Application of Chipericumin D in preparation of medicine for treating calcified aortic valve diseases

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to an application of Chipercumin D in preparation of a medicine for treating calcified aortic valve diseases. The core purpose of the invention is to provide a large-scale drug screening method based on Dot-blotting, and the drug is searched for inhibiting osteogenic differentiation of aortic valve interstitial cells and reducing expression of osteogenic specific genes ALP and RUNX2 and formation of calcified nodules, so that the purpose of preventing or treating calcified aortic valve diseases (CAVD) is achieved.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and in particular relates to an application of chipericumin D in the preparation of a medicine for treating calcific aortic valve disease. Background Art

[0002] Calcific aortic valve disease (CAVD) is a serious heart valve disease with an increasing global prevalence. Currently, transcatheter aortic valve implantation (TAVI) or surgical aortic valve replacement (SAVR) are the only treatment options. However, surgical treatment brings many complications, high medical costs, and a lower quality of life. Currently, there are no effective drugs to reverse or treat CAVD. Therefore, it is urgent to find drugs to prevent or treat CAVD through large-scale drug screening.

[0003] Studies have shown that the main pathological features of CAVD include endothelial damage, lipid deposition, inflammatory response, and osteoblastic differentiation of valvular interstitial cells, ultimately leading to collagen deposition, extracellular matrix remodeling, and the formation of calcified nodules. Among them, osteoblastic differentiation of aortic valvular interstitial cells plays a crucial role in the pathogenesis of CAVD, accompanied by upregulation of the osteoblast-specific genes ALP and RUNX2. Prior art has not explored the application of Chipericumin D, an extract from the Chinese herbal medicine Hypericum perforatum, in CAVD.

[0004] Defects of existing technology:

[0005] 1. Currently, there is a lack of effective treatments for calcific aortic valve disease;

[0006] 2. There is no large-scale drug screening targeting osteoblastic differentiation of aortic valve interstitial cells;

[0007] 3. The application of chipericumin D in calcific aortic valve disease has not been explored. Summary of the Invention

[0008] The core purpose of the present invention is to provide a large-scale drug screening method based on Dot-blotting to find drugs that inhibit the osteoblastic differentiation of aortic valve interstitial cells, reduce the expression of osteoblast-specific genes ALP and RUNX2, and the formation of calcified nodules, thereby achieving the purpose of preventing or treating calcific aortic valve disease (CAVD).

[0009] The present invention provides an application of Chipericumin D in preparing a medicine for preventing or treating calcific aortic valve disease.

[0010] Furthermore, the drug can reduce the expression of osteogenesis-related genes ALP and RUNX2.

[0011] Furthermore, the Chipericumin D is used to prepare a drug for inhibiting osteogenic differentiation of VIC.

[0012] Furthermore, Chipericumin D is the only active ingredient in the drug.

[0013] The present invention provides a medicine for preventing or treating calcific aortic valve disease, wherein the medicine contains chipericumin D.

[0014] Furthermore, the drug treats calcific aortic valve disease by targeting EGFR and inhibiting EGFR phosphorylation activation.

[0015] Furthermore, the dosage forms of the drug include oral solution, injection, tablet, pill, dispersant, capsule, dripping pill, granule, suspension, and emulsion.

[0016] Compared with the prior art, the present invention has the following beneficial effects:

[0017] 1. A large-scale screening of a natural product compound library was performed using Dot-blotting, and candidate compounds were further identified using ALP and Alizarin Red staining. Chipericumin D inhibited the osteogenic differentiation of aortic valve interstitial cells (VICs) by ALP and Alizarin Red staining, Western blotting, and immunofluorescence. Network pharmacology, molecular docking, DARTS, CETSA, SPR analysis, and rescue experiments confirmed that chipericumin D inhibited VIC osteogenic differentiation by targeting EGFR and influencing the downstream PI3K / AKT signaling pathway.

[0018] 2. Treatment of aortic valve interstitial cells with the candidate compound significantly inhibited their osteoblastic differentiation, including decreased expression of the osteoblast-specific genes ALP and RUNX2, and a reduction in the amount of calcium nodules stained with ALP and Alizarin red. Chipericumin D significantly reduced the positive area of ​​calcified nodules stained with ALP and Alizarin red, significantly reduced the expression levels of the osteoblast-specific genes ALP and RUNX2, and significantly reduced the nuclear translocation ratio of the osteoblast-specific transcription factor RUNX2. Chipericumin D inhibits EGFR phosphorylation and activation by targeting EGFR. After inhibiting EGFR phosphorylation and activation, Chipericumin D inhibits the downstream PI3K / AKT signaling pathway. Network pharmacology and molecular docking revealed potential targets for Chipericumin D; DARTS, CETSA, SPR analysis, and rescue experiments confirmed that Chipericumin D targets EGFR.

