Application of expression inhibitor of Gadd45a in preparation of medicine for preventing or reducing cisplatin ototoxicity

Abstract

The invention provides an application of an expression inhibitor of Gadd45a in preparation of a medicine for preventing or reducing the ototoxicity of cis-platinum. In-vivo and in-vitro experiments prove that by locally applying the Gadd45a inhibitor, cochlea hair cells and auditory functions can be effectively protected by activating autophagy, inhibiting an NF-kappaB1 inflammation pathway and regulating JNK-mediated apoptosis, so that hearing loss caused by cis-platinum is relieved. The invention provides a brand-new effective medicine product and a treatment strategy for clinically preventing and treating the ototoxicity of cis-platinum, and has important clinical application value for preventing or relieving the ototoxicity of a patient receiving cis-platinum chemotherapy.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of Gadd45a expression inhibitors in the preparation of drugs for preventing or mitigating cisplatin ototoxicity. Background Technology

[0002] Cisplatin is a highly effective and broad-spectrum platinum-based anti-tumor drug that has been widely used in clinical practice since its introduction. It has demonstrated remarkable therapeutic effects against various solid tumors, such as testicular cancer, ovarian cancer, bladder cancer, and head and neck tumors. Cisplatin's main anti-cancer mechanism involves entering cells and forming activated hydrated molecules that cross-link with DNA strands, disrupting DNA replication and transcription, thereby inducing tumor cell apoptosis. However, the clinical application of cisplatin is significantly limited by its severe toxic side effects, particularly nephrotoxicity and ototoxicity. Cisplatin ototoxicity often manifests as bilateral, progressive, and irreversible sensorineural hearing loss, accompanied by tinnitus, severely impacting the quality of life of patients, especially long-term surviving children with cancer. Its pathological mechanism is mainly related to the accumulation of cisplatin in the cochlea, particularly in the outer hair cells, stria vascularis, and spiral ganglion, triggering oxidative stress, inflammatory responses, and various programmed cell deaths (including apoptosis and ferroptosis), ultimately leading to irreversible damage to auditory cells. Because the outer hair cells of the basal gyrus of the cochlea are more sensitive to cisplatin, hearing loss typically begins at high frequencies and gradually extends to the mid and low frequencies with increasing dosage. Currently, there are no effective drugs approved by the US FDA or China NMPA for the prevention and treatment of cisplatin ototoxicity. Some studies have explored the use of antioxidants (such as amifostine) or other cytoprotective agents, but these strategies often have one or more inherent drawbacks: (1) Limited protective effect: The protective effect of many drugs has failed to achieve the expected effect in clinical studies, or the results are inconsistent.

[0003] (2) Risk of potential interference with anticancer efficacy: While systemically administered protective agents protect normal tissues, there is a concern that they may weaken the killing effect of cisplatin on tumor cells, which is unacceptable to clinicians and patients.

[0004] (3) Inconvenient or invasive administration routes: Some exploratory protocols require complex administration methods, which limits their clinical translation and application prospects.

[0005] Therefore, there is an urgent need in this field to develop a new drug that can effectively and specifically prevent or reduce the ototoxicity of cisplatin, while ensuring that its core anti-tumor efficacy is not affected. Summary of the Invention

[0006] To address the aforementioned technical problems, this invention provides the application of Gadd45a expression inhibitors in the preparation of drugs for preventing or mitigating cisplatin ototoxicity. Gadd45a expression inhibitors have significant application prospects in the preparation of drugs for preventing or mitigating cisplatin ototoxicity.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect of the invention, the use of Gadd45a as a target in screening for drugs to prevent or mitigate cisplatin ototoxicity is provided, wherein the screening method includes screening for substances that can inhibit the expression of the Gadd45a gene.

[0008] In a second aspect of the invention, the use of an inhibitor of Gadd45a expression in the preparation of a medicament for preventing or mitigating cisplatin ototoxicity is provided.

[0009] Furthermore, the Gadd45a expression inhibitor comprises at least one of the following components: Gadd45a inhibitors; Knockdown reagent for Gadd45a.

[0010] Furthermore, the Gadd45a knockdown reagent comprises: shRNA and / or gRNA and / or siRNA targeting the target gene, wherein the siRNA nucleotide sequence is shown in SEQ ID NO:1 (5'-GGAUCCUGCCUUAAGUCAAtt-3').

