Use of glycopeptide of medlar in preparation of medicine for preventing or treating presbycusis

Abstract

The present application belongs to the technical field of biological medicine, and particularly relates to the use of glycopeptide of medlar in preparation of a medicine for preventing or treating presbycusis. Experimental results show that the glycopeptide of medlar has a protective effect on hearing loss of a mouse in a rapid aging mouse model. In addition, the glycopeptide of medlar has a protective effect on cell aging of a hair cell in a mouse cochlea cultured in vitro and a HEI-OC1 mouse cochlea hair cell line. It can be seen that the glycopeptide of medlar, as a traditional Chinese medicinal material extraction product, can be effectively used for prevention and treatment of presbycusis, has a prospect of being developed into a medicine for preventing and treating presbycusis, provides more treatment options for preventing or treating presbycusis in clinic, and has important social benefits and economic value.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the use of Lycium barbarum glycopeptides in the preparation of drugs for the prevention or treatment of age-related hearing loss. Background Technology

[0002] Hearing impairment is one of the most prevalent ear diseases worldwide. According to statistics from the World Health Organization (WHO), approximately 466 million people globally currently suffer from hearing impairment, a number projected to rise to over 900 million by 2050. With an aging population, the overuse of ototoxic drugs, noise pollution, and environmental pollution, the number of people experiencing deafness and hearing loss is increasing annually. Deafness has become a global health problem affecting society and the economy. Sensorineural hearing loss accounts for approximately 63% of all hearing loss cases, and damage to cochlear hair cells is one of the main causes of sensorineural hearing loss. Currently, while hearing aids and cochlear implants, commonly used in clinical practice, improve hearing to some extent, they have limitations and cannot fundamentally solve the problem. Furthermore, their effectiveness depends entirely on the quantity and quality of hair cells. Therefore, identifying the causes and mechanisms of hair cell damage is crucial for the prevention and treatment of sensorineural hearing loss.

[0003] Presbycusis, also known as age-related hearing loss (AHL), is a complex degenerative disease in which hearing declines symmetrically and progressively in both ears with increasing age. It is characterized by a decline in auditory function, including an increase in hearing threshold and a decrease in frequency. The World Health Organization (WHO) estimates that by 2025, the population aged 60 and over will reach 1.2 billion, and an estimated 500 million of these individuals will suffer from presbycusis. Presbycusis has serious consequences for the physical, cognitive, emotional, behavioral, and social health of older adults, and can lead to social isolation, depression, and low self-esteem.

[0004] Age-related hearing loss (AHL) is a complex, multifactorial disease, with contributing factors including environment, genetics, health, and nutrition. Loss of cochlear hair cells or degeneration of the spiral ganglion are the main pathological features of AHL. The primary pathological change in the cochlea during AHL is the loss of hair cells, especially the outer hair cells of the parietal rotation, with the loss extending from the parietal to the posterior rotation and from the outer hair cells to the inner hair cells. In mammals, this hair cell loss is irreversible, and the molecular basis of hair cell death associated with AHL is not yet fully understood. Given the serious prevalence of AHL, developing effective protective drugs has become a key focus of auditory research in recent years. However, there are currently no effective treatments or drugs to prevent or treat AHL clinically.

[0005] Goji berries are the dried, ripe fruit of *Lycium barbarum* L., a plant in the Solanaceae family. They are believed to nourish the liver and kidneys, improve eyesight, and alleviate symptoms such as weakness, fatigue, lower back and knee pain, dizziness, and tinnitus. Goji berry polysaccharides are natural, large-molecule, water-soluble polysaccharides extracted from goji berries. Studies have shown that goji berry polysaccharides possess pharmacological effects such as immunomodulation, hypoglycemia, lipid reduction, anti-aging, antioxidant, and anti-tumor activity. Other studies have indicated that goji berry polysaccharides can protect against neuronal cell damage and exert neuroprotective effects such as antidepressant effects. In recent years, goji berry glycopeptide (LBGP), extracted from goji berry polysaccharides, has attracted attention. Compared to goji berry polysaccharides, LBGP has removed inorganic salts and various monosaccharides, and studies have shown that LBGP has strong immune activity and anti-aging effects.