[0019] 3. This study significantly accelerates drug screening and development, enabling rapid drug screening and clinical application in calcified aortic valve disease (CAVD). Low doses of Chipericumin D can inhibit VIC osteoblastic differentiation, providing a new option for the clinical treatment of CAVD. This study clarifies the role of the EGFR / PI3K / AKT axis in CAVD, providing a theoretical basis for subsequent drug development. EGFR can be used as an independent target for the development of other anti-tumor and osteoblastic differentiation inhibitory therapies. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Figures 5 and 6 are the results of the initial screening experiment in Example 1, wherein AC are experimental results of verifying the expression level of the osteogenic differentiation-specific gene RUNX2 in VIC at 1, 3, and 5 days of OM induction by Dot-blotting and Western blotting, wherein B and C are statistical graphs of RUNX2 expression; D is a flow chart of drug screening by Dot-blotting; E is a Dot-blotting screening result graph, including a graph of RUNX2 and β-actin expression; F is a statistical heat map of the results based on Figure E; and G is a statistical scatter plot of the results based on Figure E.

[0021] Figure 2 Figures 5 and 6 are secondary screening graphs in Example 1, wherein AC are graphs showing the anti-calcification effects of the top 10 candidate compounds screened by Alizarin Red staining and ALP staining, wherein B is a statistical graph showing Alizarin Red staining, and C is a statistical graph showing ALP staining; DF are graphs showing the inhibition of osteogenesis-specific genes ALP and RUNX2 expression by the top 10 candidate compounds screened by Western blotting, wherein E is a statistical graph showing ALP protein expression, and F is a statistical graph showing RUNX2 protein expression; and G is the structural formula of Chipericumin D.

[0022] Figure 3 The results of the in vitro experiment of Chipericumin D inhibiting osteoblastic differentiation of aortic valve interstitial cells in Example 2 are shown in Figure A, wherein A is the result of the CCK8 experiment, showing the IC 5027.57 μM; BD are the experimental results of the concentration gradient experiment of chipericumin D inhibiting VIC osteogenic differentiation-specific genes ALP and RUNX2 verified by Western blotting, showing that the inhibitory effect can be achieved at 3 μM, where C is the statistical graph of ALP protein expression, and D is the statistical graph of RUNX2 protein expression; EF are the experimental results of chipericumin D inhibiting VIC osteogenic differentiation-specific genes ALP and RUNX2 transcriptional expression verified by PCR; GI are the experimental results of chipericumin D inhibiting VIC osteogenic differentiation-specific genes ALP and RUNX2 verified by Western blotting, where H is the statistical graph of ALP protein expression, and I is the statistical graph of RUNX2 protein expression; JK are the experimental results of chipericumin D inhibiting the nuclear entry ratio of VIC osteogenic differentiation-specific genes RUNX2 verified by immunofluorescence staining, where J is the statistical graph; LM are the experimental results of chipericumin D inhibiting late calcification verified by alizarin red staining, where M is the statistical graph; NO is the experimental results of chipericumin D inhibiting VIC osteogenic differentiation-specific genes ALP and RUNX2 verified by ALP staining. D is the result of the early calcification inhibition experiment, where N is the statistical graph;

[0023] Figure 4 A is a Venn diagram for predicting potential targets of CAVD and chipericumin D by network pharmacology in Example 3; Figure 4 B is the network diagram of the common targets of CAVD and Chipericumin D in Example 3;

[0024] Figure 5 This is a diagram of the shared target interaction between CAVD and Chipericumin D in Example 3;

[0025] Figure 6 In Example 3, based on Figure 5 Molecular docking diagram of Chipericumin D and EGFR;

[0026] Figure 7 In Example 3, based on Figure 5 Molecular docking diagram of Chipericumin D and NF-KB1;

[0027] Figure 8 In Example 3, based on Figure 5 Molecular docking diagram of Chipericumin D and TLR4;

[0028] Figure 9 In Example 3, based on Figure 5 Molecular docking diagram of Chipericumin D and STAT3;