[0011] The gRNA nucleotide sequence is shown in SEQ ID NO:2-5. In a third aspect of the invention, a pharmaceutical composition for preventing or mitigating cisplatin ototoxicity is provided, the pharmaceutical composition comprising at least one of a Gadd45a inhibitor and a Gadd45a knockdown agent.

[0012] The pharmaceutical composition comprises a pharmaceutically acceptable carrier. The choice of the carrier depends on the type of Gadd45a inhibitor and the route of administration. When the inhibitor is a nucleic acid molecule (such as siRNA), the carrier preferably comprises a delivery carrier, such as lipid nanoparticles (such as Invivofectamine® 3.0 reagent), polymer nanoparticles, cationic liposomes, or viral vectors (such as adeno-associated virus vectors), to facilitate its entry into cells. More broadly, for pharmaceutical compositions suitable for topical inner ear administration, the carrier may comprise excipients conventional in the art, such as: Solvents or diluents: such as water for injection, physiological saline, phosphate buffer; Isotonic agents: such as glucose, mannitol, glycerin; pH adjusters and buffers: such as hydrochloric acid, sodium hydroxide, phosphate buffer, Tris-HCl buffer; Preservatives: such as benzalkonium chloride; Thickeners or gel matrices: such as hyaluronic acid, methylcellulose, poloxamer 407 (used to prepare thermosensitive gels); Sustained-release materials: such as polylactic acid-glycolic acid copolymer.

[0013] The dosage form of the pharmaceutical composition can be prepared in various forms suitable for local inner ear administration, including but not limited to: injection solutions, in-situ gels for injection, sustained-release microspheres, implantable films, or sponges. Preferred routes of administration include intratympanic injection, posterior semicircular canal injection, or round window membrane puncture injection.

[0014] The dosage form of the drug includes one of the following: injection, gel, and sustained-release formulation.

[0015] The beneficial effects of the above-described technical solution of the present invention are as follows: This invention provides the application of a Gadd45a expression inhibitor in the preparation of drugs for preventing or mitigating cisplatin ototoxicity. In vitro and in vivo experiments demonstrate that targeted inhibition of Gadd45a produces significant and multifaceted technical effects, effectively overcoming the problem of the lack of effective means to prevent cisplatin ototoxicity in existing technologies. Specifically, the effects are reflected in the following four aspects: 1. Significant cell protection effect: Knocking out the Gadd45a gene in the cochlear hair cell line (OC1) can significantly enhance the cell's tolerance to cisplatin.

[0016] Cell viability: CCK-8 assays showed that, under different concentrations of cisplatin treatment, the cell viability of the Gadd45a knockout group was significantly higher than that of the control group. Figure 1 B).

[0017] Inhibition of apoptosis: Flow cytometry (Annexin V / PI staining) and TUNEL staining results jointly confirmed that Gadd45a knockout significantly reduced the proportion of cisplatin-induced early apoptosis from approximately 35% to approximately 10%. Figure 1 C-1F clearly demonstrates its powerful anti-apoptotic effect.

[0018] 2. Clear in vivo hearing protection effect in animals: In vivo experiments demonstrated its therapeutic potential by specifically knocking down Gadd45a expression in the inner ear of mice through injection of siRNA into the posterior semicircular canal.

[0019] Hearing function preservation: Auditory brainstem response (ABR) tests showed that, compared with the control group, mice treated with si-Gadd45a experienced significantly smaller increases in hearing thresholds at different frequencies (8, 16, 32 kHz) after cisplatin chemotherapy. Figure 3 H) indicates that hearing function has been effectively preserved.

[0020] Cochlear structure protection: Immunofluorescence staining of the cochlear basilar membrane showed that the loss of cochlear hair cells was significantly reduced in the si-Gadd45a treatment group. Figure 3 JK) has confirmed its protective effect on the cochlear structure from a histological perspective.

[0021] 3. Advantages of multi-pathway synergistic effect: This invention reveals for the first time the multiple mechanisms by which Gadd45a regulates cisplatin ototoxicity, rather than relying on a single pathway, which ensures the robustness and efficiency of the protective effect.

[0022] Activation of protective autophagy: Gadd45a knockout significantly enhanced cellular autophagy levels. Figure 2 A-2C helps remove damaging substances caused by cisplatin.

