Application of alpha-galactosidase inhibitor in preparation of medicine for treating myocardial injury related diseases

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to application of an alpha-galactosidase inhibitor in preparation of a medicine for treating myocardial injury related diseases. According to the invention, GLA-mediated Gal2Cer / GalCer imbalance found by space metabonomics is taken as a core mechanism, and a precise intervention scheme is provided. The targeting property is strong, the myocardial specificity is prominent, the used gene knock-down vector can focus on myocardial tissues to play a role, and the peripheral side effect risk is reduced; meanwhile, the GLA inhibitor is applied to myocardial I / R injury intervention for the first time, and the GLA inhibitor and gene knock-down synergistically prove that the cardiac function can be effectively protected and pathological injuries such as myocardial infarction and fibrosis can be relieved by regulating GLA, and related effects are verified by animal experiments; a new target is provided for treatment of heart diseases such as myocardial I / R injury, the clinical application scene of the GLA inhibitor is expanded, and the GLA inhibitor has innovation value and clinical transformation potential.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological medicine, and particularly relates to application of an alpha-galactosidase inhibitor in preparation of a medicament for treating myocardial injury related diseases. BACKGROUND

[0002] Acute myocardial infarction (MI) is one of the diseases with the highest mortality and disability rates in global cardiovascular diseases. The core pathological mechanism is acute myocardial necrosis caused by interruption of coronary artery blood flow, which further leads to serious complications such as heart failure and malignant arrhythmia, significantly reducing the quality of life and long-term prognosis of patients. At present, reperfusion therapy (such as percutaneous coronary intervention and thrombolytic therapy) is the core strategy for clinical management of MI, which can quickly restore blood flow to ischemic myocardium and save endangered myocardium. However, reperfusion can induce ischemia-reperfusion (I / R) injury, which is manifested as expansion of myocardial necrosis, endothelial dysfunction, calcium homeostasis imbalance and increased oxidative stress, ultimately offsetting the benefits of treatment. Although a large number of studies have been carried out on the mechanism of I / R injury at present, there is still a lack of a consensus treatment that can effectively reduce its adverse effects. The existing candidate strategies are mostly focused on non-specific pathways such as anti-oxidative stress and inhibition of inflammatory response, which have problems such as dispersed action targets, low regulation efficiency, significant systemic side effects, and cannot achieve precise intervention on the core mechanism of the disease.

[0003] Recent studies have confirmed that the occurrence and development of myocardial I / R injury are closely related to global metabolic disorders and local metabolic reprogramming, and lipid metabolism imbalance is the core link of regulating myocardial cell survival and tissue repair. Although traditional metabolomics research has revealed energy metabolism abnormalities during I / R, the analysis method based on tissue homogenate cannot capture the regional specific metabolic differences of myocardial "ischemic core area (irreversible injury) - boundary area (reversible injury) - distal normal area" (i.e. "metabolic spatial heterogeneity"), resulting in a blind spot in the understanding of key regulatory pathways of I / R injury.

[0004] However, there is no scheme in the prior art for treating myocardial I / R injury by targeting the "GLA-Gal2Cer / GalCer axis". On the one hand, although existing GLA small molecule inhibitors (such as GR181413A and migalastat hydrochloride) have been proven to inhibit GLA enzyme activity in vitro, their application scenarios are limited to Fabry disease (a lysosomal storage disease caused by GLA gene mutation), and are used to correct substrate accumulation caused by GLA activity defects in the disease. No research has revealed that they can be used to treat myocardial I / R injury by inhibiting GLA activity and correcting Gal2Cer / GalCer imbalance. Moreover, existing research has completely ignored the abnormal up-regulation and pathogenic role of GLA in myocardial I / R injury. On the other hand, existing schemes for regulating lipid metabolism for myocardial I / R injury all focus on global sphingolipid metabolism (such as ceramide enzyme inhibitors and sphingosine kinase activators), and lack specific regulation of the key target GLA. Accordingly, the present application aims to: first, provide a new application of GLA small molecule inhibitors in treating myocardial I / R injury; and second, provide a MyoAAV1A-shGLA recombinant vector (combined with a myocardium-specific promoter) to achieve myocardial cell-specific GLA knockdown and correct sphingolipid imbalance in I / R injury myocardium. Both of them take "targeting the core pathogenic mechanism mediated by GLA" as the innovation point, effectively inhibit myocardial necrosis and reduce fibrosis, thereby improving cardiac function. SUMMARY

