Use of inhibitor np-g2-044 in the manufacture of a medicament for breast cancer

Abstract

This invention relates to the application of the inhibitor NP-G2-044 in the preparation of breast cancer drugs, belonging to the field of biomedical technology. It aims to address the problems of drug resistance and significant toxic side effects in existing breast cancer treatments. This invention provides the application of NPG2044 in inhibiting breast cancer cell proliferation and regulating apoptosis, and constructs a combination therapy system of NPG2044 and doxorubicin. Cell proliferation inhibition rate was detected by CCK8 assay, and the expression of apoptosis-related proteins Bax, cleaved PARP, and Bcl2 was detected by Western blot. NPG2044 can specifically inhibit breast cancer cell proliferation and induce apoptosis. Its combination with doxorubicin exhibits a synergistic effect, reducing the single-drug dosage and toxic side effects, providing a new target and combination therapy strategy for breast cancer treatment.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology and relates to the application of the inhibitor NP-G2-044 in the preparation of breast cancer drugs. Background Technology

[0002] Breast cancer is the leading cause of cancer-related death among women worldwide. It is characterized by insidious onset, high malignancy, strong invasiveness, high recurrence and metastasis rates, and treatment resistance, resulting in poor long-term prognosis for patients. Uncontrolled proliferation, apoptosis tolerance, enhanced invasiveness and metastasis, and immunosuppression in the tumor microenvironment are the core pathological reasons for clinical treatment failure and disease recurrence. Currently, routine clinical treatments include surgery, chemotherapy, radiotherapy, endocrine therapy, and targeted therapy. However, these treatments generally suffer from high rates of drug resistance, significant side effects, limited applicability, and difficulty in eliminating minimal residual disease, failing to fundamentally inhibit tumor progression and recurrence. Clinical treatment needs remain unmet. Therefore, developing novel small-molecule drugs with high efficacy, low toxicity, and a clear mechanism of action, possessing both anti-tumor cell proliferation, apoptosis-inducing, and anti-metastatic activities, has significant clinical value and application prospects for overcoming the bottlenecks in breast cancer treatment.

[0003] NP G2 044 is a first-in-class oral small molecule compound developed by Novita Pharmaceuticals, a highly selective inhibitor targeting Fascin (FSCN1). This compound specifically binds to the Fascin protein, blocking its actin-binding activity and inhibiting the formation of filopodia and lamellar pseudopodia in tumor cells, thereby significantly inhibiting tumor cell migration and invasion. Fascin is highly expressed in various tumors, including breast cancer, lung cancer, pancreatic cancer, and ovarian cancer. Its expression level is closely related to poor tumor prognosis, metastasis risk, and treatment resistance. Furthermore, it is expressed at low levels or not at all in normal epithelial tissues, making it an ideal tumor-specific therapeutic target. Existing research largely focuses on NP... G2 The anti-metastasis and immunomodulatory effects of 044 are still in the early stages of research.

[0004] Doxrubicin is a classic first-line chemotherapy drug for breast cancer, widely used in neoadjuvant chemotherapy, adjuvant chemotherapy, and palliative care for advanced stages. It can effectively inhibit tumor cell proliferation and shrink tumor lesions. However, long-term clinical use can easily lead to primary and acquired drug resistance, resulting in decreased efficacy or even treatment failure. It is also accompanied by adverse reactions such as cardiotoxicity and bone marrow suppression, which severely limits its clinical dosage and course of treatment. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide the application of the inhibitor NP-G2-044 in the preparation of breast cancer drugs.

[0006] To achieve the above objectives, the present invention provides the following technical solution: This invention provides the application of the inhibitor NP-G2-044 in the preparation of breast cancer drugs, and the structural formula of the inhibitor NP-G2-044 is shown below: ; The breast cancer described is ER-positive breast cancer; Preferably, the concentration of the inhibitor NP-G2-044 is 1-40 μM; Furthermore, NP G2 044 Application of combination therapy with doxorubicin in the preparation of ER-positive breast cancer drugs; Preferably, the concentration of the inhibitor NP-G2-044 is 20 μM, and the concentration of doxorubicin is 0.1 μM. The present invention also provides a pharmaceutical composition for treating ER-positive breast cancer, the pharmaceutical composition comprising the inhibitor NP-G2-044 and doxorubicin, the structural formula of the inhibitor NP-G2-044 being shown below: ; Preferably, the pharmaceutical composition further includes a pharmaceutically acceptable carrier and excipients; Preferably, the pharmaceutical composition is an oral or injectable formulation.