[0006] Currently, there are no reports of wolfberry glycopeptides being used to prevent or treat age-related hearing loss. Summary of the Invention

[0007] To address the current shortage of effective drugs for the prevention or treatment of age-related hearing loss in clinical use, the inventors have employed modern pharmaceutical research methods to conduct in-depth development and utilization of natural products. Combined with extensive pharmacodynamic experiments, this invention provides the use of Lycium barbarum glycopeptides in the preparation of drugs for the prevention or treatment of age-related hearing loss.

[0008] Specifically, the present invention is achieved through the following technical solutions: This invention provides the use of Lycium barbarum glycopeptides in the preparation of medicaments for the prevention or treatment of age-related hearing loss.

[0009] Preferably, the present invention provides the use of Lycium barbarum glycopeptide as the sole active ingredient in the preparation of a medicament for the prevention or treatment of age-related hearing loss.

[0010] Alternatively, in the above-described uses, the symptoms of age-related hearing loss may include one or more of the following: tinnitus, hearing loss, or hearing impairment.

[0011] Alternatively, in the above-described uses, the goji berry glycopeptide reduces the upward shift of the hearing threshold in patients with age-related hearing loss and improves the survival rate of cochlear hair cells.

[0012] Alternatively, in the above-mentioned applications, the goji berry glycopeptide contains ≥60% sugar, ≥5% uronic acid, and 20-33% protein by weight percentage, and the goji berry glycopeptide is in the form of a powder with a purity of ≥90%.

[0013] Alternatively, in the above-mentioned uses, the molecular weight of the wolfberry glycopeptide is determined by SDS-PAGE to be 88 kD, the sugar content is 70% by weight, and the sugar composition is ArA:GA1:Glc = 2.5:1.0:1.0 by molar ratio.

[0014] Alternatively, in the above-described uses, the medicament comprises a preventive or therapeutically effective amount of lycine glycopeptide and a pharmaceutically acceptable carrier.

[0015] Alternatively, in the above-described uses, the drug may be an oral preparation, an injection, or a topical preparation.

[0016] Preferably, the oral preparation is selected from oral liquid, tablets, powder, capsules or granules.

[0017] Preferably, the injection is selected from aqueous injections or sterile powders for injection.

[0018] Preferably, the topical preparation is selected from drops, sprays, or gels.

[0019] Alternatively, in the above-described uses, the drug is suitable for various administration methods, including oral administration, parenteral administration, or intraocular administration.

[0020] Preferably, the parenteral administration includes intravenous injection, intramuscular injection, and subcutaneous injection.

[0021] Compared with the prior art, the present invention has the following advantages: This invention, leveraging my country's strengths in natural product research, has for the first time discovered a novel application of the natural product Lycium barbarum glycopeptide in the prevention or treatment of age-related hearing loss. Experimental results show that Lycium barbarum glycopeptide has a protective effect against hearing loss in a rapidly aging mouse model. Furthermore, Lycium barbarum glycopeptide has a protective effect against cellular senescence in cochlear hair cells from cultured mice and in the HEI-OC1 mouse cochlear hair cell line. Therefore, Lycium barbarum glycopeptide, as an extract from a traditional Chinese medicine, can be effectively used for the prevention and treatment of age-related hearing loss, showing promise for development into drugs for the prevention and treatment of age-related hearing loss. This provides more treatment options for the clinical prevention and treatment of age-related hearing loss, and has significant social and economic value. Detailed Implementation

[0022] The inventors employed modern pharmaceutical research methods to deeply develop and utilize natural products. Through extensive screening, they discovered for the first time that wolfberry glycopeptides can prevent or treat age-related hearing loss. Based on this discovery, the present invention was completed.

[0023] The preferred dosage form of the drug of the present invention is an oral preparation, an injection, or a topical preparation.

[0024] Preferably, the oral preparation is selected from oral liquids, tablets, powders, capsules, or granules. More preferably, the injectable preparation is selected from aqueous injections or sterile powders for injection. More preferably, the topical preparation is selected from drops, sprays, or gels.

