Application of coptis chinensis active ingredient jatrorrhizine in preparation of medicine for resisting EGFR-TKIs drug-resistant non-small cell lung cancer
Through the intervention of the PI3K-Akt signaling pathway by the active ingredient of Coptis chinensis, the treatment problem of EGFR-TKIs resistant non-small cell lung cancer was solved, and effective inhibition and apoptosis promotion of T790M mutation was achieved, and new ideas for traditional Chinese medicine were provided.
Patent Information
- Application Number
- CN202510654561.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-21
- Publication Date
- 2025-08-12
AI Technical Summary
The prior art is difficult to effectively intervene in non-small cell lung cancers resistant to EGFR-TKIs, especially the resistance caused by T790M mutations, resulting in poor treatment effects and high cost.
The active ingredient of Coptis chinensis was used to intervene in T790M mutant EGFR-TKIs-resistant non-small cell lung cancer through the PI3K-Akt signaling pathway, regulate the activity of amino acid kinases such as tyrosine and serine, inhibit cell growth, migration and invasion, and regulate the PI3K/Akt/mTOR pathway.
Significantly inhibiting the growth, proliferation and invasion of non-small cell lung cancer cells resistant to T790M mutant EGFR-TKIs, improving the apoptosis rate, and reducing the expression of key target proteins PI3K and mTOR, providing a new therapeutic strategy.
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Figure CN120459092A_ABST
Abstract
Description
Technical Field
[0001] The present application relates to the field of biomedicine technology, and in particular to the use of jatrorrhizine, an active ingredient of Coptis chinensis, in the preparation of drugs for treating EGFR-TKIs-resistant non-small cell lung cancer. Background Art
[0002] Approximately 90% of lung cancers are non-small cell lung cancer (NSCLC), which is highly malignant, has a low survival rate, and a poor prognosis. Because early-stage NSCLC patients have no obvious tumor symptoms, by the time they are diagnosed, most have already metastasized. Furthermore, early diagnosis and treatment of NSCLC remain underdeveloped. Consequently, the five-year survival rate for NSCLC patients is less than 15%.
[0003] Currently, targeted biological therapy is recommended as the first-line standard treatment for non-small cell lung cancer (NSCLC). Gefitinib, a first-generation targeted drug and an epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), offers the advantages of minimal side effects and high targeted efficacy in the clinical treatment of NSCLC. However, most patients develop resistance after 10-16 months, leading to disease progression, local recurrence, or distant metastasis. Studies have shown that the primary cause of acquired resistance to gefitinib in NSCLC is the secondary T790M mutation in exon 20 of the EGFR gene. Osimertinib, as a second-line salvage therapy for patients with the EGFR driver gene T790M mutation, is an important option for overcoming resistance to EGFR-TKIs. However, resistance is inevitable after first-line or second-line use of osimertinib, limiting its long-term clinical benefit. Furthermore, osimertinib is relatively expensive.
[0004] Overcoming EGFR-TKI resistance could effectively prolong lung cancer patients' survival and reduce their economic burden. Currently, there is little research and drug development targeting delaying, reversing, or killing resistant tumor cells. Due to the continuous development and evolution of tumors and their selective adaptation to EGFR inhibitors, treatment strategies for patients with resistant non-small cell lung cancer remain an ongoing research challenge. Therefore, there is an urgent need to identify new drugs that target T790M mutation-resistant non-small cell lung cancer, and there is significant potential for developing new drugs against EGFR-TKI-resistant non-small cell lung cancer. Summary of the Invention
[0005] The technical problem to be solved by the present invention is to seek new drugs for intervening in T790M mutation-resistant non-small cell lung cancer, and to provide the application of jatrorrhizine, an active ingredient of Coptis chinensis, in the preparation of drugs against EGFR-TKIs-resistant non-small cell lung cancer.
[0006] The above-mentioned purpose of the present invention is achieved through the following technical solutions:
[0007] The present study shows that jatrorrhizine, an active ingredient in Coptis chinensis, exerts its efficacy against EGFR-TKIs-resistant non-small cell lung cancer by acting on PI3K and through the PI3K-Akt signaling pathway. In vitro cell experiments showed that jatrorrhizine can significantly inhibit the growth, proliferation, migration, and invasion of H1975 cells. As the concentration of jatrorrhizine increases, the apoptosis rate of H1975 cells gradually increases, and the invasive ability of H1975 cells gradually weakens, showing a concentration-dependent effect. At the same time, network pharmacology technology was used to predict the targets of jatrorrhizine on T790M mutation-resistant non-small cell lung cancer. Further pathway enrichment analysis of these effective targets predicted that jatrorrhizine intervenes in T790M mutation-resistant non-small cell lung cancer through the PI3K / Akt / mTOR signaling pathway. Research on the regulatory effect of jatrorrhizine on the PI3K / Akt / mTOR pathway showed that jatrorrhizine can downregulate the phosphorylation of PI3K and mTOR proteins, indicating that jatrorrhizine has the effect of intervening in targeted drug-resistant non-small cell lung cancer, which not only provides a new direction for the treatment of targeted drug-resistant lung cancer, but also provides new ideas for the research of traditional Chinese medicine.