[0023] Suppressing harmful inflammation: Gadd45a knockout leads to decreased expression of the key inflammatory transcription factor NF-κB1 and its downstream inflammatory factors. Figure 2 D-2E) reduced cisplatin-induced inflammatory damage.

[0024] Blocking apoptosis signaling: Experiments have shown that Gadd45a overexpression activates the JNK apoptosis pathway, while inhibiting Gadd45a can block this pathway. Figure 2 These three pathways work synergistically to form a robust cell protection network.

[0025] 4. High targeting specificity and safety: The local inner ear administration method (such as posterior semicircular canal injection) adopted in this invention precisely limits drug delivery to the target organ (cochlea), minimizing the side effects that may be caused by systemic administration and fundamentally eliminating the risk of interfering with the systemic anticancer efficacy of cisplatin, thus exhibiting extremely high clinical translational safety. Attached Figure Description

[0026] To more clearly illustrate the technical solutions in the embodiments of this application, the following detailed description of non-limiting embodiments is provided using accompanying drawings, making their features, objectives, and advantages more apparent: Figure 1: Validation of the protective effect of Gadd45a knockout against cisplatin toxicity at the cellular level. (A) qRT-PCR detection of Gadd45a gene knockout efficiency. (B) CCK-8 assay detection of cell viability in the control group and Gadd45a knockout group under different concentrations of cisplatin treatment. (CD) Flow cytometry detection of cisplatin-induced apoptosis. (EF) TUNEL staining detection of DNA fragmentation.

[0027] Figure 2 : Mechanism of Gadd45a regulation of cisplatin ototoxicity. (A) Changes in the expression levels of autophagy-related proteins. (BC) Immunofluorescence showing an increase in autophagosomes in Gadd45a knockout cells. (DE) Detection of NF-κB1 and downstream inflammatory cytokines expression levels. (FG) Gadd45a overexpression activates the JNK pathway and induces cytotoxicity.

[0028] Figure 3 : In vivo animal studies to verify the effect of siRNA knockdown of Gadd45a. (A) Schematic diagram of injection into the posterior semicircular canal and fluorescence image of successful siRNA delivery. (B) Detection of siRNA residence time in the cochlea. (C) Anatomical structure of cochlear hair cells after injection. (DF) Western blotting and immunofluorescence verification of downregulated Gadd45a protein expression in cochlear tissue. (GK) Mouse hearing test (ABR) and cochlear basilar membrane staining results.

[0029] Figure 4 The present invention relates to a signaling pathway pattern of Gadd45a regulating cisplatin ototoxicity. Detailed Implementation

[0030] The present invention will now be described in detail with reference to the accompanying drawings. The present invention and its embodiments are described below; however, this description is not restrictive, and actual embodiments are not limited thereto. If those skilled in the art, inspired by this description, design similar structures and embodiments without departing from the spirit of the invention, such designs should fall within the protection scope of the present invention.

[0031] Example 1: Cell-level target validation 1.1 Construction and validation of Gadd45a gene knockout cell lines We first knocked out the Gadd45a gene in the cochlear hair cell line OC1 using CRISPR-Cas9 technology.

[0032] Construction of OC1 gene knockout cell lines (1) Purchase the PX459 plasmid vector designed by Zhang Feng’s laboratory from Addgene, and obtain sticky ends after digestion with Bbs1 enzyme for later use.

[0033] (2) Use the MIT online design tool http: / / crispr.mit / edu / to design CRISPR / CAS combinatorial sequences.

[0034] (3) The gene sequences used in this paper are shown below: Table 1

[0035] (4) The two single strands synthesized above are annealed to form a double-stranded DNA containing Bbs1 sticky ends, which is then ligated with the PX459 (Bbs1 digestion) vector and transformed to obtain the target plasmid vector. After sequencing, the plasmid is extracted, and after removing endotoxin, it is transfected into cells.

[0036] (5) OC1 cells were seeded on 60 mm plates and transfected with plasmids when the cells reached about 60% of their length.

[0037] (6) In accordance with the instructions for use of the transfection reagent, the plasmid vector and transfection reagent were coupled at a ratio of 1 mg: 2 μl. After standing for 16 min in a clean bench, the mixture was added to a cell culture dish.