[0029] Figure 10 In Example 3, based on Figure 5 Molecular docking diagram of Chipericumin D and HIF1A;

[0030] Figure 11 The figure shows the results of the DARTS experiment in Example 3, verifying the binding of chipericumin D to the top five potential targets, showing that chipericumin D binds to the target EGFR, with the statistical bar graph on the right;

[0031] Figure 12 The results of the CETSA assay for verifying the binding of Chipericumin D to the target EGFR in Example 3 are shown in the figure, wherein the right side of the figure is a statistical scatter plot;

[0032] Figure 13 The SPR analysis results of Chipericumin D and target EGFR in Example 3, K D The value is 5.11 μM;

[0033] Figure 14 Figure 4 is a result diagram showing that chipericumin D inhibits osteogenic differentiation of aortic valve interstitial cells by inhibiting EGFR phosphorylation in Example 4; wherein, AF is a graph showing the experimental results of whether chipericumin D affects the EGFR / PI3K / AKT signaling pathway in the presence of the EGFR agonist NSC228155 verified by Western blotting, wherein B is a statistical histogram of the p-EGFR / EGFR ratio, C is a statistical histogram of the p-PI3K / PI3K ratio, D is a statistical histogram of the p-AKT / AKT ratio, E is a statistical histogram of ALP expression, and F is a statistical histogram of RUNX2 expression; GI is a graph showing whether chipericumin D affects the EGFR / PI3K / AKT signaling pathway and thus affects calcification in the presence of the EGFR agonist NSC228155 and the AKT agonist SC79 verified by alizarin red staining and ALP staining, wherein H is a statistical histogram of alizarin red staining, and I is a statistical histogram of ALP staining;

[0034] Figure 15 These are the in vivo experimental results of the chipericumin D treatment of the mouse calcific aortic valve disease (CAVD) model in Example 5. AB are the observations of the thickness of the mouse aortic valve by HE staining under different treatment groups, wherein A is a representative image of the mouse aortic valve by HE staining, and B is a statistical graph of the thickness of the mouse aortic valve; CD are the observations of the calcification area of ​​the mouse aortic valve by Alizarin red staining under different treatment groups, wherein C is a representative image of the mouse aortic valve by Alizarin red staining, and D is a statistical graph of the calcification area of ​​the mouse aortic valve by Alizarin red staining. DETAILED DESCRIPTION

[0035] Example 1

[0036] Drug screening and Chipericumin D target validation program

[0037] The differences in Dot-blotting, ALP staining, and Alizarin red staining between the compound intervention group and the positive control group under osteogenic differentiation medium (OM) culture conditions were compared to determine the best candidate compound for inhibiting osteogenic differentiation of aortic valve interstitial cells (VICs), such as Figure 1 、 2 shown.

[0038] 1. Initial screening: The top 10 candidate compounds that inhibit the osteogenic differentiation of aortic valve interstitial cells (VICs) were preliminarily screened from a self-built natural product library of 88 compounds using the Dot-blotting method. The specific process is as follows:

[0039] The treated 96-well plate cell lysate was "spotted" onto the nitrocellulose membrane using a pipette, dried at 37°C for 10 minutes, and then blocked with 5% skim milk at room temperature for 1 hour. Finally, the membrane was incubated with antibodies overnight and developed the next day after incubation with the corresponding species secondary antibody at room temperature for 1 hour.

[0040] 2. Secondary screening: Alizarin red staining, ALP staining, and Western blotting were performed using the top 10 candidate compounds from the initial screening. Commercial Alizarin red staining and ALP staining were used to assess the degree of calcification after compound intervention.

[0041] 3. Grouping and compound intervention:

[0042] Negative control group: aortic valve interstitial cells (VIC) were cultured in 2% FBS high glucose medium only;

[0043] Positive control group: VIC cultured in osteogenic differentiation medium (OM);

[0044] Compound intervention group: VICs were treated with compounds under OM culture conditions;

[0045] Detection indicators:

[0046] Dot-blotting: evaluate the expression changes of the osteogenic-specific gene RUNX2;

[0047] ALP staining: evaluate the degree of early calcification;

[0048] Alizarin red staining: to evaluate the content of late calcium nodules;

[0049] Experimental results:

[0050] Dot-blotting:

[0051] Negative control group: low expression of the osteogenic-specific gene RUNX2;

[0052] Positive control group: high expression of the osteogenic-specific gene RUNX2;

[0053] Compound intervention group: The top 10 candidate compounds reduced the expression of the osteoblast-specific gene RUNX2;

[0054] ②ALP staining, Alizarin red staining:

[0055] Negative control group: negative ALP staining and alizarin red staining, with little or no calcium nodules;

[0056] Positive control group: ALP staining and alizarin red staining were positive, with more calcium nodules;

[0057] Compound intervention group: ALP staining and Alizarin red staining were positive after Chipericumin D intervention, but significantly lower than those in the positive control group.