[0007] The beneficial effects of this invention are as follows: 1. Fills a research gap and demonstrates outstanding innovation. Existing research has only confirmed the efficacy of NP-G2-044 in triple-negative breast cancer, and there are no reports on its application in MCF-7 hormone-dependent breast cancer. At the same time, this invention is the first to propose the combination of NP-G2-044 and doxorubicin, opening up a new direction for the treatment of hormone-dependent breast cancer with targeted drugs combined with chemotherapy drugs, and the research is significantly innovative.

[0008] 2. Complementary mechanisms of action, synergistically enhancing overall efficacy. The two drugs have different mechanisms of action and functional advantages, and their combined use achieves complementary benefits. Doxorubicin strongly inhibits cell proliferation and induces apoptosis, while NP-G2-044 exerts anti-migration and colony formation effects. The combined use of these drugs significantly outperforms the single-drug therapy in all aspects, and can comprehensively intervene in malignant behaviors such as tumor proliferation, apoptosis, migration, and long-term tumorigenesis.

[0009] 3. Reduce medication dosage and alleviate drug toxicity and side effects. By leveraging the synergistic effect of combined use, the dosage of doxorubicin can be reduced while ensuring the same anti-tumor efficacy, effectively reducing adverse reactions such as cardiotoxicity and bone marrow suppression caused by the drug, and significantly improving patient tolerance and treatment safety.

[0010] 4. The experimental system is comprehensive, and the results are true and reliable. This invention employs a variety of classic in vitro pharmacodynamic evaluation techniques, such as CCK-8, Western blot, scratch assay, and colony formation assay, to complete the verification from multiple dimensions, including cell function and molecular proteins. The experimental design is standardized, the data has good reproducibility, and the experimental results can serve as a reliable basis for subsequent basic research and clinical translation.

[0011] 5. The finished drug has a promising future and a wide range of applications. This pharmaceutical composition has a simple formulation and a mature preparation process. The two active ingredients are chemically stable, facilitating industrial production and formulation development. This combination can be used as a standalone treatment or in conjunction with other treatments depending on the patient's condition, thus adapting to the individualized treatment needs of different patients and demonstrating high clinical translational value.

[0012] Other advantages, objectives, and features of the invention will be set forth in part in the description which follows, and in part will be apparent to those skilled in the art from the following examination, or may be learned from practice of the invention. The objectives and other advantages of the invention can be realized and obtained through the following description. Attached Figure Description

[0013] To make the objectives, technical solutions, and advantages of the present invention clearer, the preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings, wherein: Figure 1 Gradient concentration NP G2 044 Effect on the proliferation activity of breast cancer cells MCF-7.

[0014] Figure 2 For different concentrations of NP G2 044. Regulation of the expression of apoptosis-related proteins in MCF-7 breast cancer cells.

[0015] Figure 3 NP G2 044, when used in combination with doxorubicin, has a synergistic effect in inhibiting breast cancer cell proliferation and promoting apoptosis.

[0016] Figure 4 For different concentrations of NP G2 Effect of 044 on the migration ability of MCF-7 breast cancer cells.

[0017] Figure 5 NP G2 Effects of 044 combined with doxorubicin on the migration ability of MCF-7 cells. Detailed Implementation

[0018] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that the illustrations provided in the following embodiments are only schematic representations of the basic concept of the present invention. Unless otherwise specified, the following embodiments and features can be combined with each other.

[0019] The accompanying drawings are for illustrative purposes only and are schematic diagrams, not actual pictures. They should not be construed as limiting the invention. To better illustrate the embodiments of the invention, some parts in the drawings may be omitted, enlarged, or reduced, and do not represent the actual product dimensions. It is understandable to those skilled in the art that some well-known structures and their descriptions may be omitted in the drawings.

[0020] In the accompanying drawings of the embodiments of the present invention, the same or similar reference numerals correspond to the same or similar components. In the description of the present invention, it should be understood that if terms such as "upper," "lower," "left," "right," "front," and "rear" indicate the orientation or positional relationship based on the orientation or positional relationship shown in the drawings, they are only for the convenience of describing the present invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, the terms used to describe positional relationships in the drawings are only for illustrative purposes and should not be construed as limiting the present invention. For those skilled in the art, the specific meaning of the above terms can be understood according to the specific circumstances.