[0005] Our team, using matrix-assisted laser desorption / ionization mass spectrometry (MALDI-MSI, a spatial metabolomics technique that enables micron-level resolution metabolite localization while preserving tissue morphology) combined with spatial clustering analysis, has for the first time discovered that the sphingolipid metabolic pathway is one of the most spatially heterogeneous pathways in myocardial I / R injury: Six hours after reperfusion, galactosylceramide (GalCer) significantly accumulated in the ischemic core and boundary regions, while digalactosylceramide (Gal2Cer) was significantly depleted in these regions. Furthermore, the degree of this Gal2Cer / GalCer imbalance was positively correlated with the area of ​​myocardial necrosis and the degree of fibrosis. Further mechanistic verification revealed that α-galactosidase (GLA) is the key enzyme driving this imbalance—GLA specifically hydrolyzes Gal2Cer to generate GalCer, and this process occurs 24 hours after I / R injury. Within hours, its expression level was upregulated 2.4 times in the myocardial boundary region (a key intervention area for reversible injury) compared to normal myocardium (based on the GSE138201 dataset and immunofluorescence validation), directly leading to an imbalance in the Gal2Cer / GalCer ratio, ultimately exacerbating I / R damage by promoting cardiomyocyte apoptosis. This finding clarifies for the first time that GLA-mediated sphingolipid imbalance is the core regulatory mechanism of myocardial I / R injury, providing a novel target for precise intervention of the disease.

[0006] In response, the present invention proposes the following technical solution: The first aspect of this invention aims to provide the use of α-galactosidase inhibitors in the preparation of medicaments for treating myocardial injury-related diseases.

[0007] A second aspect of the present invention aims to provide the use of α-galactosidase as a target in the development of medicaments for treating myocardial ischemia-reperfusion injury.

[0008] To achieve the above-mentioned objectives of this invention, the technical solution adopted by this invention is as follows: In a first aspect, the invention provides the use of α-galactosidase inhibitors in the preparation of medicaments for treating myocardial injury-related diseases.

[0009] The transcript number of α-galactosidase (GLA) in the NCBI database is NM_013463.3, and the protein ID in the UniProt database is P51569.

[0010] In some embodiments of the present invention, the α-galactosidase inhibitor comprises at least one of the following: (a1) Substances that inhibit the activity of α-galactosidase protein; (a2) Substances that reduce the content of α-galactosidase protein; (a3) Substances that knock out the α-galactosidase gene; (a4) Substances that inhibit the expression of α-galactosidase gene.

[0011] In some embodiments of the present invention, the α-galactosidase inhibitor includes nucleic acid molecules, protein molecules, and small molecule compounds.

[0012] In some embodiments of the present invention, the nucleic acid molecule is microRNA, siRNA, shRNA, dsRNA, sgRNA and / or antisense oligonucleotide.

[0013] In some embodiments of the present invention, the target sequence of the shRNA is 5′-GCTAGGGATTTATGCAGAT-3′ (SEQ ID NO: 1).

[0014] In some embodiments of the present invention, the protein molecule is a specific antibody against α-galactosidase.

[0015] In some embodiments of the present invention, the small molecule compound includes migastrol.

[0016] Migalastat has the chemical formula C6H. 13 NO4, CAS No.: 108147-54-2, structural formula as shown in (Formula I): (Formula I).

[0017] In some embodiments of the invention, the migasstat comprises a pharmaceutically acceptable salt or a pharmaceutically acceptable modification. For example, migasstat hydrochloride: CAS No.: 75172-81-5, with the structural formula shown in (Formula II): (Formula II).

