Use of methyl jasmonate in the preparation of a drug for delaying age-related hearing loss

Abstract

This invention relates to the field of novel pharmaceutical uses, disclosing the application of methyl jasmonate in the preparation of drugs for delaying age-related hearing loss. This invention is the first to clearly discover that methyl jasmonate can significantly improve the auditory function of aged mice, significantly enhance the pure tone hearing threshold and speech recognition ability of aged mice, effectively protect the structural integrity of the cochlea, reduce apoptosis of cochlear hair cells and spiral ganglion cells, inhibit the inflammatory response and oxidative stress damage of the auditory system, and delay the degenerative process of the auditory system from a pathological perspective, which has important clinical translational potential and market value.

Description

Technical Field

[0001] This invention relates to the field of novel uses of pharmaceuticals, specifically to the use of methyl jasmonate in the preparation of drugs for delaying age-related hearing loss. Background Technology

[0002] Age-related hearing loss (ARHL), clinically known as presbycusis, is a progressive degenerative disease of the auditory system that occurs with the aging process. It is one of the most common sensory impairments in the elderly, and its onset is closely related to aging. It is also influenced by a variety of factors, including genetic susceptibility, noise exposure, use of ototoxic drugs, metabolic diseases (such as hypertension, diabetes, and hyperlipidemia), and unhealthy lifestyle habits. The pathological features of ARHL mainly include damage to cochlear hair cells, apoptosis of spiral ganglion cells, atrophy of the stria vascularis, and decline in the function of the auditory central conduction pathway. The typical clinical symptom is progressive hearing loss, initially manifesting as high-frequency hearing loss, gradually affecting all frequencies. Patients often experience the phenomenon of "hearing but not understanding clearly," with a significant decline in speech recognition ability in noisy environments. It is also often accompanied by persistent high-pitched tinnitus, which can severely affect sleep quality, creating a vicious cycle of "tinnitus-sleep disorder-further hearing loss." ARHL not only causes hearing and speech communication disorders, but also triggers mental and psychological problems such as loneliness, anxiety, and depression in the elderly. It is even closely related to cognitive decline, dementia and other diseases, further exacerbating social isolation and reducing the quality of life of the elderly.

[0003] Currently, there are no approved drugs worldwide that can effectively slow the progression of ARHL or reverse the degenerative changes in the auditory system. Routine clinical interventions primarily focus on symptom compensation, including hearing aid fitting, cochlear implantation, and basic lifestyle interventions and management of the underlying disease. Hearing aids are the preferred intervention for patients with mild to moderate ARHL, amplifying sound signals to compensate for hearing loss and help improve speech recognition. However, rigorous medical evaluation and audiological examinations are required before fitting, including pure-tone audiometry, acoustic impedance testing, and speech audiometry. Personalized fitting based on the patient's hearing loss characteristics is also essential; otherwise, improper fitting may worsen auditory burden. Cochlear implantation is mainly suitable for patients with severe to profound ARHL. It involves implanting an electronic device to directly stimulate the auditory nerve, helping patients regain some auditory function. However, this method is invasive, carries surgical risks, and requires long-term auditory rehabilitation training post-surgery. Furthermore, its accessibility is limited in rural and remote areas due to the distribution of medical resources.

[0004] In addition to the core interventions mentioned above, clinical practice also includes auxiliary interventions for the accompanying symptoms and underlying diseases of ARHL patients. For example, patients with tinnitus may use medications such as ginkgo biloba extract to relieve symptoms, and patients with comorbid chronic diseases such as hypertension and diabetes may actively treat their primary diseases to reduce further damage to the auditory system.

[0005] Current conventional treatments for ARHL have significant limitations, with core issues concentrated in three areas: First, the treatment philosophy is limited. Existing interventions are all symptomatic compensation rather than etiological treatment, unable to reverse or prevent damage and apoptosis of key auditory cells such as cochlear hair cells and spiral ganglion cells, nor can they slow down the degenerative changes in the auditory system, making it difficult to fundamentally address the root cause of ARHL. Second, drug development is lagging. To date, there are still no approved drugs worldwide that can effectively slow the progression of ARHL. Although some traditional Chinese medicine formulas and neuroprotective agents are undergoing clinical trials, no conclusive data has been published, and the field of drug intervention is still in the exploratory stage. Third, the accessibility of intervention is uneven. High-quality hearing intervention resources (such as professional fitting institutions and cochlear implant surgery teams) are mostly concentrated in first- and second-tier cities. Patients in rural and remote areas face problems such as inconvenient fitting, high surgical costs, and a lack of rehabilitation resources. At the same time, some elderly people lack awareness of ARHL and neglect early screening and intervention, resulting in missing the best intervention opportunity.