[0058] in conclusion:

[0059] The feasibility of large-scale drug screening using Dot-blotting was clarified; through two composite screenings, the natural product Chipericumin D was identified as a candidate compound for inhibiting the osteoblastic differentiation of aortic valve interstitial cells.

[0060] Among them, the molecular weight of Chipericumin D is 460.6; the molecular formula is: C 27 H 40 O6; PubChem CID: 57381512; structural formula Figure 2 As shown in G.

[0061] Chipericumin D's targeting mechanism:

[0062] Targeting EGFR: Chipericumin D binds to the EGFR protein, forming a hydrogen bond with its Asp-800 amino acid site, inhibiting EGFR phosphorylation;

[0063] Downregulate the EGFR / PI3K / AKT signaling pathway: Chipericumin D inhibits EGFR phosphorylation, thereby inhibiting downstream PI3K / AKT phosphorylation activation, reducing the expression of osteoblast-specific genes ALP and RUNX2, and reducing calcium nodule deposition.

[0064] Example 2

[0065] Chipericumin D inhibits osteoblastic differentiation of aortic valve interstitial cells in vitro

[0066] Chipericumin D was verified to inhibit the osteoblastic differentiation of aortic valve interstitial cells in vitro, and the expression of related genes was detected by qPCR, Western blotting, ALP staining, Alizarin red staining, and immunofluorescence staining.

[0067] 1. Cell culture: Primary valvular interstitial cells were isolated from aortic valves obtained from patients with dilated cardiomyopathy and cultured in DMEM medium containing 10% FBS.

[0068] 2. Osteogenic differentiation induction: Aortic valve interstitial cells are divided into 3 groups:

[0069] Negative control group: aortic valve interstitial cells (VIC) were cultured in 2% FBS high glucose medium only;

[0070] Positive control group: VIC were cultured in osteogenic differentiation medium (OM) to induce osteogenic differentiation;

[0071] Chipericumin D intervention group: VICs were treated with chipericumin D under OM culture conditions;

[0072] 3. Sample collection:

[0073] Cells treated in step 2 were collected and RNA was extracted using an RNA extraction kit (TIANGEN, A0508A) according to the manufacturer's instructions. RNA was reverse-transcribed into cDNA using a reverse transcriptase mix (Vazyme, R323-01). qRT-PCR was then performed using SYBR Green (Vazyme, 7E782J3). VIC cells were lysed using RIPA cell lysis buffer and sonicated for 10 cycles at 8% power for 1 second, 1 second pause, and then centrifuged at 12,000 rpm for 15 minutes. The supernatant was aspirated, 5x loading buffer was added, and the cells were boiled at 95°C for 5 minutes to denature the proteins. RNA and protein were extracted. Cells were fixed with 4% paraformaldehyde for 10 minutes and then stained with ALP, Alizarin Red, and immunofluorescence.

[0074] 4. qPCR detection: SYBR Green assay was used to detect the mRNA expression of osteogenic-specific genes (ALP, RUNX2);

[0075] 5. Western blot: detect the expression levels of ALP and RUNX2 proteins;

[0076] 6. ALP staining: evaluate the degree of early calcification;

[0077] 7. Alizarin red staining: evaluate the content of late calcium nodules;

[0078] 8. Immunofluorescence staining: evaluate the nuclear translocation of the osteogenic-specific gene RUNX2 transcription factor;

[0079] The results are as follows Figure 3 As shown:

[0080] The mRNA expression of bone-specific genes ALP and RUNX2 in the chipericumin D intervention group was significantly decreased; the protein expression levels of bone-specific genes ALP and RUNX2 in the chipericumin D intervention group were significantly decreased; the ALP-positive area in the chipericumin D intervention group was significantly reduced; the content of calcium nodules stained with alizarin red in the chipericumin D intervention group was significantly reduced; and the nuclear entry of the transcription factor of the bone-specific gene RUNX2 in the chipericumin D intervention group was significantly reduced.