[0025] The drug of this invention is suitable for various routes of administration, including oral, parenteral, and intraauricular administration. Parenteral administration includes intravenous injection, intramuscular injection, and subcutaneous injection.

[0026] The pharmaceutically acceptable carrier refers to a conventional drug carrier in the field of pharmaceutical formulations, selected from one or more of fillers, binders, disintegrants, lubricants, suspending agents, wetting agents, pigments, flavoring agents, solvents, and surfactants.

[0027] The fillers described in this invention include, but are not limited to, starch, microcrystalline cellulose, sucrose, dextrin, lactose, powdered sugar, glucose, etc.; the lubricants include, but are not limited to, magnesium stearate, stearic acid, sodium chloride, sodium oleate, sodium lauryl sulfate, poloxamer, etc.; the binders include, but are not limited to, water, ethanol, starch paste, syrup, hydroxypropyl methylcellulose, sodium carboxymethyl cellulose, sodium alginate, polyvinylpyrrolidone, etc.; the disintegrants include, but are not limited to, starch effervescent mixtures, i.e., sodium bicarbonate and citric acid, tartaric acid, low-substituted hydroxypropyl cellulose, etc.; the suspending agents include, but are not limited to, polysaccharides such as farnesian gum, agar, alginic acid, cellulose ether, and carboxymethyl chitosan, etc.; and the solvents include, but are not limited to, water, balanced salt solutions, etc.

[0028] The above dosage forms can be prepared using conventional processes in the pharmaceutical formulation field.

[0029] "Goji berry glycopeptides" can be extracted and isolated from plants such as goji berries containing this active ingredient using conventional natural product extraction methods in this field, or they can be purchased from commercially available products. Specifically, goji berry glycopeptides can be prepared using the preparation method described in CN107021995B. As the most effective active ingredient in goji berry pulp, goji berry glycopeptides are the essence of goji berries. Modern pharmacological experiments have fully confirmed that goji berry glycopeptides can significantly improve the body's immune function, participate in the immune regulation of organisms, and participate in various activities of living cells. They have certain effects such as lowering blood pressure, lowering blood lipids, lowering blood sugar, anti-inflammation, anti-oxidation, anti-aging, and anti-tumor effects. The high biological activity of goji berry glycopeptides makes them hundreds of times more immunomodulatory than commonly used oral polysaccharides in clinical practice. Further experiments have also demonstrated that goji berry glycopeptides possess various pharmacological activities such as anti-oxidation, anti-aging, immunomodulation, and inhibition of tumor cell replication.

[0030] In the pharmaceutical applications described herein, the timing, frequency, and duration of administration of the wolfberry glycopeptide should be determined based on the specific diagnostic results of the condition, which is within the technical scope of those skilled in the art.

[0031] Applying treatment plans used on animals to humans allows for the conversion of the effective human dose of all drugs to the effective animal dose, which is easily achievable for those skilled in the art.

[0032] To better understand the essence of the present invention, the following detailed description section uses pharmacodynamic experiments and their results to further illustrate the novel use of Lycium barbarum glycopeptides in the pharmaceutical field for the prevention or treatment of age-related hearing loss.

[0033] The present invention will be further described below with reference to specific embodiments. It should be understood that the specific embodiments described herein are for illustrative purposes only and are not intended to limit the scope of the invention.

[0034] Where specific techniques or conditions are not specified in the examples, they shall be performed in accordance with the techniques or conditions described in the literature in this field, or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels.

[0035] Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods. Unless otherwise specified, the experimental materials used in the following embodiments are commercially available products.

[0036] Example 1: Protective effect of Lycium barbarum glycopeptides on hearing loss in a rapidly aging mouse model. 1. Animal grouping and experimentation Samp8 mice (purchased from Hangzhou Ziyuan Experimental Animal Technology Co., Ltd.), a rapid aging model, were randomly divided into two groups: the Samp8 control group and the Lycium barbarum glycopeptide administration group (20 mg / kg). The Lycium barbarum glycopeptide group was administered the drug via gavage starting on day 30. Auditory brainstem response (ABR) tests were performed on days 30, 45, 60, 75, 90, 105, 120, 135, 150, 165, and 180.

