Use of a small molecule compound targeting phosphorylated ARMC10 S43 in the preparation of a medicament for treating neurological damage caused by tin exposure

Abstract

This invention relates to the field of pharmaceutical technology for treating diseases related to heavy metal exposure, specifically to the application of a small molecule compound targeting the phosphorylated serine 43 site of ARMC10 in the preparation of drugs for treating neurological damage caused by tin exposure. This technical solution is the first to discover that trimethyltin chloride exposure can specifically induce abnormal phosphorylation of ARMC10, thereby causing neuronal death and cognitive impairment. Based on this mechanism, a small molecule compound targeting a specific phosphorylation site of ARMC10 is proposed, which effectively reverses neurological damage and protects neuronal structure and function by blocking its mediated abnormal mitochondrial division and dysfunction. This solution solves the problem of existing technologies lacking effective treatments for tin exposure-related neurological damage due to the lack of clear drug targets, and has ideal application prospects. This small molecule compound has a well-defined structure and a clear mechanism of action, is easy to further develop, and possesses good drug development and commercialization prospects.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology for treating diseases related to heavy metal exposure, specifically to the application of a small molecule compound that targets phosphorylated ARMC10 S43 in the preparation of a drug for treating neurological damage caused by tin exposure. Background Technology

[0002] Against the backdrop of escalating environmental health risks, the dual threat posed by toxic pollutants to both ecology and human health has become a focus of international concern. Trimethyltin chloride (TMT), a typical organotin compound, was once widely used. Although its use has been somewhat restricted, it cannot be completely avoided in industrial processing such as PVC plastics manufacturing, and poisoning cases still occur annually in some areas. TMT continues to cause pollution through various pathways, including wastewater discharge and processed products. It is frequently detected not only in drinking water, food, and daily chemical products, but can even be found in human tissues, thus posing an exposure risk.

[0003] Numerous studies have demonstrated that TMT (tumor thrombosis) possesses significant neurotoxicity, particularly targeting the hippocampus to induce cognitive impairment. Its toxic effects are closely related to mitochondrial damage: TMT exposure can induce excessive reactive oxygen species (ROS) production, disrupt intracellular calcium homeostasis, and consequently damage mitochondrial structural integrity and respiratory chain function. Recent research has further revealed that mitochondrial dynamic imbalance (dysregulation of fusion and division processes) is the core pathological mechanism of TMT-induced neuronal dysfunction. For example, TMT can lead to mitochondrial swelling, vacuolation, and a reduction in the number of mitochondria in hippocampal neurons. Interventions such as artemisinin, by restoring mitochondrial morphology and content, effectively reduce neuronal apoptosis and improve behavioral deficits, highlighting the potential of targeting mitochondrial dynamic regulation in neuroprotection.

[0004] Against this backdrop, ARMC10 (containing armadillo repeat protein 10), a newly identified mitochondrial outer membrane protein, has gradually been identified as a key factor in regulating mitochondrial dynamics. ARMC10 can dynamically regulate its activity through post-translational modifications (such as phosphorylation), thereby affecting mitochondrial morphology, energy metabolism, and neuronal survival. However, a crucial gap remains: it is currently unclear whether ARMC10 is involved in TMT-induced mitochondrial dysfunction and its downstream neurotoxic processes, nor is there evidence that it can serve as a molecular target for antagonizing organotin neurotoxicity.

[0005] Specifically, whether TMT exposure affects the expression, localization, or modification status of ARMC10, and whether changes in ARMC10 function are sufficient to determine neuronal sensitivity to TMT toxicity, remain unanswered. Consequently, the lack of mechanistic analysis and intervention strategies based on the ARMC10 pathway has become a major technical bottleneck in the development of targeted neuroprotective therapies. Summary of the Invention

[0006] The present invention aims to provide the application of a small molecule compound that targets phosphorylated ARMC10 S43 in the preparation of a drug for treating tin exposure-induced neurological damage, in order to solve the technical problem that there are no effective drugs for treating tin exposure-related neurological damage due to the lack of a clear drug target.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] The application of small molecule compounds in the preparation of drugs for treating tin exposure-induced neurological damage, wherein the structural formula of the small molecule compounds is shown as Formula II or Formula V:

[0009]

[0010] Formula II;

[0011]

[0012] Formula V.

[0013] Furthermore, small molecule compounds with structures such as Formula II or Formula V are used to target the ARMC10 protein, which is phosphorylated at serine position 43.

[0014] Furthermore, the affinity of the small molecule compound with structure II to the ARMC10 protein phosphorylated at serine position 43 is -9.783 kcal / mol; the affinity of the small molecule compound with structure V to the ARMC10 protein phosphorylated at serine position 43 is -9.291 kcal / mol.

[0015] Furthermore, tin exposure-induced neurological damage refers to acute / subacute neurological damage caused by organotin exposure.

[0016] Furthermore, the organotin is trimethyltin chloride.

[0017] Furthermore, the neurological damage manifests as reduced neuronal activity, impaired neuronal mitochondrial function, and abnormal neuronal mitochondrial morphology.

[0018] Furthermore, the structural formula of the small molecule compound is shown in Formula II:

[0019]

[0020] Formula II.

[0021] This technical solution also provides a drug that alleviates neurological damage caused by tin exposure, the structural formula of which is shown in Formula II or Formula V:

[0022]

[0023] Formula II;

[0024]

[0025] Formula V.

[0026] Furthermore, the neurological damage caused by tin exposure is trimethyltin chloride-induced neurological damage; the neurological damage manifests as reduced neuronal activity, impaired neuronal mitochondrial function, and abnormal neuronal mitochondrial morphology.

[0027] Furthermore, the active ingredient of the drug achieves relief of tin exposure-induced neurological damage by binding to the ARMC10 protein, which is phosphorylated at serine position 43.

[0028] This technical solution also provides a compound that antagonizes neuronal damage caused by acute / subacute trimethyltin chloride exposure, the structural formula of which is shown in Formula II or Formula V:

[0029]

[0030] Formula II;

[0031]

[0032] Formula V.

[0033] The technical principle of this technical solution is as follows:

[0034] This technical solution first discovered the pathological regulatory axis of "trimethyltin chloride-phosphorylation of serine at position 43 of ARMC10 protein-mitochondrial dynamics imbalance-nerve damage", and combined with targeted molecular screening, obtained small molecule compounds that can be used to treat tin exposure-induced nervous system damage.