[0018] In some embodiments of the present invention, the pharmaceutically acceptable salt includes acid addition salts and base addition salts.

[0019] "Pharmaceutically acceptable acid addition salts" refer to salts that retain the biological effectiveness and properties of the free base, are not undesirable in biological or other respects, and are formed from an inorganic acid and an organic acid, such as, but not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc., and such as, but not limited to, acetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, camphoric acid, camphor 10 sulfonic acid, decanoic acid, hexanoic acid, caprylic acid, carbonic acid, cinnamic acid, citric acid, cyclohexanesulfonic acid, dodecyl sulfate, ethane 1,2 disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, fumaric acid, galactopyric acid, gentian acid, gluconic acid, glucuronic acid, glutamic acid, glutamate, 2-oxoglutamate, glycerophosphate, glycolic acid, hippuric acid, isobutyric acid, lactic acid, lactobionic acid, lauric acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, etc.

[0020] "Pharmaceutically acceptable base addition salts" refer to salts that retain the biological effectiveness and properties of the free acid and are not undesirable in biological or other respects. These salts are prepared by the addition of an inorganic or organic base to a free acid. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Preferred inorganic salts are ammonium, sodium, potassium, calcium, and magnesium salts. Salts derived from organic bases include, but are not limited to, the following: primary, secondary, and tertiary amines; substituted amines (including naturally occurring substituted amines); cyclic amines; and basic ion exchange resins such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, diethanolamine, ethanolamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, ethylenediamine, glucosamine, methylglucosamine, theobromine, triethanolamine, thiazoline, purine, piperazine, piperidine, N-ethylpiperidine, and polyamine resins. Particularly preferred organic bases are isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine.

[0021] In some embodiments of the present invention, the pharmaceutically acceptable modification includes at least one of phosphorylation, sulfonation, acylation, glycosylation, ubiquitination, acetylation, methylation, sulfation, phospholipidation, and halogenation.

[0022] In some embodiments of the present invention, the medicament includes pharmaceutically acceptable excipients and / or any one or more other active ingredients.

[0023] In some embodiments of the present invention, the pharmaceutically acceptable excipients include at least one of the following: solvents, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, flow aids, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesion agents, integrators, penetration enhancers, pH adjusters, buffers, plasticizers, surfactants, defoamers, thickeners, encapsulating agents, humectants, absorbents, diluents, flocculants and anti-flocculators, filter aids, release inhibitors, and carriers.

[0024] The pharmaceutically acceptable excipients mentioned above are generally recognized for use in this purpose and as inactive ingredients in the pharmaceutical preparation. Compilations of pharmaceutically acceptable excipients can be found in reference books such as the *Handbook of Pharmaceutical Excipients* (2nd edition, edited by A. Wade and PJ Weller; published by the American Pharmaceutical Association, Washington and The Pharmaceutical Press, London, 1994) and the *Pharmacopoeia of the People's Republic of China - List of Pharmaceutical Excipients*.

[0025] In some embodiments of the present invention, the dosage form of the drug includes a gastrointestinal dosage form or a non-gastrointestinal dosage form.

[0026] In some embodiments of the present invention, the gastrointestinal dosage form includes at least one of powder, tablet, granule, capsule, sustained-release, solution, dry suspension, effervescent tablet, emulsion, suspension, syrup, drops, and chewable tablet; In some embodiments of the present invention, the non-gastrointestinal drug delivery dosage form includes at least one of injection dosage form, respiratory dosage form, skin dosage form, mucosal dosage form, and cavity dosage form.

[0027] In some embodiments of the present invention, the dosage form of the drug is a dosage form suitable for children or a dosage form suitable for adults.

[0028] In some embodiments of the present invention, the product is applied to mammals; In some embodiments of the invention, the mammal includes humans.