[0006] Therefore, finding candidate drugs that can target and intervene in the progression of ARHL, protect auditory structures, and improve auditory function has become an urgent task in the field of auditory medicine. Summary of the Invention

[0007] The present invention aims to provide the application of methyl jasmonate in the preparation of drugs for delaying age-related hearing loss, in order to solve the problem of the lack of effective drugs for treating age-related hearing loss in the prior art.

[0008] To achieve the above objectives, the present invention adopts the following technical solution: the application of methyl jasmonate in the preparation of drugs for delaying age-related hearing loss.

[0009] Preferably, as an improvement, the drug comprises methyl jasmonate and one or more pharmaceutically acceptable excipients.

[0010] Preferably, as an improvement, the dosage form of the drug is an injection.

[0011] Preferably, as an improvement, the solvent for the injection is physiological saline containing 1% DMSO.

[0012] Preferably, as an improvement, the effective dosage of the drug is 25 mg / kg / day.

[0013] Preferably, as an improvement, the drug is administered continuously for at least 12 weeks.

[0014] Preferably, as an improvement, the drug is used to protect the structural integrity of the cochlea and reduce apoptosis of cochlear hair cells and spiral ganglion cells.

[0015] Preferably, as an improvement, the drug is used to suppress inflammatory responses and oxidative stress damage to the auditory system.

[0016] The principle and advantages of this approach are as follows: In practical applications, the core pathology of age-related hearing loss (presbycusis) in existing technologies is the degenerative death of cochlear hair cells, loss of spiral ganglion neurons, and chronic inflammation / oxidative stress damage to the auditory pathway. Existing technologies can only compensate for symptoms and do not address the root cause with appropriate medications. This technical approach is the first to propose that methyl jasmonate can inhibit the inflammatory response and oxidative stress damage of the auditory system, thus slowing down the degenerative progression of the auditory system from a pathological perspective. For a long time, ARHL has been considered an age-related, irreversible degenerative condition. Clinically, only physical compensation methods such as hearing aids and cochlear implants are available. Globally, there are no approved drugs that can slow down or reverse the progression of ARHL. The industry generally holds a pessimistic view on finding "small molecule drugs that can intervene in ARHL," believing that the aging of the auditory system is "hardware damage" that is difficult to repair with chemical drugs.

[0017] Methyl jasmonic acid, a natural cyclopentanone derivative, is an important stress hormone in plants and a core messenger in plant responses to external environmental stresses (such as drought, ultraviolet radiation, and pests). Traditionally, it is considered to primarily participate in plant defense responses, regulating various physiological processes including growth, development, defense mechanisms, and apoptosis. Currently, there is no literature or technological insight regarding the effects of methyl jasmonic acid on the auditory system, particularly its intervention effects and mechanisms of action in age-related hearing loss (ARHL). Its application in ARHL intervention is a completely new research area. Existing technologies widely suggest that such plant secondary metabolites are rapidly metabolized and inactivated in mammals, or cannot penetrate the blood-labyrinthine barrier to reach inner ear target tissues, making it difficult to produce meaningful pharmacological effects. Furthermore, as a non-endogenous natural product mainly extracted from plants (such as jasmine, tomato, and grape), methyl jasmonic acid faces inherent safety biases in human use, with concerns about potential immunogenicity, allergic reactions, or unknown chronic toxicity, often leading to its exclusion in drug development. This invention, through systematic animal experiments, is the first to clearly discover and demonstrate that methyl jasmonate can significantly improve auditory function in aged mice. Specifically, it enhances pure-tone hearing thresholds and speech recognition abilities in aged mice, while effectively protecting the structural integrity of the cochlea, reducing apoptosis of cochlear hair cells and spiral ganglion cells, inhibiting inflammatory responses and oxidative stress damage in the auditory system, and pathologically slowing the degenerative progression of the auditory system. This discovery fills a gap in the research of methyl jasmonate in the field of hearing, provides a novel candidate molecule for drug intervention in ARHL, and offers important experimental evidence and research directions for subsequent targeted drug development for ARHL, breaking through existing treatment bottlenecks. It successfully bridges the research gap from plant physiology to mammalian audiology.