[0035] This study found that ARMC10, an armadillo repetitive sequence protein located on the outer mitochondrial membrane, can affect mitochondrial morphology and function, ATP synthesis efficiency, and the homeostatic clearance of reactive oxygen species (ROS). Trimethyltin chloride (TMT) exposure induces phosphorylation modification of serine residue 43 (S43) in ARMC10. Customized antibodies specifically recognizing ARMC10 (S43) phosphorylation and parallel reaction detection techniques confirmed the increased phosphorylation level. Phosphorylated and non-phosphorylated mutant plasmids were constructed to further clarify that altered phosphorylation at this site induces changes in ARMC10 function, thereby affecting normal mitochondrial function, including: inducing mitochondrial fragmentation, membrane potential collapse, impaired ATP synthesis, and suppressed ROS clearance system function, ultimately leading to neuronal death and manifesting as nervous system damage.

[0036] This study employed precise virtual screening of targeted compounds. Focusing on the S43 phosphorylation site of ARMC10, a three-dimensional structural model of the phosphorylation modification was first created using software to ensure the structural accuracy of the active site (including the electrostatic distribution of the phosphate group and the spatial arrangement of surrounding residues). Subsequently, molecular docking was performed on a ChemDiv compound library containing over 100,000 molecules, using the S43 phosphorylation site as the core. After multi-step screening, five candidate high-affinity compounds were obtained. These high-affinity compounds underwent further experimental studies. The small molecule compounds represented by formulas II (R2) and V (R5) not only specifically bind to the phosphorylated ARMC10 (S43) site but also inhibited the decline in neuronal cell viability, abnormal mitochondrial membrane potential, abnormal mitochondrial ROS levels, and abnormal cellular ATP levels caused by TMT exposure, ultimately achieving a therapeutic effect against TMT-induced neuronal damage. Among them, formula II (R2) showed better efficacy than formula V (R5) and possessed greater potential for drug development.

[0037] Compared with the prior art, the present invention has the following outstanding advantages:

[0038] (1) It fills the technical gap in pathological mechanisms and targets, and for the first time reveals the neurotoxicity regulatory axis of "TMT-ARMC10(S43) phosphorylation-mitochondrial dynamics imbalance", and clarifies that ARMC10(S43) phosphorylation is the key pathogenic target of organotin neurotoxicity. It solves the core bottleneck of existing technologies that lack clear targets for tin exposure-related nerve damage and cannot develop targeted therapeutic drugs. At the same time, it provides a new theoretical framework for the study of the mechanism of mitochondrial-related nerve damage induced by environmental pollutants.

[0039] (2) The strategy of “precise modeling of phosphorylated target proteins + virtual screening” is adopted. Compared with the traditional single high-throughput screening, the targeting specificity (only for S43 phosphorylated ARMC10 protein) and binding activity of candidate compounds are greatly improved. At the same time, the interference of non-specific binding molecules is avoided, and the cost and cycle of subsequent experimental verification are reduced.

[0040] (3) The compounds obtained by screening have practical development potential and binding stability.

[0041] Compounds of Formulas II and V can be directly obtained from commercial compound libraries (ChemDiv). Their chemical structures are well-defined and their synthetic routes are mature, eliminating the need for complex new structural synthesis development. This facilitates rapid advancement of preclinical studies such as dosage form optimization and pharmacodynamic validation. Furthermore, both compounds exhibit high affinity for phosphorylated ARMC10 (S43), providing a molecular basis for sustained neuroprotective effects in vivo.

[0042] (4) The drugs screened in this scheme can intervene in nerve damage in multiple dimensions, and can reverse TMT-induced nerve damage from multiple aspects such as neuronal activity recovery, mitochondrial function repair, and mitochondrial morphological remodeling. Specifically, it can enhance neuronal vitality, restore mitochondrial membrane potential and ATP synthesis capacity, reduce mitochondrial ROS levels, and repair mitochondrial tubular structures, thereby achieving multi-stage blocking of the nerve damage cascade.

[0043] (5) Low off-target risk and broad application scope

[0044] The compound of this invention specifically targets ARMC10 in the S43 phosphorylated state, not the normal unphosphorylated ARMC10 protein, thus avoiding interference with the mitochondrial physiological regulation of normal cells and significantly reducing the off-target toxicity risk of traditional neuroprotective drugs. Furthermore, its targeting effect on ARMC10 (S43) phosphorylation can potentially be extended to neurological damage caused by other organotin compounds that induce ARMC10 phosphorylation, such as tributyltin and dibutyltin, significantly broadening the drug's applicable scenarios. Attached Figure Description

[0045] Figure 1The experimental results of TMT reducing cell viability and inducing mitochondrial dysfunction in Example 1 are shown below: (A: Statistical results of cell confluence and typical images after treatment with different concentrations of TMT for 0, 6, 12, 18, and 24 hours; scale bar: 200 μm; B: Statistical results of mitochondrial membrane potential and typical images; sample size n=3; scale bar: 50 μm; mitochondria in a polarized state exhibit red fluorescent aggregates, i.e., polymeric state; depolarized mitochondria emit green monomeric fluorescence, i.e., monomeric state; C: Statistical results of mitochondrial reactive oxygen species (MitoROS) production and flow cytometry images; sample size n=3; D: Statistical results of ATP levels; sample size n=3; Except for confluence, all the above indicators were detected after Neuro-2a cells were exposed to 0, 2, 4, and 8 μM TMT for 24 hours; the bar charts all use the control group (0 μM TMT) value as 1.0; compared with the control group (0 μM TMT)...) Compared with TMT, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001; ns indicates no statistical significance).

[0046] Figure 2 The results of the TMT-induced excessive mitochondrial fragmentation experiment in Example 1 are shown below (A: Electron micrographs of cell sections treated with 8 μM TMT for 24 hours (treatment group) and untreated (control group); scale bars: 5 μm and 1 μm; B: Representative images and quantitative analysis of mitochondrial morphology in cells treated with 8 μM TMT for 24 hours (treatment group) and untreated (control group); sample size n=10; scale bars: 10 μm and 5 μm; bar charts are all based on the control group value as 1.0; compared with the control group, *** p<0.001; ns indicates no statistical significance).

