) of this yellow color was measured at a wavelength of 450 ± 2 nm. Since the OD value is proportional to the concentration of mouse α-Klotho, the concentration of mouse α-Klotho in the blood sample was calculated by comparing the OD of the sample with the standard curve. As shown in Figure 19, the α-Klotho blood concentration was approximately 22% higher in the glycyrrhizic acid administration group than in the non-administration group. This value was close to that of young mice (10 weeks old) at the start of the study.

[0062] Blood samples were collected from mice in the glycyrrhizic acid-treated group and the non-treated group 10 weeks and 12 months after the start of the cystatin C level test, and cystatin C levels were measured. The glycyrrhizic acid-treated group had approximately 37% lower levels than the non-treated group (Figure 20).

[0063] The following is an outline of the measurement method for the effect of the test substance on preventing the decline in α-Klotho levels in aging-accelerated mice. Specifically, 40-week-old SAMP8 mice were used as test animals and were reared for 12 weeks. SAMR1 mice were used as the control group. SAMP8 mice and SAMR1 mice (both available from Nippon SLC Co., Ltd.) are often used in animal experiments on age-related decline in learning and memory abilities, with the latter serving as the control. SAMP8 mice are an established aging-accelerated mouse model that shows age-related learning and memory impairments, emotional disorders, etc., earlier than normal mice. SAMR1 mice originate from the same mouse colony as SAMP8 mice, but show a normal aging profile. These mice were divided into experimental and control groups, and the experimental group mice were orally administered 15 mg / kg of 18α-glycyrrhizic acid or 18β-glycyrrhizic acid three times a week. Water and food were provided as usual. The following measurements and tests were performed on these test animals. Blood samples were collected from mice in the glycyrrhizic acid administration group and the non-administration group 12 weeks after the start of the experiment, and α-Klotho blood concentrations were measured. As shown in Figure 20, the glycyrrhizic acid administration group showed higher α-Klotho blood concentrations in approximately 90% of 18α-forms and approximately 69% of 18β-forms compared to the non-administration group, and it was found that the same was true in aging-accelerated mice (Figure 21).

[0064] The composition of the present invention, which contains a glycyrrhizic acid analog, is useful in preventing age-related declines in motor function and internal organ functions such as the liver, heart, and kidneys in animals, particularly humans, and is also useful in preventing and treating diseases caused by these declines. It is also useful in maintaining the blood concentration of the longevity protein α-Klotho, which similarly declines with age.

[0065] The disclosure of Japanese Patent Application No. 2025-007094 (filing date: January 17, 2025) is incorporated herein by reference in its entirety. All documents, patent applications, and technical standards described herein are incorporated herein by reference to the same extent as if each individual document, patent application, and technical standard were specifically and individually noted to be incorporated by reference.

Claims

1. A pharmaceutical composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in motor function and / or preventing or treating diseases induced by age-related decline in motor function in a subject.

2. A pharmaceutical composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in visceral function and / or preventing or treating diseases induced by age-related decline in visceral function in a subject.

3. A pharmaceutical composition comprising glycyrrhizic acid and its derivatives, and at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its salts and solvates that are pharmaceutically acceptable, for use in preventing age-related decline in α-Klotho blood concentration in a subject, and / or preventing or treating diseases induced by age-related decline in α-Klotho blood concentration.

4. The pharmaceutical composition according to any one of claims 1, 2, and 3, wherein the glycyrrhizic acid analog is at least one selected from the group consisting of 18β-glycyrrhizic acid, 18β-glycyrrhetinic acid, and 18β-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

5. The pharmaceutical composition according to any one of claims 1, 2, and 3, wherein the glycyrrhizic acid analog is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

6. The pharmaceutical composition according to any one of claims 1 to 5, wherein the subject is a human or a non-human animal.

7. A food composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in motor function and / or preventing or improving diseases induced by age-related decline in motor function in a subject.

8. A food composition comprising at least one glycyrrhizic acid analog selected from the group consisting of glycyrrhizic acid and its derivatives, and pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in organ function and / or preventing or improving diseases induced by age-related decline in organ function in a subject.

9. A food composition comprising glycyrrhizic acid and its derivatives, and at least one glycyrrhizic acid analog selected from the group consisting of pharmaceutically acceptable salts and solvates thereof, for use in preventing age-related decline in α-Klotho blood concentration and / or preventing or improving diseases induced by age-related decline in α-Klotho blood concentration in a subject.

10. The food composition according to any one of claims 7, 8, and 9, wherein the glycyrrhizic acid analog of claims 7, 8, and 9 is at least one selected from the group consisting of 18β-glycyrrhizic acid, 18β-glycyrrhetinic acid, and 18β-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

11. The food composition according to any one of claims 7, 8, and 9, wherein the glycyrrhizic acid analog of claims 7, 8, and 9 is at least one selected from the group consisting of 18α-glycyrrhizic acid, 18α-glycyrrhetinic acid, and 18α-glycyrrhetinic acid-3-O-monoglucuronide, and pharmaceutically acceptable salts and solvates thereof.

12. A food composition according to any one of claims 7 to 11, which is ingested by humans or non-human animals.

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