[0029] A second aspect of the invention provides the use of α-galactosidase as a target in the development of medicaments for treating myocardial ischemia-reperfusion injury. The beneficial effects of this invention are: This invention utilizes the GLA-mediated Gal2Cer / GalCer imbalance, discovered through spatial metabolomics, as its core mechanism to provide a precise intervention strategy. It exhibits strong targeting and significant myocardial specificity; the gene knockdown vector used focuses its effect on myocardial tissue, reducing the risk of peripheral side effects. Furthermore, it marks the first application of GLA inhibitors in the intervention of myocardial I / R injury, demonstrating, in synergy with gene knockdown, that regulating GLA can effectively protect cardiac function and alleviate pathological damage such as myocardial infarction and fibrosis. These effects have been validated in animal experiments. This not only provides a new target for the treatment of myocardial I / R injury and other heart diseases but also expands the clinical application scenarios of GLA inhibitors, possessing both innovative value and clinical translational potential. Attached Figure Description

[0030] The present invention will be further described below with reference to the accompanying drawings and embodiments, wherein: Figure 1 The spatial distribution of key substances in the ceramide metabolic pathway after IR surgery is shown; dark red indicates that the substances in this region are significantly higher than those in other regions, and dark blue indicates that the substances in this region are significantly lower.

[0031] Figure 2 The results are thermal images of ceramides.

[0032] Figure 3 The results of quantitative and statistical analysis of ceramide substances in the injury zone (purple bar), the boundary zone near the injury zone (dark blue bar), and the healthy zone far from the injury (yellow bar).

[0033] Figure 4 The results show a significant increase in α-galactosidase (GLA) at the injury boundary. A represents the transcriptional level of ceramide metabolic enzymes in human myocardial infarction, with data from dataset (#GSE138201). B represents the immunofluorescence staining of GLA and cTNI 3 days after myocardial ischemia-reperfusion injury in mice. cTNI is a cardiomyocyte-specific troponin that can be used to label cardiomyocytes. The healthier the myocardium, the brighter the cTNI fluorescence.

[0034] Figure 5 The results of the experiment on knocking down α-galactosidase in the heart are shown. A is the Western blot of GLA protein, indicating that the protein was successfully knocked down; B is a representative echocardiogram of mouse heart, indicating that knocking down GLA can protect cardiac function; C is TTC (red area is the area of ​​myocardial survival, white area is the area of ​​myocardial death) and Evans Blue staining (blue area is the area of ​​healthy myocardium without ischemia).

[0035] Figure 6The results show the protective effect of α-galactosidase inhibitors (GLAi) on cardiac function. A is a representative echocardiogram of a mouse heart, indicating that GLA inhibitors can protect cardiac function. B is a Masson staining of the heart of a mouse 7 days after ischemia-reperfusion injury (red indicates the surviving myocardial area, and blue indicates the fibrotic area). Detailed Implementation

[0036] The following will describe the concept and technical effects of the present invention clearly and completely with reference to embodiments, so as to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are all within the scope of protection of the present invention.

[0037] This technical approach is based on spatial metabolomics research (confirming that GLA-mediated Gal2Cer / GalCer imbalance is the core mechanism of myocardial I / R injury). Two types of GLA-targeted regulatory technologies were designed, and their ameliorative effects on myocardial I / R injury were verified through animal experiments. Specific core information is as follows: In the functional validation technology of GLA small molecule inhibitors, the GLA competitive inhibitor GR181413A (migalastat hydrochloride, catalog number HY-14929A) was selected. C57BL6 mice were continuously injected intraperitoneally with GR181413A (3 mg / kg) for 10 days to inhibit GLA activity. A myocardial infarction / reperfusion (I / R) model was constructed. 24 hours after I / R injury, the Gal2Cer / GalCer ratio was determined by TTC staining and mass spectrometry to confirm the inhibitor's effect on reducing infarct size and correcting metabolic imbalance. One week after I / R, the protective effect on cardiac function was demonstrated by mouse echocardiography, Masson staining, and hemodynamic monitoring.