[0047] Figure 3The following are experimental results of the expression of mitochondrial dynamics-related proteins in cells treated with TMT in Example 1: (A, B: Representative immunoblot images and quantitative analysis results of protein levels of optic atrophy protein 1 (OPA1), mitochondrial fusion protein 1 (MFN1), mitochondrial fusion protein 2 (MFN2), dynamics-related protein 1 (DRP1), and fibrillation protein 1 (FIS1) after cells were exposed to 0, 2, 4, and 8 μM TMT for 24 hours; bar charts are set with the 0 μM TMT group (as control group) as 1.0; sample size n=3; ACTB is the internal reference β-actin; ns indicates no statistical significance; C: Representative immunofluorescence staining images of OPA1, MFN1, MFN2, DRP1, and FIS1 protein expression after cells were exposed to 0 (control group) and 8 μM (treatment group) TMT for 24 hours; TOM20 is the mitochondrial marker mitochondrial outer membrane translocase 20; DAPI is the nuclear staining agent; D: Cells exposed to 0, 2, 4, and 8 μM TMT) Quantitative analysis results of OPA1, MFN1, MFN2, DRP1 and FIS1 protein expression after 24 hours; bar charts were performed with the 0 μM TMT group (as control group) value as 1.0; sample size n=3; scale bar: 40 μm; ns indicates no statistical significance).

[0048] Figure 4 The results of TMT-treated cells in Example 2 are shown in the phosphorylated proteomics analysis (A: number of differentially expressed modified peptides in cells of the treatment group (8 μM TMT, 24 hours) compared with the control group (0 μM TMT, 24 hours); B: cluster heatmap of differentially expressed phosphorylated peptides; red: upregulated expression; blue: downregulated expression).

[0049] Figure 5 The results of the study on the effect of TMT treatment on the phosphorylation level of ARMC10 protein at the S43 site in Example 2 (A: 0 μM TMT, 24 hours in the treated group and the untreated group, 0 μM TMT, 24 hours, 0-log phosphokinase levels of the phosphorylated peptides). 10 Volcano plot of (P-value) versus log2FC (fold change); B: Detailed information on phosphorylation at S43 site of ARMC10 protein; C: Evolutionary conservation of S43 site of ARMC10 protein.

[0050] Figure 6The results of phosphorylated ARMC10 (S43) protein level detection in Example 2 are as follows: (A: Representative immunoblot and quantitative analysis of phosphorylated ARMC10 (S43) protein levels in Neuro-2a cells after treatment with 0, 2, 4, and 8 μM TMT for 24 hours; p-ARMC10 (S43) is phosphorylated ARMC10 (S43) protein; ARMC10 is ARMC10 protein; ACTB is internal reference β-actin; B: Quantitative analysis of A, showing the relative phosphorylation level of phosphorylated ARMC10 (S43) protein; with the control group (0 μM TMT) value as 1.0; compared with the control group (0 μM TMT), ** is p<0.01, *** is p<0.001).

[0051] Figure 7 The phosphorylated ARMC10 (S43) protein level obtained by parallel reaction monitoring in Example 2 (8 μM TMT-treated cells for 24 hours were the experimental group, and 0 μM TMT-treated cells for 24 hours were the control group, with the control group value as 1; sample size n=3; compared with the control group, ***p<0.001).

[0052] Figure 8 The results of Example 2 are as follows: Study on the effects of ARMC10 phosphorylation on TMT-treated Neuro-2a cells and their mitochondrial function (Neuro-2a cells were all transfected with empty plasmid and ARMC10). S43A Plasmid (simulating non-phosphorylated state) or ARMC10 S43D Plasmids (simulating phosphorylation) were then treated with 8 μM TMT or 0 μM TMT for 24 hours before subsequent detection; A: Changes and statistical analysis of Neuro-2a cell confluence; Scale bar: 200 μm; B: Representative images of mitochondrial morphology and quantitative analysis of mitochondrial length; Scale bar: 10 μm and 5 μm; C: Electron micrographs of Neuro-2a cell sections; Scale bar: 1 μm and 5 μm; D-1, D-2: Statistical results and typical images of mitochondrial membrane potential; Sample number n=3; Scale bar: 40 μm; Mitochondria in a polarized state appear as red fluorescent aggregates, i.e., polymeric state; Depolarized mitochondria emit green monomeric fluorescence, i.e., monomeric state; E: ATP level statistical graph; Sample number n=3; F: Statistical results of mitochondrial reactive oxygen species (ROS) production; Sample number n=3; S43A indicates transfection with ARMC10. S43A Experimental group of plasmids; S43D indicates transfection with ARMC10 S43DThe experimental group of plasmids; except for fusion, all bar charts use the empty plasmid (0μm) group (as control group) value as 1.0; compared with the empty plasmid (0μm) group, ** is p<0.01; compared with the empty plasmid (8μm) group, ## is p<0.01).

[0053] Figure 9 This is a schematic diagram of the process for identifying small molecule compounds through virtual screening in Example 3.

[0054] Figure 10 The following are the compound information and experimental verification of compounds R1-R5 selected by virtual screening in Example 3: (A: The top 5 compounds ranked based on affinity value; B: Statistical results of cell viability of Neuro-2a cells after 24 hours of treatment with 8 μM TMT and 10 μM of different candidate compounds or no treatment; with the control group (no candidate compounds and TMT added) as 1.0; C: Cell confluence and statistical results (percentage of cells on the culture surface) of Neuro-2a cells after 24 hours of treatment with 8 μM TMT and 10 μM of different candidate compounds or no treatment; D: Compound structural formulas; Scale bar: 200 μm; Compared with the trimethyltin chloride group (only TMT added, no candidate compounds added), **p<0.01).

[0055] Figure 11 The experimental results of Example 3 on the antagonistic effect of R2 on TMT-induced neurotoxicity, mitochondrial fragmentation, and dysfunction (A: ATP level statistics; B, C: mitochondrial membrane potential (Δψ)). m Images and statistical graphs; scale bar: 20 μm; D, E: images and statistical graphs of mitochondrial reactive oxygen species (MitoROS) generation; F, G: representative images and quantitative analysis statistical graphs of mitochondrial morphology; combined 3 data points, totaling 9 independent cells; scale bar: 10 μm and 5 μm; all the above indicators were detected in Neuro-2a cells after treatment with 8 μM TMT and 10 μM R2 or no treatment for 24 hours; all bar charts were set with the control group (without R2 and TMT) as 1.0; compared with the control group (without R2 and TMT) or the trimethyltin chloride group (only TMT added, no R2 added), **p < 0.01). Detailed Implementation

[0056] The present invention will be further described in detail below with reference to embodiments, but the implementation of the present invention is not limited thereto. Unless otherwise specified, the technical means used in the following embodiments and experimental examples are conventional means well known to those skilled in the art, and the materials and reagents used can all be obtained commercially.