[0008] The jatrorrhizine used in the present invention is a safe source, derived from the traditional Chinese medicine Coptis chinensis. Jatrorrhizine is abundant in the herb, relatively readily available, and inexpensive. Furthermore, it is non-toxic and relatively safe. The CAS number for jatrorrhizine is 3621-38-3, and its molecular structure is shown below:
[0009]
[0010] Therefore, the present invention protects the use of jatrorrhizine in preparing drugs against non-small cell lung cancer.
[0011] Preferably, jatrorrhizine is used in the preparation of a drug for treating EGFR-TKIs-resistant non-small cell lung cancer.
[0012] Furthermore, the drug can inhibit the growth and proliferation of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
[0013] Furthermore, the drug can inhibit the migration of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
[0014] Furthermore, the drug can promote apoptosis of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
[0015] Furthermore, the drug can weaken the invasive ability of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
[0016] Furthermore, the drug can reduce the expression of PI3K and mTOR, key target proteins of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer.
[0017] Furthermore, the drug can regulate the phosphorylation of the PI3K / Akt / mTOR pathway.
[0018] Preferably, the drug further comprises a pharmaceutically acceptable carrier or excipient.
[0019] Preferably, the dosage form of the drug is tablets, granules, injections, powders, capsules or suspensions.
[0020] The present invention has the following beneficial effects:
[0021] The present invention provides a new use of jatrorrhizine, an active ingredient of the traditional Chinese medicine Coptis chinensis, in the treatment of non-small cell lung cancer. It was found that jatrorrhizine can significantly inhibit the growth, proliferation, migration and invasion of H1975 cells, and as the concentration of jatrorrhizine increases, the apoptosis rate of H1975 cells gradually increases, and the invasive ability gradually weakens, which is concentration-dependent. Jatrorrhizine intervenes in T790M mutant EGFR-TKIs-resistant non-small cell lung cancer via the PI3K-Akt signaling pathway, and kills targeted drug-resistant non-small cell lung cancer cells by regulating the activity of amino acid kinases such as tyrosine and serine, regulating the cell cycle, etc. The jatrorrhizine provided by the present invention has the effect of intervening in targeted drug-resistant non-small cell lung cancer, which not only provides a new direction for the treatment of targeted drug-resistant lung cancer, but also provides a new idea for the research of traditional Chinese medicine. BRIEF DESCRIPTION OF THE DRAWINGS
[0022] Figure 1 The diagram shows the cytotoxicity of jatrorrhizine to H1975 cells (A in the figure is 24 hours; B is 48 hours).
[0023] Figure 2 This is the result of jatrorrhizine inhibiting the migration of H1975 cells.
[0024] Figure 3 This is the result of jatrorrhizine-induced apoptosis of H1975 cells.
[0025] Figure 4 This is the result of jatrorrhizine inhibiting the invasion of H1975 cells.
[0026] Figure 5 The effect of jatrorrhizine on the phosphorylation of PI3K and mTOR proteins in H1975 cells (Note: x±s, n=3, compared with the blank control group, *P<0.05, **P<0.01). DETAILED DESCRIPTION
[0027] The present invention will be further described below with reference to the accompanying drawings and specific examples, but the examples do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.
[0028] Unless otherwise specified, the reagents and materials used in the following examples were commercially available.
[0029] The human lung adenocarcinoma H1975 cells used in the examples (T790M mutation EGFR-TKIs resistant non-small cell lung cancer cell line, batch number: CL-0298) were purchased from Wuhan Punosai Life Science Technology Co., Ltd.; jatrorrhizine (mass fraction ≥98%; batch number: 3621-38-3) was purchased from Shanghai Yuanye Biotechnology Co., Ltd.; RPMI-1640 medium (batch number: C11875500BT), fetal bovine serum (batch number: 10099141), and 1% penicillin / streptomycin double antibody (batch number: 15140-122) were all purchased from Gibico, USA; CCK-8 kit (batch number: BS350B) was purchased from Anhui Lanjieke Technology Co., Ltd.
[0030] Example 1 Effect of jatrorrhizine on H1975 cell growth
[0031] Cell culture: H1975 cells were cultured in RPMI-1640 medium containing 10% fetal bovine serum and 1% double-antibody at 37°C and 5% CO2 saturated humidity. Cells were passaged when they reached the logarithmic phase.