[0038] In the construction and functional validation of the AAV9-cTnT-shGLA vector, the GLA transcript used was NM_013463.2, the target sequence of sh-GLA. Two weeks before I / R injury, GLA knockout in mouse cardiomyocytes was achieved by intravenous injection of myoAAV1a-cTnT-shRNA. Western blotting experiments confirmed that this vector could reduce myocardial GLA expression. A myocardial I / R model was constructed, and the infarct area was measured by TTC 24 hours after I / R injury. One week after surgery, its protective effect on the heart was demonstrated by small animal ultrasound and Masson staining.

[0039] Example 1: GLA-mediated Gal2Cer / GalCer imbalance is a core mechanism of myocardial I / R injury. 1. Establishment of a myocardial ischemia-reperfusion injury model: Mice were anesthetized with 4.0% isoflurane, followed by maintenance with 1.5–2.0% isoflurane, placed on a heated surgical board, and cannulated with a 22-gauge plastic catheter. The catheter was connected to a circulatory system providing supplemental oxygen at a tidal volume of 225–250 mL and a respiratory rate of 120–130 strokes / min. A left thoracotomy was performed on a volume-circulating Harvard Rodent ventilator under mechanical ventilation. Under direct visualization, the pericardial sac was opened, and the left anterior descending (LAD) artery was occluded near its origin using an 8-0 suture. Myocardial ischemia was confirmed by myocardial whitening and ST segment elevation on continuous ECG monitoring. After 30 minutes of ischemia, the sutures were loosened to induce reperfusion injury, confirmed by ST segment depression on ECG. The same procedure was performed on the sham-operated group, with the sutures inserted under the LAD without obstruction. Body temperature was monitored with a rectal probe and maintained using a heated surgical platform. The chest wall was sealed in layers, and the mice were transferred to a temperature-controlled mat for recovery.

[0040] 2. Experimental Results This study used 8-week-old mice as the research subjects. A model was established through sham surgery and I / R surgery. Myocardial tissue was prepared into 10 μm frozen sections, and MALDI-MSI technology was used to analyze changes in mouse cardiac metabolism. The results showed that the sphingolipid metabolic pathway underwent significant spatial metabolic remodeling after I / R injury. Sphingolipids are a general term for lipids with a sphingosine backbone. Sphingomyelin (SM) is an important component of the phospholipid bilayer of myocardial cell membranes and can be hydrolyzed by sphingomyelinase to generate ceramide (Cer). Cer, as a key signaling hub in the sphingolipid metabolic network, can be galactosylated to generate galactosylceramide (GalCer) and digalactosylceramide (Gal2Cer). This study used spatial metabolomics analysis to map the spatiotemporal distribution of ceramides and glycosphingolipids after myocardial ischemia-reperfusion (I / R) injury. Figure 1 ).

[0041] The results showed that 24 hours after I / R injury, the levels of ceramide metabolites such as Cer (d18:0 / 22:1, d18:0 / 24:1) and glucosylceramide (GlcCer) (d18:1 / 12:0, d18:1 / 14:0, d18:0 / 16:0) in the myocardium were significantly increased, and remained higher than those in the sham surgery group until 7 days after I / R. Figure 2 Gal2Cer (d18:1 / 16:0) showed unique fluctuations, with its I / R ratio dropping below that of the sham surgery group after 30 minutes of ischemia, and the decrease was more significant after 6 hours of I / R. Although it recovered afterward, it did not return to the baseline throughout the entire process.

[0042] Relative quantitative analysis of Cer and representative glycosphingolipids in the distal ischemic zone, border zone, and infarct zone after myocardial I / R injury revealed that, starting 6 hours after I / R, the levels of sphingomyelin (SM, d18:1 / 20:0, d18:1 / 24:0), Cer (d18:1 / 24:1), sphingosine-1-phosphate (S1P), and galactosylceramide (GalCer, d18:1 / 20:0) in the ischemic injury zone and border zone were significantly higher than those in the distal ischemic zone. Gal2Cer (d18:1 / 16:0) showed a differentiated pattern: it was significantly reduced in the ischemic boundary and injury areas 6 hours after I / R (suggesting initial disruption of glycosphingolipid homeostasis), compensatory upregulation occurred in the distal myocardium and ischemic boundary areas 24 hours after I / R, and then decreased again on day 7 after I / R. Furthermore, Gal2Cer levels remained consistently suppressed in the ischemic injury areas throughout the entire process. Figure 3 This suggests that GalCer plays a crucial role in the spatial regulation of myocardial repair. In summary, GalCer and Gal2Cer exhibit unique spatiotemporal dynamics in myocardial I / R injury, and their differential regulation may affect the balance between injury and repair.