[0057] The key technical terms used in this solution are explained as follows:

[0058] Tin-induced nervous system damage (tin exposure leading to nervous system damage): This refers to the toxic process by which tin (especially organotin compounds) exposure causes pathological changes and functional abnormalities in the nervous system by interfering with cell metabolism and damaging neural structure and function. The core issue is the direct damage of organotin to neurons, mitochondria, etc. Further accumulation of tin-induced nervous system damage can lead to various subsequent harms. Tin (organotin)-induced nervous system damage can trigger a variety of further acute and chronic diseases, including:

[0059] (1) Chronic neurodegenerative disease

[0060] Long-term low-dose exposure can accelerate neuronal degeneration and increase the risk of developing neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease (the mechanism is related to oxidative stress and mitochondrial dysfunction).

[0061] (2) Neurasthenia syndrome

[0062] Symptoms include chronic headaches, insomnia, memory loss, and difficulty concentrating, which affect daily work and quality of life.

[0063] (3) Peripheral neuropathy

[0064] Organotin can damage peripheral nerves, leading to symptoms such as numbness, decreased sensation, and weakness in the limbs.

[0065] In summary, tin-induced nervous system injury (tin exposure leading to nervous system damage) is a type of poisoning disease with clinical significance, its core manifestation being nervous system damage. Therefore, nervous system damage is both a "symptom" of poisoning and a "major clinical manifestation" or "diagnostic criterion" of this poisoning disease. This protocol uses Neuro-2a cells as a research model to study and screen tin-induced nervous system injury (tin exposure leading to nervous system damage) and related drugs.

[0066] Neuro-2a cells:

[0067] The mouse neuroblastoma cell line, derived from neuroblastoma in the adrenal medulla of mice, is a commonly used in vitro model in neuroscience research. It possesses strong proliferative capacity and neuronal functional characteristics, and is frequently used in studies of neurotoxicity, neurodevelopment, neurodegenerative diseases, and drug screening. Compared to primary neurons (which are difficult to culture and have short survival periods), Neuro-2a has simpler culture conditions, stronger resistance to interference, and facilitates experimental procedures such as toxicity treatment and drug intervention. It can simulate the neuronal response to toxins (such as trimethyltin chloride), reflecting the damage of toxicity to nerve cells (mitochondrial function, morphology, etc.); it allows for rapid assessment of the protective / repairing effects of drugs on nerve cells; and it facilitates experiments such as gene editing and protein detection, elucidating neuronal molecular pathways.

[0068] According to literature reports, trimethyltin chloride (TMT) exhibits unique hippocampal selective neurotoxicity, particularly affecting the IC50 of Neuro-2a cells. 50 Typically, 6-10 μM (24h) and 8 μM can induce significant damage while preserving some cell viability. This protocol used 8 μM TMT to treat Neuro-2a cells for 24h to establish an in vitro model of TMT-induced neuronal injury in Neuro-2a cells. This model stably exhibited a TMT-specific neurotoxic phenotype; more specifically, it was an in vitro model of neuronal injury caused by acute / subacute TMT exposure, reflecting the state of nerve cells during acute / subacute organotin (TMT) poisoning. Its core principle is to simulate neuronal damage caused by TMT exposure (characterized by moderate to high toxicity and reproducible mitochondrial / signaling pathway abnormalities), suitable for neurotoxicological mechanism research, drug screening, and biomarker validation. This model directly corresponds to the core pathological aspects of neuronal injury caused by acute / subacute TMT (occupational, environmental) exposure or accidental poisoning, providing cellular-level experimental support for the mechanism analysis of acute poisoning and the validation of therapeutic targets. Without intervention in cases of acute / subacute organotin (TMT) poisoning, it may develop into neurological diseases such as Alzheimer's and Parkinson's.

[0069] ARMC10 protein (Armadillo repeat containing 10; UniProt database unique identifier UniProtID: Q9D0L7): ARMC10, or Armadillo repeat-containing protein 10, is a conserved protein expressed in eukaryotes. Its core characteristic is the presence of multiple armadillo repeat (ARM) domains in its amino acid sequence. These domains, typically composed of 40-45 amino acid residues, mediate protein-protein interactions and are key functional domains for intracellular signal transduction and structural support. ARMC10 proteins are highly conserved across species, especially at key functional sites such as mitochondrial localization signals. This characteristic is a crucial structural basis for maintaining the consistency of its core biological functions across species.

[0070] Phosphorylated ARMC10 (S43) protein: can be abbreviated as p-ARMC10 (S43); a phosphorylated protein formed by phosphorylation of serine at position 43 of the ARMC10 protein.

[0071] Example 1: Trimethyltin chloride (TMT) exposure promotes mitochondrial fragmentation and dysfunction in Neuro-2a cells

[0072] Mouse neuroblastoma cells (Neuro-2a cells) were obtained from the cell bank of the Institute of Biochemistry and Cell Biology (Shanghai, China, TCM29). Cells were treated with 0, 2, 4, or 8 μM TMT for 24 hours (or other specified treatment durations) before subsequent assays. Neuro-2a cells were cultured in DMEM / H (Gibco, C11995500BT) supplemented with 10% fetal bovine serum (AUSGENEX, FBS500-S) and 1% penicillin / streptomycin (Beyotime, C0222) at 37°C and 5% CO2. The TMT stock solution (Sigma-Aldrich, 146,498) was prepared with distilled deionized water and diluted with culture medium.