[0021] Example 1: Different concentrations of NP G2 044 Effects on breast cancer cell proliferation This embodiment uses the human breast cancer cell line MCF7 as an in vitro research model. This cell line can stably maintain the malignant biological phenotype of breast cancer and is suitable for in vitro efficacy evaluation of small molecule compounds. The cells are prepared using a solution containing 10% fetal bovine serum and 1% penicillin. Streptomycin-containing DMEM complete medium was routinely passaged in a constant temperature incubator at 37 ℃, 5% CO2, and saturated humidity. Cells in the logarithmic growth phase and in good growth condition were selected for experiments.

[0022] Breast cancer cells were uniformly seeded onto cell culture plates and randomly divided into a blank control group and groups receiving different concentrations of NP. G2 044 (0, 2, 4, 6, 8, 16, 32 μM) treatment groups. The blank control group was cultured in an equal volume of drug-free complete culture medium. Each drug group was treated with a gradient concentration of NP. G2 044. Continuous intervention culture with working solution. After the drug treatment ends, CCK is used. 8. Cell proliferation assay kit was used to detect the cell proliferation activity of each group. The assay kit was followed according to the instructions. The absorbance at the corresponding wavelength was measured using an ELISA reader. The cell viability and proliferation inhibition rate of each group were calculated.

[0023] Experimental results show (e.g.) Figure 1 As shown), NP G2 044 significantly inhibited the proliferation of breast cancer cells in a concentration-dependent manner; as the drug concentration increased, the survival rate of breast cancer cells gradually decreased, while the inhibitory effect on proliferation continued to increase, confirming that NP G2 044 exhibits clear in vitro anti-breast cancer proliferation activity.

[0024] Example 2NP G2 044 Effects on the expression of apoptosis-related proteins in breast cancer cells In this embodiment, the aforementioned human breast cancer cells were used for routine culture and drug intervention, and a blank control group and NP were set up. G2 The 044 (0, 10, 20, 40 μM) treatment group was subjected to appropriate culture and drug incubation conditions.

[0025] After drug intervention, cell pellets from each group were collected, and total cell protein was extracted using RIPA protein lysis buffer. Protein quantification was performed using a BCA kit to ensure uniform protein loading across all groups. Following protein denaturation and SDS-PAGE... The standard Western blot procedure, including PAGE electrophoresis, wet transfer, room temperature blocking, overnight incubation with primary antibody at 4°C, and room temperature incubation with secondary antibody, was used to detect the expression levels of key proteins in the apoptosis pathway, including pro-apoptotic proteins Bax and cleaved proteins. PARP and the anti-apoptotic protein Bcl-2.

[0026] Experimental results show (e.g.) Figure 2 As shown), compared with the blank control group, NP G2 The 044 treatment group showed significant upregulation of pro-apoptotic proteins Bax and cleaved proteins. PARP expression was significantly reduced, while the expression of the anti-apoptotic protein MCL1 was also significantly downregulated. This indicates that NP... G2 044 can break tumor cell apoptosis tolerance and induce breast cancer cell apoptosis by regulating the expression of key apoptosis-related proteins, thus clarifying its molecular mechanism of action.

[0027] Example 3NP G2 044 Synergistic effect of combined use with doxorubicin Using NP-G2-044 as the core, supplemented with pharmaceutically acceptable carriers and excipients (such as lactose, starch, disintegrants, etc.), an anti-breast cancer drug containing NP-G2-044 was prepared according to pharmaceutical preparation specifications. Simultaneously, this drug was combined with doxorubicin (0.1 μM) to construct an "NP-G2-044 + doxorubicin" combination therapy system. This system leverages the synergistic effect of NP-G2-044's targeted regulatory effect and doxorubicin's chemotherapeutic effect to enhance the inhibition of breast cancer cell proliferation and apoptosis induction, reduce the dosage of single drugs, decrease toxic side effects, and improve treatment safety and efficacy. This provides a new drug combination scheme and experimental evidence for clinical combination therapy of breast cancer.