[0038] (7) The next day, the supernatant was discarded and puromycin at a concentration of 3.5 μg / ml was added for pressure screening.

[0039] (8) After 72 hours of treatment with puromycin, most of the cells had died and did not develop puromycin resistance because they had not been successfully transfected.

[0040] (9) Remove the cell culture supernatant, wash twice with sterile PBS, add 1 ml of trypsin for digestion, and centrifuge to count the cells.

[0041] (10) After dilution, reseed the cells in a 96-well plate, ensuring that there is only one cell in each well.

[0042] (11) Place the 96-well plate in a cell culture incubator for 2 weeks, select cell clusters with single clones, digest and seed them on a 60 mm plate, and collect samples to extract proteins after they have grown to 80%.

[0043] Efficiency was determined by qRT-PCR and Western blotting, and the results showed that the gene knockout efficiency of Gadd45a was approximately 80%. Figure 1 A).

[0044] 1.2 Cell viability assay (CCK-8 assay) Cell viability was assessed using a CCK-8 assay kit (Vazyme, A311-02-AA).

[0045] On the first day, OC1 and Gadd45a gene knockout cells were stored at a rate of 5 × 10⁶ cells per well. 3Cells were seeded at a density of [number] cells per well in 96-well plates and cultured overnight to allow cell adhesion. The next day, the medium was replaced with fresh medium containing different concentrations of cisplatin and treated for 24 hours (TOCRIS, #2251). After establishing the cisplatin-induced injury model, 10 μL of CCK-8 reaction solution was mixed with 90 μL of fetal bovine serum-free DMEM medium and added to each well. The 96-well plates were then incubated at 37°C for 1–2 hours. Absorbance was measured at 450 nm using a multi-mode microplate reader (HH3500; PerkinElmer, USA).

[0046] The results are as follows Figure 1 As shown in B, the cell viability of both the control group and the Gadd45a gene knockout group decreased with increasing cisplatin concentration, but the cell viability of the Gadd45a gene knockout group was significantly higher than that of the control group.

[0047] 1.3 Apoptosis detection (flow cytometry) The apoptosis rate of cells exposed to cisplatin was assessed using the Annexin V-FITC / PI kit (BD Biosciences Pharmingen, 556547). On day 1, OC1 and Gadd45a knockout cells were seeded at a density of 60% in 6-well plates. On day 2, the medium was replaced with fresh medium containing 25 μM cisplatin and incubated at 37°C for 24 h. Cells were then washed twice with pre-chilled PBS, digested with trypsin, collected by centrifugation at 1000 rpm for 5 min, and resuspended at 1x10⁶ cells / well. 6 A concentration of cells / mL was determined. Annexin V-FITC and PI were added to the cell suspension and incubated at room temperature in the dark for 15 minutes. Data were acquired using a Beckman flow cytometer and analyzed using FlowJo software.

[0048] The results are as follows Figure 1 As shown in CD, compared with the control group, Gadd45a gene knockout significantly reduced cisplatin-induced early apoptosis, from about 35% to about 10%.

[0049] 1.4 Apoptosis detection (TUNEL staining) TUNEL assays were performed according to the protocol of the in situ cell death assay kit (Roche, Indianapolis, IN, USA). On day 1, OC1 and Gadd45a gene knockout cells were seeded in confocal culture dishes and treated with cisplatin. After modeling, cells were washed with pre-chilled PBS and fixed with formaldehyde at room temperature for 15 min. Cell permeation was then performed by treatment with 0.2% Triton X-100 for 15 min. Subsequently, 100 μL of TUNEL reaction mixture was added to each confocal culture dish and incubated at 37°C for 60 min. After TUNEL staining, DAPI was added at a concentration of 1:500 and incubated at room temperature for 10 min. Finally, images were acquired using a Leica confocal laser scanning microscope (Leica TCS SP8, Leica Microsystems GmbH, Wetzlar, Germany).

[0050] The results are as follows Figure 1 As shown in EF, TUNEL staining confirmed a decrease in cisplatin-induced apoptosis in the Gadd45a gene knockout group, manifested as a reduction in the number of TUNEL-positive cells.

[0051] Example 2: Study on the mechanism of action 2.1 Validation of Autophagy Pathway Activation Autophagy was significantly activated after Gadd45a gene knockout. Western blot results ( Figure 2 A) This provides evidence for further verification of the activation of the autophagy signaling pathway in Gadd45a gene knockout cells.