[0081] It shows that Chipericumin D can significantly inhibit the osteogenic differentiation of aortic valve interstitial cells.

[0082] Example 3

[0083] Validation of potential targets for Chipericumin D in inhibiting osteogenic differentiation of aortic valve interstitial cells

[0084] The potential targets of Chipericumin D in inhibiting the osteogenic differentiation of aortic valve interstitial cells were identified through network pharmacology, molecular docking, DARTS, CETSA, and SPR.

[0085] 1. Network pharmacology: Potential targets of chipericumin D were predicted using the SEA, Swiss Target Prediction, and super-prep databases, and potential targets for calcific aortic valve disease (CAVD) were predicted using the DisGeNET, GeneCards, and NCBI databases. Protein-protein interaction network analysis was performed on the String database after intersecting chipericumin D targets with CAVD targets, identifying five core targets.

[0086] 2. Molecular docking: Molecular docking was performed on the five core targets and chipericumin D to predict the binding energy;

[0087] 3. DARTS experiment: Chipericumin D (1000 μM) was incubated with VIC cell lysate (1 μg / μl, 40 μl) at room temperature for 1 hour. Pronase E was then added (the protein mass ratio of Pronase E to VIC cell lysate was 1:200) and lysed at room temperature for 5 minutes. 5x Loading buffer (10 μl) was then added and the protein was denatured at 95°C for 5 minutes. Finally, the binding of chipericumin D to the target was verified by Western blotting.

[0088] 4. CETSA assay: After VIC cells were treated with chipericumin D (3 μM) for 24 hours, the VIC cell lysate was divided into two groups. Each group was heated at 42-67°C (5°C interval) for 5 minutes, followed by centrifugation at 4°C, 12,000 rpm / min for 15 minutes. The supernatant was collected, 5x loading buffer was added, and the protein was denatured at 95°C for 5 minutes. Finally, the binding of chipericumin D to the target EGFR was verified by Western blotting.

[0089] 5. SPR analysis: SPR binding analysis can analyze the binding of Chipericumin D to the target EGFR;

[0090] 6. Detection indicators: Western blotting was used to detect the expression of core target proteins.

[0091] The results are as follows Figure 4-13 As shown:

[0092] Network pharmacology analysis showed that the core targets included EGFR, NF-KB1, TLR4, STAT3, and HIF1A; molecular docking showed that the binding energies of chipericumin D with EGFR, NF-KB1, TLR4, and STAT3 were all less than -6 kcal / mol; DARTS experimental results showed that chipericumin D bound to EGFR; CETSA experimental results further verified the binding of chipericumin D to EGFR; SPR analysis revealed that chipericumin D directly bound to EGFR, with a binding energy of 5.11 μM.

[0093] These results indicate that Chipericumin D directly binds to EGFR.

[0094] Example 4

[0095] Chipericumin D inhibits osteogenic differentiation of aortic valve interstitial cells by inhibiting EGFR phosphorylation

[0096] Rescue experiments further verified that Chipericumin D targeted and bound to EGFR, inhibiting its phosphorylation and downstream PI3K / AKT signaling pathway, thereby inhibiting the osteogenic differentiation of aortic valve interstitial cells.

[0097] Negative control group: aortic valve interstitial cells (VIC) were cultured in 2% FBS high glucose medium only;

[0098] Positive control group: VIC were cultured in osteogenic differentiation medium (OM) to induce osteogenic differentiation;

[0099] Chipericumin D intervention group: VICs were treated with chipericumin D (3 μM) under OM culture conditions;

[0100] NSC 228155 intervention group: VIC were treated with NSC 228155 (100 nM) under OM culture conditions;

[0101] NSC 228155 and Chipericumin D intervention group: VICs were co-treated with NSC 228155 (100 nM) and Chipericumin D (3 μM) under OM culture conditions;

[0102] SC79 intervention group: VICs were treated with SC79 (5 μM) under OM culture conditions;

[0103] SC79 and Chipericumin D intervention group: VICs were co-treated with SC79 (5 μM) and Chipericumin D (3 μM) under OM culture conditions.