[0073] After treating cells according to the above TMT treatment method, cell confluence, mitochondrial membrane potential, mitochondrial reactive oxygen species (MitoROS) production, mitochondrial morphology, and mitochondrial dynamics-related proteins were measured. The detection methods were standard existing techniques, and are roughly described as follows: For cytotoxicity detection (detecting cell proliferation), Neuro-2a cells were used at a density of 1 × 10⁻⁶ cells per well. 4Cells were seeded at density in 96-well plates. Cell images were recorded after different treatment times. Cell confluence was analyzed using the Induced Cell Zoom live cell analysis system (EssenBioScience, USA) (characterized by confluence: percentage of cells covering the culture dish surface). For cell membrane potential detection, JC-1 staining was performed on cells treated with different TMT concentrations for 24 hours according to the manufacturer's instructions (Beyotime, C2006). Cellular ATP levels were quantified using an ATP assay kit (Thermo Fisher, A22066). MitoSOX was used to analyze mitochondrial ROS. TM Mitochondrial reactive oxygen species (mtROS) were quantified using a red fluorescent probe (ThermoFisher Scientific, M36008). Mitochondrial morphology was analyzed using the MitoTracker Deep Red FM fluorescent probe (Invitrogen, M22426). Neuro-2a cells were cultured at a density of 1×10⁻⁶. 4 Cells were seeded at a density of 100 cells / well, treated with TMT for 24 h, and then fixed, dehydrated, embedded, sectioned, and observed under an electron microscope. Total protein was extracted from cells after TMT treatment and analyzed by Western blotting (WB) to determine the expression of proteins related to mitochondrial dynamics. Immunocytochemistry was also performed on cells after TMT treatment to determine the expression of these proteins. Statistical analysis was performed on the results, expressed as mean ± SEM. Unpaired two-sided t-tests were used for comparisons between two experimental groups, and one-way ANOVA was used for comparisons among multiple groups. Each experiment was repeated at least three times, and *p < 0.05 was defined as statistically significant.

[0074] The experimental results are as follows: This study used a live-cell analysis system to assess the neurotoxicity of TMT. Specifically, after Neuro-2a cells were exposed to TMT at concentrations of 0, 2, 4, and 8 μM for 24 hours, their survival rate decreased in a dose- and time-dependent manner. Figure 1 A). Given that mitochondrial dysfunction is considered to play a key role in TMT-induced neurotoxicity, we further investigated whether TMT exposure impaired mitochondrial function in Neuro-2a cells. This was determined by measuring mitochondrial membrane potential (Δψ). m The functional status of mitochondria was assessed by measuring mitochondrial reactive oxygen species (mtROS) levels and adenosine triphosphate (ATP) production. Results showed that after 24 hours of treatment with 8 μM TMT, the mitochondrial membrane potential of Neuro-2a cells significantly decreased. Figure 1 B), mtROS levels were significantly elevated ( Figure 1 C) and intracellular ATP levels were significantly reduced ( Figure 1D). In summary, these results indicate that TMT exposure is cytotoxic to Neuro-2a cells and impairs their mitochondrial function. Mitochondrial dynamics dysregulation is a key pathogenic mechanism in a range of diseases characterized by mitochondrial dysfunction. To clarify whether TMT affects mitochondrial dynamics, we evaluated mitochondrial morphology after TMT treatment. MitoTracker staining results showed that after 24 hours of exposure to 8 μM TMT in Neuro-2a cells, mitochondria exhibited significant fragmentation and a significant reduction in mitochondrial length. Figure 2 A). Electron microscopy revealed structural damage in the mitochondria of Neuro-2a cells in the TMT-treated group, including division, swelling, cristae breakage, and mitochondrial membrane rupture. Figure 2 B).

[0075] Mitochondrial dynamics are precisely regulated by a set of key proteins: dyskinetic-associated protein 1 (DRP1) and fission protein 1 (FIS1) dominate mitochondrial fission, while mitochondrial fusion proteins (MFN1 and MFN2) and optic atrophy protein 1 (OPA1) regulate mitochondrial fusion. Notably, although we demonstrated impaired mitochondrial dynamics, the expression levels of the key proteins OPA1, MFN1, MFN2, DRP1, and FIS1 remained unchanged. Figure 3 A and B). Immunofluorescence staining results further validated this finding, showing no statistically significant difference in the overall fluorescence signal intensity of these proteins in the cell population (A and B). Figure 3 (C and D). In summary, these results strongly demonstrate that TMT impairs mitochondrial dynamics; furthermore, the fact that the expression levels of key mitochondrial proteins remained unchanged suggests the existence of other mechanisms regulating TMT-induced neurotoxicity.

[0076] Example 2: Study on the effect of phosphorylation at serine site 43 (S43) of ARMC10

[0077] Neuro-2a cells were treated with 8 μM TMT for 24 hours. Then, phosphorylation modification proteomics analysis based on tandem mass tag was performed on the treated cells (treated group) and untreated cells (control group) (three replicates were set for each group). The analysis process is a routine procedure of existing technology and can be outsourced to a biotechnology company. The detection process will not be described in detail here.

[0078] Mitochondrial homeostasis and functional status are highly dependent on the regulation of protein phosphorylation. Phosphoryptiform proteomics analysis detected 922 upregulated phosphorylation sites and 949 downregulated phosphorylation sites (fold change >1.2 or <0.83, P<0.05). For details on the number of differentially expressed modified peptides in Neuro-2a cells of the treatment group compared to the control group, please refer to [link to relevant documentation]. Figure 4 A, For a detailed clustering heatmap of differentially expressed phosphorylated peptides, see [link to heatmap]. Figure 4 B. Analysis showed that TMT treatment significantly upregulated the phosphorylation level of a single amino acid site, serine 43 (S43), in the ARMC10 protein in Neuro-2a cells. Figure 5 A and B). ARMC10 is an anchoring protein located on the outer mitochondrial membrane, containing a classic ankyrin repeat domain that mediates protein-protein interactions. Sequence homology analysis showed that the S43 site is highly conserved across multiple species, suggesting its important role in the function of ARMC10. Figure 5 C).