[0037] 2. Auditory brainstem response (ABR) Mice were anesthetized by intraperitoneal injection of sodium pentobarbital based on body weight. Anesthetized mice were placed in a shielded room with a 37°C constant-temperature pad. A detection electrode was inserted subcutaneously at the center of the mouse's head, a reference electrode was inserted subcutaneously below the left ear, and a grounding electrode was inserted subcutaneously on the lateral thigh. An acoustic device was placed at the mouse's left ear, and stimulation was provided with pure tones at 4kHz, 8kHz, 12kHz, 16kHz, 24kHz, and 32kHz. The sound intensity was gradually decreased from 90dB to 10dB until no repeatable waveform was recorded. The intensity at which a repeatable waveform was recorded was taken as the mouse's hearing threshold.

[0038] 3. Immunofluorescence counting of mouse cochlear hair cells Mice were euthanized and decapitated quickly. The auditory bulla was opened, and the obtained cochlear tissue was fixed in 4% paraformaldehyde (PFA) for 24 hours, followed by EDTA decalcification. After complete decalcification, the cochlea was placed in PBS, and the basement membrane was dissected under a dissecting microscope. Immunofluorescence staining was performed using hair cell markers (Myosin 7a) and phalloidin (Phalloidine). The cochlear hair cells were then counted and statistically analyzed under a ZEISS 900 laser confocal microscope.

[0039] Statistical analysis methods: Graphpad Prism 9.0 software was used to calculate statistically significant differences through one-way ANOVA and Dunnett's test. A p-value <0.05 was considered statistically significant.

[0040] Example 2: Protective effect of Lycium barbarum glycopeptides on cellular senescence of hair cells in cultured mouse cochlea Experimental methods: The basement membrane of P3 C57BL / 6J mice was cultured. The basement membrane was damaged by D-gal at concentrations of 10 mg / mL, 20 mg / mL, 40 mg / mL, 80 mg / mL and 100 mg / mL for 72 h. The cultured cochlea was then collected. The number of hair cells was detected by immunofluorescence Myosin7a staining, the changes in senescence marker proteins were detected by Western blot, and the mitochondrial DNA copy number was detected by real-time PCR to determine the senescence of hair cells.

[0041] Example 3: Protective effect of Lycium barbarum glycopeptides on cellular senescence in HEI-OC1 mouse cochlear hair cell line Experimental methods: HEI-OC1 cell lines were treated with D-gal at concentrations of 1 mg / mL, 2 mg / mL, 5 mg / mL, 10 mg / mL, and 15 mg / mL. Cell death was detected by CCK-8 and TUNEL assays. Cell senescence was assessed by galactosidase staining to select appropriate D-gal concentrations for subsequent experiments.

[0042] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims of this invention and their equivalents, this invention also intends to include these modifications and variations.

Claims

1. The use of Lycium barbarum glycopeptides in the preparation of drugs for the prevention or treatment of age-related hearing loss.

2. According to the use described in claim 1, the symptoms of age-related hearing loss include one or more of the following: tinnitus, hearing loss, or hearing impairment.

3. The use according to claim 1 or 2, characterized in that: By weight percentage, the goji berry glycopeptide contains ≥60% sugar, ≥5% uronic acid, and 20-33% protein, and the goji berry glycopeptide is in the form of a powder with a purity of ≥90%.

4. The use according to claim 1, characterized in that: The drug contains a preventive or therapeutically effective amount of goji berry glycopeptide and a pharmaceutically acceptable carrier.

5. The use according to claim 4, characterized in that: The drug is an oral preparation, an injection, or a topical preparation.

6. The use according to claim 5, characterized in that: The oral preparation is selected from oral liquids, tablets, powders, capsules, or granules; the injectable preparation is selected from water-based injections or sterile powders for injection; the topical preparation is selected from drops, sprays, or gels.

7. The use according to any one of claims 4 to 6, characterized in that: The drug is suitable for oral, parenteral, or intraoral administration.