[0104] The results are as follows Figure 14 As shown:

[0105] In the positive control group, ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly higher than those in the negative control group;

[0106] Chipericumin D intervention group: ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly lower than those in the positive control group;

[0107] In the NSC 228155 intervention group, ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly higher than those in the positive control group;

[0108] In the NSC 228155 and Chipericumin D intervention groups, ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly lower than those in the NSC 228155 intervention group;

[0109] In the SC79 intervention group, ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly higher than those in the positive control group;

[0110] In the SC79 and Chipericumin D intervention groups, ALP, RUNX2, p-EGFR / EGFR ratio, p-PI3K / PI3K ratio, and p-AKT / AKT ratio were significantly lower than those in the SC79 intervention group.

[0111] It shows that Chipericumin D targets and binds to EGFR, inhibiting its phosphorylation and downstream PI3K / AKT signaling pathway, thereby inhibiting the osteogenic differentiation of aortic valve interstitial cells.

[0112] Example 5

[0113] In vivo study of Chipericumin D in treating calcific aortic valve disease (CAVD) mouse model

[0114] Through ApoE - / - Knockout mice (starting from 8 weeks old, divided into negative control group, positive control group, and chipericumin D treatment group; 12 mice in each group) were fed a high-fat diet (HFD, containing 0.25% cholesterol) to establish a CAVD disease model. At the same time, chipericumin D was injected intraperitoneally for in vivo drug treatment to verify whether chipericumin D can treat calcific aortic valve disease (CAVD) in mice.

[0115] Negative control group: Wild-type c57 mice were fed a normal diet (ND) and intraperitoneally injected with DMSO (0.1 mg / kg) every 3 days, without intraperitoneal injection of chipericumin D.

[0116] Positive control group: ApoE - / - Mice were fed a high-fat diet (HFD, containing 0.25% cholesterol) and intraperitoneally injected with DMSO (0.1 mg / kg) every 3 days. Chipericumin D was not injected intraperitoneally.

[0117] Chipericumin D spraying group: ApoE - / -Mice were fed a high-fat diet (HFD, containing 0.25% cholesterol) and intraperitoneally injected with chipericumin D (0.1 mg / kg) diluted in DMSO solution every 3 days;

[0118] The above process lasts for 6 months.

[0119] The results are as follows Figure 15 As shown:

[0120] Negative control group: HE and Alizarin red staining of mouse aortic valve sections showed that the aortic valve was thin and no obvious calcified nodules were observed.

[0121] Positive control group: HE and Alizarin red staining of mouse aortic valve sections showed that the aortic valve was significantly thickened, with obvious calcified nodules and an increased calcified area.

[0122] Chipericumin D treatment group: HE and Alizarin red staining of mouse aortic valve sections showed that the aortic valve thickness was thinner than that of the positive control group, with a small number of calcified nodules and a smaller calcification area.

[0123] It was shown that Chipericumin D can treat calcific aortic valve disease (CAVD) in mice in vivo.

[0124] The present invention solves the current problem of lack of large-scale drug screening for osteoblastic differentiation of aortic valve interstitial cells (VIC) through Dot-blotting large-scale drug screening, and identifies the candidate compound Chipericumin D that inhibits VIC osteoblastic differentiation, which preliminarily solves the core problem of the current lack of effective therapeutic drugs for calcific aortic valve disease (CAVD).

[0125] Experimental data show that low-dose Chipericumin D significantly inhibits VIC osteoblastic differentiation by targeting EGFR and inhibiting its phosphorylation and downstream PI3K / AKT signaling pathway activation. This comprehensive strategy not only provides a new drug screening method for calcific aortic valve disease, but also provides a new strategy for the treatment of other vascular diseases.

[0126] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art shall fall within the scope of protection defined by the claims of the present invention.

Claims

1. Application of Chipericumin D in the preparation of a drug for preventing or treating calcific aortic valve disease.

2. The use according to claim 1, characterized in that The drug can reduce the expression of osteogenesis-related genes ALP and RUNX2.

3. The use according to claim 1, characterized in that The chipericumin D is used to prepare a drug for inhibiting osteogenic differentiation of VIC.

4. The use according to claim 1, characterized in that Chipericumin D is the only active ingredient in the drug.

5. A drug for preventing or treating calcific aortic valve disease, characterized in that: The medicine contains Chipericumin D.

6. The drug according to claim 5, characterized in that The drug treats calcific aortic valve disease by targeting EGFR and inhibiting EGFR phosphorylation activation.

7. The drug according to claim 5, characterized in that The dosage forms of the drug include oral solution, injection, tablet, pill, dispersant, capsule, dripping pill, granule, suspension and emulsion.

Images