[0043] As the rate-limiting enzyme catalyzing the hydrolysis of digalactosylceramide (Gal2Cer) to galactosylceramide (GalCer), transcriptome data analysis (GSE138201 dataset) showed that GLA mRNA expression levels in myocardial tissue 24 h after myocardial infarction were significantly upregulated compared to normal myocardium. Furthermore, immunofluorescence staining results confirmed that in the ischemic boundary region following cardiac I / R injury, GLA protein expression was significantly higher than in distal normal myocardium. Figure 4These results suggest that I / R injury may accelerate the hydrolysis of Gal2Cer into GalCer by inducing abnormally high expression of GLA in ischemic critical areas, thereby disrupting the metabolic balance between GalCer and Gal2Cer in the local myocardium. This region-specific sphingolipid metabolic imbalance may be a key factor driving the pathological progression of I / R injury.

[0044] Example 2: Cardioprotective effect of α-galactosidase 1. Establishment of a myocardial ischemia-reperfusion injury model: Same as in Example 1. Simultaneously, a GLA knockdown group and a GLA competitive inhibitor GR181413A treatment group were established (3 mg / kg intraperitoneally for 10 consecutive days).

[0045] 2. Knockdown of GLA expression in mouse hearts The GLA transcript used in the experiment was numbered NM_013463.2, and the target sequence of the GLA interfering small RNA (sh-GLA) was 5′-GCTAGGGATTTATGCAGAT-3′ (SEQ ID NO: 1). Two weeks before constructing the myocardial ischemia-reperfusion (I / R) injury model, the GLA gene was knocked down in mouse cardiomyocytes by intravenous injection of myoAAV1a-cTnT-shRNA (myocardial-specific interference vector).

[0046] 3. Western blot (WB) detection of target protein levels 1) Electrophoresis: Prepare a 12% gel using the kit and electrophore at a suitable voltage for about 1 hour.

[0047] 2) Transfer: Protein blotting was performed using the Tris-glycine system. The 0.22 μm PVDF membrane was used and the membrane was transferred at a constant current of 220 mA for 2 h under ice bath conditions.

[0048] 3) Blocking: Use 3% BSA solution (solvent is TBST solution) to block for 2 h at room temperature.

[0049] 4) Primary antibody incubation: Dilute the primary antibody 1000 times with 3% BSA solution and add it to the transferred PVDF membrane. Incubate overnight at 4°C in a rocker incubator.

[0050] 5) Wash, discard the primary antibody solution, wash with an appropriate amount of TBST solution, at room temperature, wash 3 times, 5 min each time.

[0051] 6) Secondary antibody incubation: Dilute the secondary antibody 3000 times with 3% BSA solution and add it to the transferred PVDF membrane. Incubate at room temperature in a rocker shaker.

[0052] 7) Wash, discard the secondary antibody solution, and wash with an appropriate amount of TBST solution at room temperature for 3 times, 5 min each time.

[0053] 8) Imaging: Using Bio Rad's chemiluminescence analyzer and ECL developer, develop and image the data and save the images.

[0054] 4. Evas blue and TTC staining Twenty-four hours after I / R injury in mice, the LAD was re-ligated, and 3% Evas blue was perfused into the aorta. Heart tissue was removed, blood was pumped out in preheated physiological saline at 37°C, and the tissue was placed in a cardiac fixation device and cut into 5 pieces. The 5 pieces of heart tissue were directly placed in 2% 2,3,5-triphenyltetrazolium chloride solution and incubated at 37°C for 15 minutes. After fixation with 4% PFA for 10 minutes, the tissue was examined under a stereomicroscope.