[0032] H1975 cells in logarithmic growth phase were collected and 5×10 4 Cells were seeded at a density of 1 μg / well in a 96-well plate and cultured in a 37°C, 5% CO2 incubator for 24 hours. The cells were then divided into four groups according to the drug concentration, with six replicates per group. Different concentrations of jatrorrhizine solution (0, 3.75, 7.5, 15, 30, and 60 μg / mL) were administered according to the grouping. After 24 and 48 hours of treatment, the medium was replaced. 10 μL of CCK-8 solution was added to each well in the dark. After 2 hours of incubation in a cell culture incubator, the absorbance (OD) at 450 nm was measured using a multi-function microplate reader (Biotek, USA, model: Biotek Synergy H1). The experiment was repeated three times.
[0033] The results of the test are as follows Figure 1 As shown in the figure, the cell survival rate of H1975 cells decreased significantly with the increase of jatrorrhizine concentration, and was also significantly negatively correlated with time. The effective concentration at 24 hours and 48 hours was 3.75 μg / mL. The half-maximal inhibitory concentration (IC 50) was 6.434 μg / mL, indicating that jatrorrhizine could significantly inhibit the growth and proliferation of H1975 cells.
[0034] Example 2 Effect of jatrorrhizine on H1975 cell migration
[0035] H1975 cells were evenly seeded at an appropriate density in a 12-well plate. A scratch was made on the surface of the culture dish with a 10 μL pipette tip. The cells were cultured in serum-free jatrorrhizine medium with different concentrations for 24 h and 48 h. The healing of the scratches in each group was observed, and the width of the cell scratches and the number of cells distributed in the scratches were measured. Cells with smaller widths and more cells migrating into the scratches had stronger relative migration ability.
[0036] The results of the test are as follows Figure 2 As shown in the figure, the wound healing area decreased with increasing jatrorrhizine concentration and was positively correlated with time. At the same time, cell morphology was observed and cell density decreased with time. Therefore, it was concluded that jatrorrhizine can inhibit the migration of H1975 cells.
[0037] Example 3 Effect of jatrorrhizine on apoptosis of H1975 cells
[0038] H1975 cells were evenly plated and treated with 0μM, 15μM, 30μM and 60μM jatrorrhizine according to group. After fixation, RNase A and propidium iodide were added sequentially for incubation. The cell cycle of each group was detected by flow cytometry.
[0039] The results of the test are as follows Figure 3 As shown, it was shown that with the increase of jatrorrhizine concentration, the apoptosis rate of H1975 cells gradually increased in a concentration-dependent manner.
[0040] Example 4 Effect of jatrorrhizine on H1975 cell invasion
[0041] H1975 cells were divided into groups and treated with 0 μM, 15 μM, 30 μM and 60 μM jatrorrhizine solutions for 24 h and 48 h, and then a tumor cell invasion experiment was performed using the Transwell invasion chamber assay to determine the invasion ability of H1975 cells.
[0042] The results of the test are as follows Figure 4 As shown, with the increase of jatrorrhizine concentration, the invasion ability of H1975 cells gradually weakened in a concentration-dependent manner, indicating that jatrorrhizine can inhibit the invasion of H1975 cells.
[0043] Example 5 Prediction of Targets of Jatrorrhizine against Non-Small Cell Lung Cancer and Its Effects on Key Targets of H1975 Cells
[0044] 1. Prediction of jatrorrhizine targets against non-small cell lung cancer and construction of PPI network analysis
[0045] The potential targets of jatrorrhizine were compared and analyzed with the genes related to non-small cell lung cancer, and a Venn diagram was drawn. Subsequently, the intersection of the potential targets of jatrorrhizine and the genes related to non-small cell lung cancer was imported into the STRING database (https: / / string-db.org, Version 11.0), and the free target genes were excluded. A PPI network was constructed for visual analysis. Then, the intersection genes were imported into the DAVID database (https: / / david.ncifcrf.gov / , Version 8.0) for GO enrichment analysis and KEGG enrichment analysis. Then, the "KEGG PATHWAY Database" online database (https: / / www.genome.jp / kegg / pathway.html) was used to search for the "EGFR tyrosine kinase inhibitor resistance pathway" (hsa01521 EGFR TYROSINE KINASE INHIBITOR RESISTANCE PATHWAY), and the genes in this pathway were exported to construct a database of genes related to EGFR-TKIs resistance.