[0052] Furthermore, immunofluorescence staining showed that autophagosomes were significantly increased in Gadd45a knockout cells after cisplatin treatment and double-labeled lentiviral transfection. Figure 2 BC).

[0053] 2.2 Validation of Inflammatory Pathway Inhibition Gadd45a gene knockout leads to a reduction in the transcription factor NF-κB1, which in turn results in a decrease in the level of CXCL family proteins, providing strong evidence for Gadd45a's regulation of inflammatory signaling pathways. Figure 2 DE).

[0054] 2.3 Validation of the JNK apoptosis pathway Overexpression of Gadd45a activates the JNK signaling pathway, which is the primary mediator of apoptosis. This suggests that Gadd45a knockout may protect cells from cisplatin-induced apoptosis by inhibiting the JNK signaling pathway.

[0055] To confirm this hypothesis, we performed immunofluorescence staining. In these experiments, DAPI was used to label all cell nuclei, while FLAG staining identified cells that successfully overexpressed Gadd45a.

[0056] The results showed that cells exhibiting green fluorescence displayed cytoplasmic fragmentation, strongly indicating the cytotoxicity of Gadd45a overexpression. Figure 2 FG).

[0057] Example 3: In vivo verification and application in animals 3.1 Animal Models and siRNA Inner Ear Delivery We used an intraocular injection method to inject siRNA (sequence GGAUCCUGCCUUAAGUCAAtt) into the inner ear of mice through the posterior semicircular canal to specifically reduce the expression of Gadd45a.

[0058] The image illustrates the surgical procedure of inserting a microcannula into the posterior semicircular canal, with the black arrow indicating the injection site within the semicircular canal. Figure 3 A). The specific surgical procedure was as follows: The animal was deeply anesthetized by intraperitoneal injection of ketamine (100 mg / kg, ip) and xylazine (25 mg / kg, ip). The skin behind the ear was prepared by clipping the hair and applying depilatory cream. Subsequently, the mouse was placed on an operating table (China Yuyan Instruments) and its body temperature was maintained using an electric heating pad. A 1 cm incision was made behind the ear, and the sternocleidomastoid muscle was bluntly dissected using micro-scissors. The facial nerve and tympanic cavity were identified, and the posterior semicircular canal (PSCC) was located above and behind the facial nerve. The muscle fibers and soft tissue covering the PSCC were removed, and a 27-gauge needle was used to drill into the PSCC lumen. Lymphatic leakage indicated successful entry into the PSCC lumen. Injection was performed using a microinjection system via a microsyringe connected to a thin tube. The tube was inserted into the PSCC with forceps. The gap between the inserted tube and the PSCC was sealed with superficial fascia and tissue adhesive to prevent lymphatic leakage. siRNA liposome solution was injected into the PSCC at a rate of 95 μL / h. After injection, the PSCC window was closed with adjacent muscle, and the incision was sutured with a crimping needle. The wound was disinfected with povidone-iodine, and then the mouse was placed on a heating pad to recover.

[0059] 3.2 siRNA delivery efficiency and residency verification To assess the stability and retention time of siRNA in the inner ear, 5'-FAM-siRNA (excitation wavelength 494 nm, emission wavelength 520 nm) was mixed with Invivofectamine 3.0 reagent according to the complex preparation protocol and then injected into the inner ear via PSCC. Retention was evaluated using an in vivo imaging system (IVIS SPECTRUM, PerkinElmer).

[0060] After injecting green fluorescently labeled 5'-FAM-siRNA into the semicircular canals of the cochlea, fluorescence microscopy images showed that the semicircular canals were filled with green fluorescence, confirming successful delivery. Figure 3 A). The figure also shows the residence time of 5'-FAM-siRNA in the cochlea after injection. Figure 3 B), while the anatomical structure of cochlear hair cells remained intact after 5'-FAM-siRNA injection, indicating that siRNA can effectively penetrate cochlear hair cells (B). Figure 3 C).

[0061] 3.3 In vivo knockdown efficiency verification To further improve the efficiency of Gadd45a suppression in the cochlea, we used Invivofectamine 3.0 Reagent.

[0062] Western blot (protein immunoblotting) Figure 3 DE and immunofluorescence staining confirmed that siRNA treatment significantly reduced the expression of Gadd45a in the cochlea. Figure 3 F).