[0079] Based on the above research results, a rabbit anti-phosphorylated ARMC10 (S43) antibody was prepared using conventional methods. This antibody can specifically recognize the aforementioned phosphorylated protein and can be used for subsequent research. Experimental results showed that the prepared rabbit anti-phosphorylated ARMC10 (S43) specific antibody selectively binds to phosphorylated peptides containing the S43 site, without reacting with the non-phosphorylated control peptide, confirming the antibody's high specificity. Western blot (WB) experiments using this site-specific antibody revealed that the phosphorylation level of ARMC10 at the S43 site in Neuro-2a cells increased in a concentration-dependent manner after TMT treatment. Figure 6 A and B). The specific protein level of phosphorylated ARMC10 (S43) is determined by... Figure 6 The relative phosphorylation level of phosphorylated ARMC10 (S43) protein in B was quantified. The calculation method was as follows: First, the relative phosphorylation level of ARMC10 (S43) protein in B was... Figure 6 The gray values ​​of the p-ARMC10(S43) band and the ACTB band in protein A are calculated as follows: the ratio represents the relative content of p-ARMC10(S43) per unit of total protein; the gray values ​​of the ARMC10 band and the ACTB band are then calculated as follows: the ratio represents the relative content of total ARMC10 per unit of total protein. Finally, the two ratios are divided again, and the result is used for statistical analysis of the relative phosphorylation level of phosphorylated ARMC10(S43) protein.

[0080] The level of p-ARMC10 (S43) in Neuro-2a cells treated with 8 μM TMT for 24 hours was quantified using parallel reaction monitoring (PRM) technology. Detailed results can be found in [link to results]. Figure 7 Further evidence confirms that TMT treatment increases phosphorylation at the S43 site of ARMC10. These results indicate that serine 43 (S43) is a key phosphorylation site in the ARMC10 protein, and its phosphorylation is involved in the TMT-mediated neurotoxicity process.

[0081] To confirm the role of the S43 phosphorylation site of ARMC10 in TMT-induced neurotoxicity, two mutant plasmids of the ARMC10 protein were constructed using conventional methods: ARMC10 S43A (The serine (S) at position 43 is replaced with alanine (A), and the corresponding codon is mutated from TCC to GCC to simulate the non-phosphorylated state) and ARMC10 S43D (Serine at position 43 (S) is replaced with aspartic acid (D), and the corresponding codon is mutated from TCC to GAC to simulate phosphorylation.) The empty plasmid is the commercially available pcDNA3.1-C-3×FLAG (a commonly used mammalian expression vector for efficient expression of recombinant proteins with a C-terminal 3×FLAG tag in eukaryotic cells). The expression plasmid is constructed by inserting the target gene sequence into the corresponding multiple cloning site. The plasmids corresponding to the two mutants are ARMC10. S43A Plasmid (simulating non-phosphorylated state) or ARMC10 S43D The preparation process of the plasmid (simulating phosphorylation) will not be detailed here. The mutant plasmid and the empty plasmid were transfected into Neuro-2a cells using standard techniques. The cells were then treated with 8 μM TMT for 24 hours or without TMT treatment. Cell confluence statistics and mitochondrial membrane potential Δψ were then analyzed. m Analysis included ATP level analysis, mitochondrial reactive oxygen species (MitoROS) production analysis, cell section preparation and electron microscopy analysis, and mitochondrial observation.

[0082] The results showed that the non-phosphorylated mimic mutant ARMC10 S43A It can significantly reverse the decrease in cell viability caused by TMT treatment, while the phosphorylation mimic mutant ARMC10 S43D Then it will not have this effect. Figure 8 A). Mitochondrial tracking probe staining and electron microscopy results indicate that overexpression of ARMC10... S43A (instead of ARMC10) S43D It can significantly improve mitochondrial dynamics and reduce mitochondrial fragmentation. Figure 8 (B and C). Furthermore, overexpression of ARMC10 S43AIt can significantly reverse TMT-induced mitochondrial dysfunction, manifested as reduced mitochondrial reactive oxygen species production (ROS). Figure 8 F), as well as ATP levels and mitochondrial membrane potential (Δψ). m ) rise ( Figure 8 D-1 Figure 8 D-2 and E). In summary, these results suggest that dephosphorylation of ARMC10 at the S43 site can mitigate TMT-induced neurotoxicity by improving mitochondrial dynamics and function.

[0083] Example 3: Screening for small molecule compounds that may inhibit ARMC10 phosphorylation and antagonize TMT-induced neurotoxicity based on ARMC10 protein structure.

[0084] (1) Virtual Filtering

[0085] Virtual screening utilizes multiple small molecule compound libraries containing over 100,000 molecules. First, phosphorylation modeling of the S43 phosphorylation modification of the ARMC10 protein was performed using chai-1 software (https: / / lab.chaidiscovery.com / ) to ensure the accuracy of the protein's active site structure. Then, AutoDockTools 1.5.7 software was used to define the docking box, with the S43 phosphorylation site as the core (X: -3.278, Y: 11.306, Z: 5.139), and a range of X: 70, Y: 126, Z: 70, providing a precise spatial range for subsequent molecular docking. Small molecules from the ChemDiv compound library were preprocessed using the LigPrep program, preserving the inherent chiral characteristics of the compounds. Each molecule generated a maximum of 32 dominant conformations, ensuring comprehensive and diverse conformational coverage during screening and providing a rich conformational basis for subsequent docking. A three-stage screening strategy was employed, sequentially performing high-throughput primary screening, precise virtual screening, and fine screening. All docking operations were completed using AutoDock Vina v1.2.5 software. To ensure the effectiveness and specificity of the screening, each screening stage ranked candidates based on their docking scores, selecting the top 10% of candidate compounds to proceed to the next stage, progressively enriching potential active molecules with high affinity for phosphorylated ARMC10 (S43) protein. Virtual screening is a standard technique in the field of biopharmaceutical development and can be outsourced to qualified biotechnology companies. The core technical points of the virtual screening described in this protocol are: based on the phosphorylation modification characteristics of the S43 site of the ARMC10 protein, molecular structure modeling (protein crystal structure prediction) of the target protein is conducted; subsequently, from a small molecule compound database with a capacity of over 100,000 molecules, small molecule compounds capable of generating high-affinity interactions with the binding pocket adjacent to the S43 phosphorylation site of the ARMC10 protein are screened as candidate drugs for antagonizing organotin neurotoxicity.