[0055] 5. Small animal cardiac ultrasound: 1) Echocardiography was performed using the Vevo® 3100 system and MS-400 Red probe. In short, the hair on the chest of each mouse was first plucked with depilatory cream, then the mice were gently bound and placed on a platform, followed by a rapid echocardiogram (<5 minutes). This rapid echocardiogram minimizes discomfort for the mice.

[0056] 2) Record the electrocardiogram of the mouse during the operation and confirm the success of the model by observing ST segment-specific elevation.

[0057] 3) Seven days after the IR procedure, perform echocardiography to record short-axis B-mode images parasternally, and then measure and analyze as described. Select the representative image that is closest to the quantified mean.

[0058] 6. Experimental Results To clarify the effects of GLA on cardiac function and tissue remodeling after myocardial I / R injury, this embodiment constructed a myocardial-specific GLA knockdown vector myoAAV1a-cTnT-shGla (AAV-shGla). The results are as follows: Figure 5 As shown, knocking down GLA specifically in the myocardium can significantly reduce the area of ​​myocardial infarction and myocardial fibrosis induced by I / R injury and improve cardiac function.

[0059] In addition, this embodiment used the highly selective GLA competitive inhibitor GR181413A (mirastat hydrochloride) to inhibit GLA activity in mice via continuous intraperitoneal injection (3 mg / kg, for 10 days). One week after I / R injury, echocardiography showed that the left ventricular ejection fraction (LVEF) and fractional shortening (LVFS) of the GR181413A group were significantly higher than those of the control group, while the degree of myocardial fibrosis was significantly reduced. Figure 6 ).

[0060] In summary, both intervention methods—knocking down GLA expression using a myocardial-specific vector and inhibiting GLA activity using GR181413A pharmacology—effectively protect cardiac function after I / R injury, further confirming that GLA is a key target for regulating myocardial I / R injury and providing a clear direction and feasible approach for clinical intervention.

[0061] The embodiments of the present invention have been described in detail above with reference to the accompanying drawings. However, the present invention is not limited to the above embodiments, and various changes can be made within the scope of knowledge possessed by those skilled in the art without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof can be combined with each other unless otherwise specified.

Claims

1. Use of an α-galactosidase inhibitor in the manufacture of a medicament for treating a myocardial injury-related disease.

2. The use according to claim 1, wherein: the α-galactosidase inhibitor comprises at least one of: (a1) a substance that inhibits the activity of an α-galactosidase protein; (a2) a substance that reduces the content of an α-galactosidase protein; (a3) a substance that knocks out an α-galactosidase gene; and (a4) a substance that inhibits the expression of an α-galactosidase gene.

3. The use according to claim 2, wherein: the α-galactosidase inhibitor comprises a nucleic acid molecule, a protein molecule, or a small molecule compound; preferably, the nucleic acid molecule is a microRNA, an siRNA, an shRNA, a dsRNA, an sgRNA, and / or an antisense oligonucleotide; and preferably, the protein molecule is a specific antibody of an α-galactosidase.

4. The use according to claim 3, wherein: the small molecule compound comprises migalastat.

5. The use according to claim 4, wherein: the migalastat comprises a pharmaceutically acceptable salt.

6. The use according to claim 1, wherein: the myocardial injury-related disease comprises, but is not limited to, myocardial ischemia-reperfusion injury, acute myocardial infarction, and heart failure.

7. The use according to claim 1, wherein: the medicament comprises a pharmaceutically acceptable excipient, and / or any one or more other active ingredients.

8. The use according to claim 7, wherein: the medicament is in a dosage form that is administered via the gastrointestinal tract, or in a non-gastrointestinal tract dosage form.

9. The use according to claim 8, wherein: the medicament is administered to a mammal; and preferably, the mammal comprises a human.

10. Use of an α-galactosidase as a target in the development of a medicament for treating cardiac ischemia-reperfusion injury. ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​

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