[0028] This embodiment uses the same human breast cancer cell in vitro model. After routine culture to the logarithmic growth phase, the cells are randomly divided into four groups: blank control group, NP... G2 044 Single-drug group, doxorubicin single-drug group, NP G2 044 + Doxorubicin combination therapy group. Each group was treated with the corresponding culture medium, single drug, or combination therapy at a specific concentration.

[0029] After the intervention, CCK was used. 8. Cell proliferation activity in each group was detected using the following method, and the cell proliferation inhibition rate was calculated. Simultaneously, cells from each group were collected, total protein was extracted and quantified, and Bax and cleaved protein were detected by Western blot. Changes in the expression of PARP and Bcl-2 apoptosis-related proteins.

[0030] Experimental results show (e.g.) Figure 3 As shown), with individual NP G2 Compared with the doxorubicin-only group, the combination therapy group showed a significantly enhanced inhibitory effect on breast cancer cell proliferation, exhibiting a clear synergistic effect; furthermore, the combination therapy further upregulated Bax and cleaved cells. PARP protein expression was significantly downregulated, and Bcl2 protein levels were more significantly reduced, resulting in apoptosis induction effects superior to monotherapy. Furthermore, the combination of these two drugs can reduce the dosage of monotherapy while maintaining equivalent tumor-suppressive efficacy, helping to mitigate the toxic side effects of doxorubicin alone. This provides new candidate drug combinations and experimental evidence for combined chemotherapy and synergistic therapy for breast cancer.

[0031] Example 4NP G2 Effect of 044 on the migration ability of MCF-7 cells Logarithmically growing MCF-7 cells were seeded into culture plates and cultured routinely until cell confluence reached 90% or higher. Straight scratches were made in the cell monolayer using a sterile pipette tip. After rinsing with PBS buffer to remove detached cells, the appropriate culture medium and drugs were added, and the cells were cultured further. After the predetermined time, the cells were observed and photographed under a microscope, and the scratch width was measured. The experimental results show (e.g.) Figure 4 As shown in the figure, NP-G2-044 can inhibit cell migration.

[0032] Example 5NP G2 044 Effects of doxorubicin on the migration ability of MCF-7 cells Logarithmically growing MCF-7 cells were seeded into culture plates and cultured routinely until cell confluence reached 90% or higher. Straight scratches were made in the cell monolayer using a sterile pipette tip. After rinsing with PBS buffer to remove detached cells, the cells were divided into a blank control group, an NP-G2-044 single-drug group, a doxorubicin single-drug group, and a combination group. The corresponding culture medium and drugs were added and the cells were cultured for a further period.

[0033] After culturing for the preset time, the cells were observed and photographed under a microscope, the scratch width was measured, and the cell migration rate was calculated. Experimental results showed (e.g.) Figure 5 As shown in the figure, the blank group and the doxorubicin monotherapy group showed strong cell migration ability, while the cell migration behavior of the combination therapy group was significantly inhibited and the scratch healing speed was greatly slowed down, which confirms that the combination therapy can effectively block the invasion and metastasis of tumor cells.

[0034] The above experimental results fully demonstrate that the NP-G2-044 and doxorubicin drug composition provided by this invention, relying on the complementary mechanisms and multiple synergistic effects of the two drugs, comprehensively optimizes the anti-tumor effect and has high clinical translational value.

[0035] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. The application of the inhibitor NP-G2-044 in the preparation of breast cancer drugs, characterized in that: The structural formula of the inhibitor NP-G2-044 is shown below: ； The breast cancer mentioned is ER-positive breast cancer.

2. The application according to claim 1, characterized in that: The concentration of the inhibitor NP-G2-044 is 1-40 μM. 3.NP G2 044 Application of combination therapy with doxorubicin in the preparation of ER-positive breast cancer drugs.

4. The application according to claim 3, characterized in that: The concentration of the inhibitor NP-G2-044 was 20 μM, and the concentration of doxorubicin was 0.1 μM.

5. A pharmaceutical composition for treating ER-positive breast cancer, characterized in that: The pharmaceutical composition comprises the inhibitor NP-G2-044 and doxorubicin, and the structural formula of the inhibitor NP-G2-044 is shown below: 。 6. The pharmaceutical composition according to claim 5, characterized in that: The pharmaceutical composition also includes pharmaceutically acceptable carriers and excipients.

7. The pharmaceutical composition according to claim 6, characterized in that: The pharmaceutical composition is an oral or injectable formulation.

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