[0086] This study found that TMT can induce phosphorylation at the S43 site of ARMC10. This modification mediates mitochondrial dynamics disorder and dysfunction, ultimately leading to neurotoxicity. This result reveals for the first time the regulatory axis of "TMT-ARMC10 S43 phosphorylation-mitochondrial morphology and function abnormality," providing key mechanistic support for neurotoxicity intervention strategies that target and inhibit S43 phosphorylation, block ARMC10-mitochondrial interaction, or both. To screen small molecule compounds targeting the ARMC10 S43 phosphorylation site, the protein crystal structure of phosphorylated ARMC10 was predicted, and a structure-based virtual screening was performed on a compound library containing over 100,000 small molecules. Figure 9 Based on molecular docking analysis and the predicted binding affinity of small molecules to the phosphorylated form of ARMC10(S43), the top five candidate compounds (named R1 to R5) were screened. Figure 10 A). Subsequently, the neuroprotective effects of these candidate compounds against TMT-induced mitochondrial fragmentation and neuronal dysfunction were further evaluated. The chemical formulas of R1, R2, R3, R4, and R5 are given in Formulas I, II, III, IV, and V, respectively. These compounds are available from ChemDiv; compound ID numbers are detailed in [link to compound ID information]. Figure 10 A. Conduct subsequent experimental verification studies.

[0087]

[0088]

[0089]

[0090] (2) Experimental screening

[0091] (2.1) Cell viability experiment

[0092] Cell viability was assessed using the CCK-8 (Cell Counting Kit-8) cell proliferation-toxicity assay kit from Tokyo Dojin. Neuro-2a cells were cultured at 1×10⁻⁶ cells / year. 4Cells were seeded in 96-well plates and incubated overnight. Three replicates were made for each experimental group. When cells reached 80% confluence, the test compounds (10 μM, represented by formulas I, II, III, IV, and V) were added to each well, with or without TMT (8 μM), and the cells were treated for 24 hours. After treatment, the culture medium was removed, and 100 μL of pre-mixed medium containing 10% CCK-8 reagent was added to each well. Three additional wells were added as blank wells to avoid air bubbles interfering with readings. The 96-well plates were incubated for 30–45 min. Absorbance was measured at 450 nm using a microplate reader. Cell viability was calculated using the formula: Cell viability = [(At-Ab) / (Ac-Ab)] × 100%. At represents the absorbance of the treatment wells (medium containing CCK-8, cells under different treatments); Ac represents the absorbance of the control wells (medium containing CCK-8, control group cells); Ab represents the absorbance of the blank wells (medium containing CCK-8, cell-free).

[0093] For detailed experimental results, please refer to Figure 10 B and Figure 10 D. From Figure 10 B (cell viability bar chart) shows that the cell viability of the group treated with only TMT was significantly lower than that of the control group (no TMT and candidate compounds), indicating that TMT treatment inhibited cell viability and established an in vitro model of neuronal injury caused by acute TMT exposure. Based on this in vitro injury model, the addition of candidate compounds R1-R5 resulted in a significant recovery in cell viability in some experimental groups (R2 and R5), while some experimental groups (R1, R3, and R4) did not show any relief from the toxic effects of TMT, and even experienced a slight decrease in cell viability. This indicates that although candidate compounds R1-R5 showed strong affinity for phosphorylated ARMC10 (S43) protein through virtual screening, only after R2 and R5 bound to phosphorylated ARMC10 (S43) protein could they effectively affect the downstream pathways of this protein and thus alleviate the toxic effects of TMT on neurons.

[0094] (2.2) Cytotoxicity detection

[0095] Neuro-2a cells were loaded at a rate of 1 × 10⁻⁶ 4Cells were seeded in 96-well plates and incubated overnight. Three replicates were performed per experimental group. When cells reached 80% confluence, the test compounds (10 μM, represented by formulas I, II, III, IV, and V) were added, with or without TMT (8 μM). The plates were placed in a cell induction Zoom system, and cell images were recorded for 24 hours. Cell confluence was analyzed using a Zoom live-cell analysis system (EssenBioScience, USA).

[0096] For detailed experimental results, please refer to Figure 10 C. The experiment uses cell confluence to reflect cell growth status; higher confluence indicates better cell condition. At 0 hours, there was no significant difference in cell confluence among the groups, all being in the initial seeding state. At 24 hours, the experimental group with only TMT showed significantly lower cell confluence compared to the control group (without TMT and candidate drugs), corresponding to sparser cells in the morphological map. After adding candidate compounds R1-R5, different experimental groups showed different states. The experimental group with R2 showed significantly increased cell confluence and increased cell density in the morphological map, indicating that compound R2 can alleviate the cytotoxicity of TMT. While R5 can also alleviate the cytotoxicity of TMT, its effect is not as strong as R2. Although R1, R3, and R4 have the potential to bind to phosphorylated ARMC10 (S43) protein, they did not show their ability to alleviate the cytotoxicity of TMT in this experiment.

[0097] In summary, among these candidate compounds, R2 exhibited the strongest protective effect, and its systematic name is:

[0098] Methyl-4-[7-chloro-1-(3-fluoro-4-methylphenyl)-4-sulfanylidene-1H,2H,3H,4H,5H-chromeno[2,3-d]pyrimidin-2-yl]benzoate;

[0099] (4-[7-chloro-1-(3-fluoro-4-methylphenyl)-4-thionyl-1H,2H,3H,4H,5H-chromeno[2,3-d]pyrimidin-2-yl]benzoate;

[0100] The linear representation of the SMILES chemical structure of R2 is as follows:

[0101] COC(=O)C1=CC=C(C=C1)C1NC(=S)C2=C(OC3=CC=C(Cl)C=C3C2)N1C1=CC=C(C)C(F)=C1.

[0102] Further research was conducted on R2, and its effects on mitochondria were measured by cell membrane potential, cellular ATP, and mitochondrial morphology.

[0103] (2.3) Detection of cellular ATP

[0104] Quantitative analysis of cellular ATP levels was performed using an ATP assay kit (Thermo Fisher, A22066). Neuro-2a cells were seeded in white-walled 96-well plates and treated with R2 (10 μM) with or without TMT for 24 hours, after which the culture medium was removed. 100 µL of the reaction mixture provided in the kit was added to each well, and the microplate was incubated under standard cell culture conditions (37°C, 5% CO2) for 15 minutes to initiate the chemiluminescence reaction. Immediately after incubation, the chemiluminescence intensity of each well was measured using a microplate reader. A known concentration of ATP stock solution was prepared; this stock solution was serially logarithmically diluted; 100 µL of each gradient of ATP dilution standards was added to individual wells in the 96-well plate (measured simultaneously with the sample wells). The measured chemiluminescence intensity values ​​from each cell sample well were then substituted into the plotted ATP standard curve.