[0018] In the initial stages of the technical solution research, peripheral blood samples from 260 community individuals underwent non-targeted metabolomics testing, identifying a total of 1883 metabolites, of which as many as 501 were differentially related to age-related hearing loss (ARHL). The first technical hurdle was how to screen out truly functionally active candidate molecules with a causal relationship to the disease from these hundreds or thousands of molecules. This technical solution employs a tiered screening strategy: first, metabolic changes solely caused by aging are excluded, retaining only metabolites significantly associated with the disease state itself; second, the ability of each metabolite to predict ARHL individually is evaluated (AUC value is calculated); third, the expression trends of metabolites at different stages of the disease (mild, moderate, and severe) are analyzed; finally, a metabolite correlation network is constructed, and nodes at the network hubs are screened. After this multi-layered screening, MeJA was ultimately identified as the only target molecule that simultaneously satisfies high predictive efficacy, disease stage-dependent changes, network centrality, and known biological activity.

[0019] Furthermore, determining the dosage of MeJA was another technical challenge in this approach. This method successfully verified that mice in the low-dose group experienced a significant reduction in hearing threshold (10-15 dB SPL in the 4-16 kHz frequency band), while the protective effect was significantly weakened in the high-dose group. This result provides crucial experimental evidence for determining the optimal treatment window.

[0020] The beneficial effects of this technical solution are as follows: 1. First Discovery of a Novel Use: This invention reveals for the first time a novel use of methyl jasmonate in delaying age-related hearing loss. No prior art reports have described the use of methyl jasmonate for the treatment or prevention of age-related hearing loss; this discovery provides a completely new candidate molecule for drug intervention in ARHL.

[0021] 2. Clear efficacy and reliable evidence: This invention, through standardized animal experimental models and dual verification using audiology and histology, confirms that methyl jasmonate can significantly improve the auditory function of elderly individuals and protect the cochlear structure. The efficacy is clear and the data is reliable.

[0022] 3. Effective at low doses with high safety: Animal experiments have confirmed that methyl jasmonate exhibits a significant hearing-protective effect at a low dose of 25 mg / kg. This dose is far below the level at which it may cause toxic side effects, indicating a high safety window for its development as a drug or health supplement. During the technology development phase, the inventors' team also tried a dosage of 40 mg / kg. Although it showed a trend of improving hearing function, its effect was far less than that of the 25 mg / kg low-dose group.

[0023] 4. Broad application prospects: There are currently no effective treatments for ARHL. The candidate molecules provided by this invention are expected to fill this gap and have significant clinical translation potential and market value. Attached Figure Description

[0024] Figure 1 This is a diagram showing the results of methyl jasmonate improving auditory function in aged mice in an embodiment of the present invention.

[0025] Figure 2 This is a fluorescent staining result of methyl jasmonate protecting cochlear hair cells and nerve fibers in an embodiment of the present invention.

[0026] Figure 3 This is a quantitative counting diagram of cochlear hair cells in an embodiment of the present invention.

[0027] Figure 4 This is an image showing the HE staining results of maintaining vascular striae with methyl jasmonate in an embodiment of the present invention.

[0028] Figure 5This is a diagram showing the HE staining results of the cochlear axis in an embodiment of the present invention.

[0029] Figure 6 This is a diagram showing the quantitative analysis results of the cochlear axis in an embodiment of the present invention.

[0030] Figure 7 The following are Western Blot results of ACSL4 and PPARα protein expression in the embodiments of the present invention.

[0031] Figure 8 This is a fluorescence staining result of 4-HNE in an embodiment of the present invention. Detailed Implementation

[0032] The following detailed description provides further details on specific embodiments, but the embodiments of the present invention are not limited thereto. Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art; the experimental methods used are all conventional methods; and the materials and reagents used are all commercially available.

[0033] Example 1 The use of methyl jasmonate in the preparation of drugs for delaying age-related hearing loss (senile deafness). The drug uses methyl jasmonate as the active ingredient and is formulated into a clinically acceptable preparation, preferably an injectable form, in combination with one or more pharmaceutically acceptable excipients. The dosage of the drug is the effective dose, preferably 25 mg / kg / day.

[0034] Note: The methyl jasmonate used in this example was purchased commercially. Purchase information: (rac)-Methyljasmonate from MCE, product number: HY-W013507, specification: 1g.

[0035] Example 1: Protective effect of methyl jasmonate on auditory function in aged mice To verify whether methyl jasmonate has a direct intervention effect on ARHL, this study conducted an animal experiment. Experimental method: Thirty 12-month-old male C57BL / 6J mice were randomly divided into three groups: control group (Vehicle), low-dose MeJA group (25 mg / kg), and high-dose MeJA group (40 mg / kg), with 10 mice in each group.