[0046] The predicted targets of jatrorrhizine against non-small cell lung cancer were compared and analyzed with the genes related to EGFR-TKIs resistance to determine the targets of jatrorrhizine against EGFR-TKIs-resistant non-small cell lung cancer. The targets of jatrorrhizine against EGFR-TKIs-resistant non-small cell lung cancer were marked on the hsa01521 pathway map through the "KEGG Mapper" database (https: / / www.genome.jp / kegg / mapper / color.html).
[0047] A total of 118 potential targets of jatrorrhizine were predicted through the Swiss Target Prediction database. By integrating and comparing the drug targets with the disease-related genes, a total of 69 targets of jatrorrhizine against non-small cell lung cancer were obtained. Further in the EGFR-TI<Is resistance pathway, 7 targets related to the action of jatrorrhizine were obtained: PIK3CB, PIK3CD, BRAF, Rafl, MAP2Kl, ERBB2, and mTOR. These 7 targets are the targets of jatrorrhizine against ECFR-TKIs-resistant non-small cell lung cancer.
[0048] The molecular simulation docking results between jatrorrhizine and targets showed that the RMSD values of the docking models of jatrorrhizine and the seven target molecules were all less than 2A, that is, the seven EGFR-TKIs resistance-related targets predicted by network pharmacology can all be docked with jatrorrhizine molecules, among which the binding energies of PIK3CB, PIK3CD, mTOR and Rafl for docking with jatrorrhizine are all less than -7kcal / mol. The smaller the binding energy, the better the binding effect of molecular docking. If it is lower than -7kcal / mol, there is a strong affinity between the two. It can be seen that PIK3CB, PIK3CD, mTOR and Raf1 all have strong affinity with jatrorrhizine molecules. Among them, PIK3CB, PIK3CD and mTOR can form hydrogen bonds with jatrorrhizine molecules, indicating that their binding has good stability. The KEGG enrichment results show that the PI3K-Akt signaling pathway is an important pathway with highly enriched targets for jatrorrhizine in the treatment of NSCLC. PIK3CB and PIK3CD are subunits of PI3K. PI3K and mTOR are both key proteins in the PI3K-Akt signaling pathway. Therefore, we selected PI3KCB, PIK3CD and mTOR as the key targets of jatrorrhizine against EGFR-TKIs-resistant NSCLC.
[0049] 2. Western blotting to detect the effect of jatrorrhizine on the expression of key targets in H1975 cells
[0050] H1975 cells were evenly plated and treated with 0μM, 15μM, 30μM and 60μM jatrorrhizine according to group. The expression of PI3K, pPI3K, pmTOR and mTOR proteins was detected by Western Blot.
[0051] Western blot was further used to determine the expression and phosphorylation of key target proteins in H1975 cells after treatment with different concentrations of jatrorrhizine. Figure 5 As shown, it was shown that jatrorrhizine could downregulate the phosphorylation of PI3K and mTOR proteins.
[0052] In summary, this study found that coptis chinensis, through its active ingredient jatrorrhizine, may intervene in EGFR-TKI-resistant non-small cell lung cancer (NSCLC) with the T790M mutation via the PI3K-Akt signaling pathway. This activity modulates the activity of tyrosine and serine amino acid kinases and the cell cycle, leading to the killing of target-resistant NSCLC cells. In vitro cytological experiments revealed that jatrorrhizine, an active ingredient in coptis chinensis, has the potential to inhibit the development of target-resistant NSCLC. This provides new avenues for the treatment of target-resistant lung cancer and new insights into Traditional Chinese Medicine research.
[0053] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.
Claims
1. The use of jatrorrhizine in the preparation of a drug for treating non-small cell lung cancer, characterized in that: The structural formula of jatrorrhizine is:
2. The application according to claim 1, characterized in that Application of jatrorrhizine in the preparation of drugs for treating EGFR-TKIs-resistant non-small cell lung cancer.
3. The application according to claim 2, characterized in that: The drug can inhibit the growth and proliferation of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
4. The application according to claim 2, characterized in that: The drug can inhibit the migration of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
5. The application according to claim 2, characterized in that: The drug can promote apoptosis of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
6. The application according to claim 2, characterized in that: The drug can weaken the invasive ability of T790M mutant EGFR-TKIs-resistant non-small cell lung cancer cells.
7. The use according to claim 2, characterized in that: The drug can reduce the expression of PI3K and mTOR, key target proteins in T790M mutant EGFR-TKIs-resistant non-small cell lung cancer.
8. The application according to claim 7, characterized in that: The drug can regulate the phosphorylation of the PI3K / Akt / mTOR pathway.
9. The use according to any one of claims 1 to 8, characterized in that: The medicine further contains a pharmaceutically acceptable carrier or excipient.
10. The use according to any one of claims 1 to 8, characterized in that: The dosage form of the medicine is tablet, granule, injection, powder, capsule or suspension.