[0105] (2.4) Detection of mitochondrial membrane potential

[0106] JC-1 staining was used to distinguish between polarized and depolarized mitochondria. The JC-1 staining method was used for detection according to the product instructions (Beyotime, C2006). Polarized mitochondria exhibited red fluorescent aggregates with excitation / emission (Ex / Em) wavelengths of 585 / 590 nm; while depolarized mitochondria emitted green monomeric fluorescence with excitation / emission (Ex / Em) wavelengths of 514 / 529 nm. Neuro-2a cells were seeded in confocal culture dishes and treated with R2 (10 μM) with or without TMT for 24 hours. The culture medium was then removed, and the cells were washed three times with PBS. After treatment, JC-1 staining working solution to a final concentration of 1 μg / mL was added to the cells under light-protected conditions at 37°C and 5% CO2, and incubated for 20 minutes. After incubation, the cells were washed twice with Hank's balanced salt solution (HBSS) to remove residual staining reagent. Finally, the red and green fluorescence signals of JC-1 were observed and captured using a Leica TCS SP8 laser confocal scanning microscope (Leica Microsystems, Germany).

[0107] (2.5) Detection of mitochondrial ROS:

[0108] Using MitoSOX TMA red fluorescent probe (Thermo Fisher Scientific, M36008) was used to quantify mitochondrial reactive oxygen species (mtROS). Neuro-2a cells were divided into groups of 1×10⁻⁶. 4 Cells were seeded at a density of 10 μM / well in 6-well plates, treated with or without R2 (10 μM) and TMT for 24 h, and washed once with PBS. Trypsin digestion was performed for 1 min, followed by addition of culture medium to terminate the process. Cells were collected, centrifuged, and washed three times with PBS. MitoSOX was then added, diluted to a final concentration of 5 μM with serum-free DMEM medium. TM Cells were treated with Red probe working solution and incubated for 30 minutes under standard cell culture conditions (37°C, 5% CO2) in the dark. After incubation, cells were washed three times with PBS to remove unbound probes. Immediately after elution, cells were analyzed by flow cytometry using an Accuri C6 flow cytometer (BD Biosciences, USA) to quantify mitochondrial reactive oxygen species (mtROS) levels.

[0109] (2.6) Mitochondrial morphology detection:

[0110] Mitochondrial morphology was analyzed using the MitoTracker Deep Red FM fluorescent probe (Invitrogen, M22426). Neuro-2a cells were seeded in confocal culture dishes and treated with R2 (10 μM) with or without TMT for 24 h. The culture medium was then removed, and the cells were washed three times with PBS. After treatment, the cells were incubated with 200 nM MitoTracker Deep Red FM fluorescent probe staining working solution at 37°C and 5% CO2 in the dark for 20 min. After incubation, the cells were washed twice with PBS to remove residual staining reagent. Finally, images were acquired using a Leica TCS SP8 laser confocal scanning microscope (Leica Microsystems, Germany).

[0111] For the experimental results of (2.3)-(2.6), please refer to [link to relevant documentation]. Figure 11 For the detection and analysis of cellular ATP ( Figure 11A) ATP is a core indicator of cellular energy metabolism. This experiment investigated changes in energy levels through "TMT damage + R2 intervention." The ATP luminescence intensity in the TMT-treated group alone was significantly reduced, reflecting insufficient cellular energy supply. However, when R2 was added to the TMT treatment, the ATP level in the experimental group significantly rebounded, approaching that of the control group (no TMT or R2 treatment). Although R2 treatment alone slightly decreased cellular ATP levels, it effectively alleviated the decline in nerve cell energy metabolism caused by TMT. This further demonstrates that R2 can alleviate the inhibition of cellular energy metabolism by TMT and protect the cell's energy supply capacity.

[0112] Analysis of mitochondrial membrane potential ( Figure 11 B and Figure 11 C) Mitochondrial membrane potential is a core marker of mitochondrial function (polarization = normal function, depolarization = functional impairment). Only the TMT-treated group showed increased green fluorescence (depolarization) and decreased red fluorescence (polarization). However, adding R2 to the TMT treatment resulted in a rebound in the proportion of red fluorescence and a decrease in green fluorescence in the experimental group. This further demonstrates that R2 can reverse TMT-induced mitochondrial depolarization and restore normal mitochondrial function.

[0113] Analysis of mitochondrial ROS in cells ( Figure 11 D and Figure 11 (E) Mitochondrial ROS is a key indicator of oxidative stress damage, and MitoSOX fluorescence intensity is positively correlated with ROS levels. Only the TMT-treated group showed a significant increase in fluorescence intensity (increased mitochondrial oxidative stress). However, the addition of R2 to the TMT treatment significantly reduced the fluorescence intensity in the experimental groups. This indicates that R2 can inhibit TMT-induced excessive production of mitochondrial reactive oxygen species and alleviate oxidative stress damage.

[0114] For mitochondrial morphology detection and analysis ( Figure 11 F and Figure 11 (G) Mitochondrial morphology reflects its structural integrity (normally tubular / reticular, fragmented after damage). Only the TMT-treated group showed fragmented, dispersed mitochondria with shortened mitochondrial length. However, with the addition of R2 to the TMT treatment, the mitochondria in the experimental group recovered to a more continuous tubular / reticular structure, and mitochondrial length was maintained to some extent. This indicates that R2 can protect the structural integrity of mitochondria and prevent mitochondrial morphological damage caused by TMT.

[0115] The above experiments, from four dimensions—energy metabolism, functional state, oxidative stress, and structural integrity—jointly demonstrate that R2 can alleviate the toxic damage of TMT to Neuro-2a cells by protecting mitochondrial function.

[0116] 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. The use of small molecule compounds in the preparation of drugs for treating tin exposure-induced nervous system damage, characterized in that: The structural formulas of small molecule compounds are shown in Formula II or Formula V: Formula II; Formula V.

2. The use of the small molecule compound according to claim 1 in the preparation of a medicament for treating tin exposure-induced nervous system damage, characterized in that: The tin exposure-induced neurological damage refers to acute / subacute neurological damage caused by organotin exposure.

3. The use of the small molecule compound according to claim 2 in the preparation of a medicament for treating tin exposure-induced nervous system damage, characterized in that: The organotin is trimethyltin chloride.

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