[0036] Mice in each group were intraperitoneally injected daily with the appropriate dose of MeJA or an equal volume of solvent (1% DMSO saline) for 12 consecutive weeks. After the administration was completed, ABR was measured.

[0037] Experimental results are as follows Figure 1As shown, the hearing thresholds of mice in the low-dose MeJA group were significantly lower than those in the control group at frequencies of 4, 8, 12, and 16 kHz (P<0.05), with a reduction of 10-15 dB SPL. Although the high-dose group showed an improvement trend, the effect was not as good as that of the low-dose group. This result clearly shows that appropriate concentrations of methyl jasmonate can indeed effectively delay hearing loss in aged mice, and this protective effect exhibits the dose-dependent characteristics we expected.

[0038] Note: p <0.05 is *. p <0.01 is **. p <0.001 is ***. p <0.0001 indicates ****, ns indicates no statistical significance. A: Before administration. B: 12 ​​weeks after administration.

[0039] Example 2: Protective effect of methyl jasmonate on cochlear hair cells and neurons Experimental Methods: Mice were sacrificed after ABR testing, and cochlear tissue (cochlear basilar membrane) was collected for immunofluorescence staining. The basilar membrane is the core structure of the auditory receptor, running spirally along the cochlear duct, and is densely packed with sensory hair cells and supporting cells, forming the initial interface for auditory signal transduction. We used Myosin7a to specifically label the cytoplasm of hair cells, which showed green fluorescence. The morphology and arrangement of hair cells were clearly shown. Nerve fibers were labeled with NF200, which showed red fluorescence. The nuclei of hair cells were labeled with DAPI, which showed blue fluorescence. This was used to assess the distribution of peripheral processes of auditory neurons and their connections with hair cells. Test results are as follows: Figure 2-3 As shown, Myo7a (hair cell marker) and NF200 (nerve fiber marker) staining revealed that the hair cells in each reposition of the cochlear basilar membrane in the low-dose MeJA group were more regularly arranged, and the number of surviving cells was significantly higher than that in the control group (P<0.05). The density and morphology of nerve fibers were also well maintained.

[0040] Figure 2 Methyl jasmonate was used to protect cochlear hair cells and nerve fibers (A: apical gyrus; B: middle gyrus; C: basal gyrus). Myosin7a (hair cells) was labeled with green fluorescence, NF200 (nerve fibers) with red fluorescence, and DAPI (nuclei) with blue fluorescence. Scale bar: 10 μm.

[0041] Figure 3 In (AB): p <0.05 is *. p <0.01 is **. p <0.001 is ***. p <0.0001 indicates ****, ns indicates no statistical significance. A: External hair cell count. B: Internal hair cell count.

[0042] The above experimental results show that the hair cell survival rate in the low-dose group was significantly higher than that in the apical, middle, and basal gyri than in the high-dose group, while the high-dose group was significantly higher than that in the control group. This indicates that methyl jasmonic acid can effectively delay age-related hair cell degeneration within a certain dosage range, with the low-dose regimen showing the most ideal cochlear protection effect. This result suggests that methyl jasmonic acid may play a protective role for the auditory organs during aging by regulating inner ear oxidative stress or anti-apoptotic pathways.

[0043] Experimental Example: Effect of Methyl Trijasmonate on the Strauss Vascularis of the Lateral Wall of the Cochlea Besides hair cells and neurons, the stria vascularis on the lateral wall of the cochlea is also a key structure for maintaining normal auditory function. The stria vascularis provides the driving force for the mechatronic conversion of hair cells by maintaining a high-potassium, low-sodium state in the cochlear endolymph. Immunofluorescence staining of the stria vascularis in this experimental case yielded the following results: Figure 4 As shown (scale bar: 20 μm): Peripheral cells of the stria vascularis are displayed using Phalloidin labeling; blue indicates DAPI-labeled cell nuclei. Typical age-related changes were observed in the stria vascularis of control mice: fusion of peripheral cells. Methyl jasmonate treatment effectively improved these structural abnormalities—the cellular layers of the stria vascularis became clearer, and the morphology of peripheral cells became healthier.

[0044] This experiment also used HE staining to observe the overall structure of the cochlea, focusing on the morphological parameters of the spiral ganglion cells and stria vascularis. Spiral ganglion neurons are first-order neurons in the auditory pathway; their cell bodies are clustered within the cochlear axon at the center of the cochlea, receiving signals from hair cells and transmitting them to the central nervous system. Results are as follows... Figure 5 As shown, the spiral ganglion cell density in the control group mice was significantly reduced, and the stria vascularis thickness was also significantly thinner. In contrast, the spiral ganglion cell density in the methyl jasmonate treatment group was higher, the cell morphology was healthier, and the stria vascularis thickness was significantly maintained.