[0063] 3.4 Evaluation of hearing protection effectiveness (ABR test) The ABR test was used to measure hearing thresholds. Specifically, to ensure normal hearing ability in mice before each treatment, we first measured the mice's weight and then, based on weight, anesthetized them by intraperitoneal injection of ketamine (100 mg / kg) and xylazine (25 mg / kg). After satisfactory anesthesia, recording electrodes were inserted subcutaneously: behind the test ear (black), behind the contralateral ear (green), and along the midline of the head (red). Subsequently, the hearing thresholds of the mice were tested at frequencies of 8, 16, and 32 kHz using a Tucker-Davis Technologies (TDT) RZ6 system (Alecithia, Florida, USA). For each frequency, the sound intensity was initially set to 90 dB and then gradually decreased by 10 dB until no ABR wave response was detected, thus determining the threshold for each frequency. To reduce variability, all tests were performed by the same person. To evaluate the efficacy of siRNA-mediated Gadd45a knockdown in alleviating cisplatin ototoxicity, we first performed hearing tests on the mice to establish a baseline for subsequent experiments. Figure 3 G).

[0064] Mice with acceptable baseline audiograms were selected and injected with si-Gadd45a via the posterior semicircular canal. The following day, the mice were injected intraperitoneally with cisplatin. Hearing tests were then performed. Figure 3 Our results indicate that siRNA-mediated downregulation of Gadd45a significantly alleviated cisplatin-induced ototoxicity (H). Figure 3H).

[0065] 3.5 Histological evaluation of the cochlea After the ABR test, the mice were euthanized, and the cochlea was removed for immunofluorescence staining of the basement membrane. Figure 3 JK). The results also showed that si-Gadd45a treatment significantly reduced cisplatin-induced cochlear hair cell loss.

[0066] The above examples clearly demonstrate that targeted inhibition of Gadd45a (whether through gene knockout or siRNA knockdown) can effectively reduce cisplatin ototoxicity at the cellular and animal levels, and its mechanism of action is related to the regulation of autophagy, inflammation, and JNK apoptosis pathways. Local delivery of Gadd45a siRNA is a feasible therapeutic strategy.

[0067] The above description represents the preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. The application of Gadd45a as a target in screening drugs to prevent or mitigate cisplatin ototoxicity, characterized in that... The screening method includes screening for substances that can inhibit the expression of the Gadd45a gene or the activity of the protein.

2. The application of Gadd45a expression inhibitors in the preparation of drugs for the prevention or mitigation of cisplatin ototoxicity.

3. The application according to claim 2, characterized in that, The Gadd45a expression inhibitor includes at least one of the following components: Gadd45a inhibitors; Knockdown reagent for Gadd45a.

4. The application according to claim 3, characterized in that, The Gadd45a knockdown reagent comprises: shRNA and / or gRNA and / or siRNA targeting the target gene, wherein the nucleotide sequence of the siRNA is shown in SEQ ID NO:

1.

5. A drug for preventing or mitigating cisplatin ototoxicity, characterized in that, The drug includes at least one of a Gadd45a inhibitor and a Gadd45a knockdown agent.

6. A medicament for preventing or mitigating cisplatin ototoxicity according to claim 5, characterized in that, The drug can inhibit the expression of the Gadd45a gene or the activity of its protein.

7. A pharmaceutical composition for preventing or mitigating cisplatin ototoxicity according to claim 5, characterized in that, The drug also includes a pharmaceutically acceptable carrier.

8. A medicament for preventing or mitigating cisplatin ototoxicity according to claim 7, characterized in that, The pharmaceutically acceptable carriers include one or more of the following: diluents, isotonic agents, pH adjusters, preservatives, thickeners, or sustained-release materials.

9. A medicament for preventing or mitigating cisplatin ototoxicity according to claim 7, characterized in that, The pharmaceutically acceptable carrier comprises a delivery carrier for delivering the nucleic acid inhibitor, wherein the delivery carrier is one of lipid nanoparticles, polymer nanoparticles, cationic liposomes, or viral vectors.

10. A medicament for preventing or mitigating cisplatin ototoxicity according to claim 5, characterized in that, The dosage form of the drug includes one of the following: injection, gel, and sustained-release formulation.

Images