[0045] Figure 6 The quantitative statistical bar charts clearly illustrate these differences.

[0046] In summary, methyl jasmonate maintains the integrity of the auditory pathway at the structural level by simultaneously protecting three key cellular structures: hair cells, spiral neurons, and stria vascularis, thus providing a solid morphological basis for improving hearing function.

[0047] Experimental Study on the Core Molecular Mechanism of Tetrajasmonate in Delaying Hearing Decline in Aged Mice Having established the protective effect of methyl jasmonate, this study further explored the mechanism by which it exerts this protective effect. This research focused on the ACSL4 protein, aiming to elucidate the molecular mechanism by which methyl jasmonate inhibits cochlear lipid peroxidation through the PPAR-ACSL4 axis. The protein level of ACSL4 in cochlear tissue was detected using Western blotting, and the results are as follows: Figure 7 As shown, consistent with the trend at the mRNA level, ACSL4 protein expression was significantly reduced after methyl jasmonate treatment, and this reduction also exhibited a dose-dependent effect: the reduction was more pronounced in the low-dose group, followed by the high-dose group. PPARα also showed a dose-dependent effect, with the PPAR pathway being significantly activated in the low-dose group.

[0048] Immunofluorescence staining was performed using 4-HNE antibody. 4-HNE is one of the end products of lipid peroxidation, and its content can directly reflect the activity of lipid peroxidation in tissues. Simultaneously, we also performed co-localization staining on ACSL4 itself. Immunofluorescence co-localization staining images show the distribution and intensity of ACSL and the lipid peroxidation end product 4-HNE in the cochlear tissues of mice in each group. Results are as follows: Figure 8 As shown, in the cochlea of ​​control mice, 4-HNE exhibited very strong fluorescence signals, appearing as bright red, in both the hair cell region and the spiral neuron region. Simultaneously, ACSL4 also showed a strong green signal, with their distributions highly overlapping, suggesting that these regions are undergoing active lipid peroxidation. However, in the methyl jasmonate treatment group, especially the low-dose group, the fluorescence intensity of both 4-HNE and ACSL4 was significantly reduced, and the previously bright red and green colors became duller, indicating a significant decrease in the degree of lipid peroxidation.

[0049] Figure 8 In the image, ACSL4 is labeled with green fluorescence, 4-HNE with red fluorescence, and DAPI is used to label the cell nucleus with blue fluorescence. Scale bar: 10 μm.

[0050] The above descriptions are merely embodiments of the present invention, and common knowledge such as specific technical solutions and / or characteristics are not described in detail here. It should be noted that those skilled in the art can make various modifications and improvements without departing from the technical solutions of the present invention, and these should also be considered within the scope of protection of the present invention. These modifications and improvements will not affect the effectiveness of the implementation of the present invention or the practicality of the patent. The scope of protection claimed in this application should be determined by the content of its claims, and the specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. Application of methyl jasmonate in the preparation of drugs to delay age-related hearing loss.

2. The application of methyl jasmonate according to claim 1 in the preparation of drugs for delaying age-related hearing loss, characterized in that: The drug comprises methyl jasmonate and one or more pharmaceutically acceptable excipients.

3. The application of methyl jasmonate according to claim 2 in the preparation of drugs for delaying age-related hearing loss, characterized in that: The drug is in the form of an injection.

4. The use of methyl jasmonate according to claim 3 in the preparation of drugs for delaying age-related hearing loss, characterized in that: The solvent for the injection is physiological saline containing 1% DMSO.

5. The use of methyl jasmonate according to claim 4 in the preparation of a drug for delaying age-related hearing loss, characterized in that: The effective dosage of the drug is 25 mg / kg / day.

6. The use of methyl jasmonate according to claim 5 in the preparation of drugs for delaying age-related hearing loss, characterized in that: The drug is administered continuously for at least 12 weeks.

7. The use of methyl jasmonate according to claim 6 in the preparation of drugs for delaying age-related hearing loss, characterized in that: The drug is used to protect the structural integrity of the cochlea and reduce apoptosis of cochlear hair cells and spiral ganglion cells.

8. The use of methyl jasmonate according to claim 7 in the preparation of a drug for delaying age-related hearing loss, characterized in that: The drug is used to suppress inflammatory responses and oxidative stress damage to the auditory system.

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