Application of EGFR inhibitor and PARP inhibitor in treatment of ovarian cancer

Abstract

The invention relates to application of an EGFR (epidermal growth factor receptor) inhibitor and a PARP (poly-ADP-ribose polymerase) inhibitor in treatment of ovarian cancer, and relates to the technical field of ovarian cancer treatment, an EGFR inhibitor lapatinib and a PARP inhibitor niraparib are obtained by screening an FDA / CFDA approved compound library, and it is found that lapatinib and niraparib have a good synergistic ovarian cancer resisting effect. In order to further research the molecular mechanism of the synergistic effect of the two, the cell death mode after the combined effect of the two is detected through apoptosis and western blotting. The combination of lapatinib and niraparib can effectively inhibit the growth of ovarian cancer in an ovarian cancer animal model, shows good clinical research and application values, and is expected to become a new tool for treating ovarian cancer.

Description

Technical Field

[0001] The present invention relates to the technical field of ovarian cancer treatment, and in particular to an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer. Background Art

[0002] Ovarian cancer is currently one of the deadliest gynecological cancers in the world and the second most common malignant tumor in women over 40 years old, second only to breast cancer. Compared with other cancers, the tumor tissue sources of ovarian cancer are diverse and complex, which makes clinical diagnosis and staging more difficult. According to the World Health Organization, there will be approximately 310,000 new cases and 210,000 deaths from ovarian cancer in 2020. Due to the difficulty in early diagnosis of ovarian cancer, more than 70% of patients are diagnosed in the late stage, and the recurrence rate after surgery and radiotherapy is still about 70%, resulting in a 5-year survival rate of less than 30%. Once the platinum-resistant phenotype appears, patients will be treated with other chemotherapy drugs, such as gemcitabine and topotecan, but the effect is weak and may even eventually develop multidrug resistance. In addition, the main targeted drugs currently approved by the FDA for widespread use are VEGF inhibitors bevacizumab and PARP inhibitors. Other targets, such as VEGF receptor inhibitors, EGFR tyrosine kinase inhibitors, folate receptor α inhibitors, and immune checkpoint inhibitors, have either been found to be ineffective in some patients, are in the early stages of clinical trials, or have been shown to have significant toxic side effects. Therefore, in response to the current dilemma in ovarian cancer treatment, it is of great significance to develop new therapies to overcome drug resistance. Although the initial response rate to PARP inhibitor treatment is good, and the progression-free survival and overall survival are significantly increased, most cancers will eventually develop drug resistance. The development of new drug combinations is not only conducive to overcoming PARP inhibitor resistance and reducing the toxic side effects of high-dose PARP inhibitors alone, but also helps to further explore the molecular mechanisms associated with the emergence of resistance. Summary of the invention

[0003] In order to solve the above problems, the present invention provides an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer.

[0004] In a first aspect, the present invention provides a use of an EGFR inhibitor combined with a PARP inhibitor in the preparation of a drug for treating ovarian cancer.

[0005] Furthermore, the EGFR inhibitor includes lapatinib, or its tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate.

[0006] Furthermore, the PARP inhibitor includes niraparib, or its tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate.

[0007] In a second aspect, the present invention provides a drug for treating ovarian cancer, wherein the drug for treating ovarian cancer comprises an EGFR inhibitor and a PARP inhibitor.

[0008] Furthermore, the EGFR inhibitor includes lapatinib, or a tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate thereof; the PARP inhibitor includes niraparib, or a tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate thereof.

[0009] Furthermore, the preparation dosage form of the drug for treating ovarian cancer includes oral liquid, capsule, oil drop, powder, tablet or injection.

[0010] Furthermore, the drug for treating ovarian cancer also includes a pharmaceutically acceptable carrier or excipient.

[0011] The above technical solution provided by the present invention has at least the following advantages compared with the prior art:

[0012] The present invention provides an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer. The EGFR inhibitor Lapatinib and the PARP inhibitor Niraparib are obtained by screening the FDA / CFDA approved compound library. The present invention finds that Lapatinib and Niraparib have a good synergistic anti-ovarian cancer effect. In order to further study the molecular mechanism of the synergistic effect of the two, the present invention also detects the cell death pattern after the combined action of the two by apoptosis and protein immunoblotting. Lapatinib combined with Niraparib effectively inhibits the growth of ovarian cancer in an ovarian cancer animal model, showing good clinical research and application value, and is expected to become a new tool for the treatment of ovarian cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the invention and, together with the description, serve to explain the principles of the invention.

[0014] In order to more clearly illustrate the technical solutions in the present invention or the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative labor.

[0015] Figure 1 Figure 3 is a diagram showing that lapatinib is identified as a target drug for combination testing with niraparib in the present invention; Figure 1Middle: (A) Screening scheme; Cell viability of niraparib and non-niraparib drugs in SKOV3 (B) and ES-2 cells (C); (B) and (C) are FDA / CFDA-approved compound libraries used alone in SKOV3 (B) and ES-2 cells (C) as control groups, and combined with the PARP inhibitor niraparib as a control group. Heat maps were drawn based on cell activity to screen for synergistic drug combinations; (DG) After treating ovarian cancer cell lines (SKOV3 / ES-2 / OVCA420 and HEYA8) with gradient niraparib and lapatinib at different concentrations, MTS was added to detect cell proliferation ability; (H) shows the IC of niraparib and lapatinib on ovarian cancer cell lines (SKOV3 / ES-2 / OVCA420 and HEYA8) 50 .

[0016] Figure 2 The effect of lapatinib combined with niraparib on the growth of ovarian cancer cells in the present invention; Figure 2 Middle: (AD) Proliferation graphs of SKOV3, ES-2, HeyA8, and OVCA420 cell lines after treatment with niraparib and lapatinib; "synergy" indicates that the CI is lower than 1, and "antagonism" indicates that the CI is higher than 1. Data are presented as mean ± SD (n = 4 per group); (EH) Niraparib and lapatinib were co-treated with SKOV3, ES-2, HeyA8, and OVCA420 cells to detect the clone formation ability and statistical significance; (I) Niraparib and lapatinib were co-treated with SKOV3 and ES-2 cells to detect their migration ability; (J) Western blot was used to detect the changes in the expression of MMP2 and vimentin in SKOV3 and ES-2 cells after co-treatment with niraparib and lapatinib.

[0017] Figure 3 The effect of lapatinib combined with niraparib on apoptosis of ovarian cancer cells in the present invention; Figure 3 Middle: (A) Cell apoptosis of ES-2 cells after 48 h of treatment with niraparib and lapatinib alone or in combination; (BC) Western blot was used to detect the expression changes of apoptosis-, cell cycle-, or proliferation-related proteins in SKOV3 and ES-2 cells after co-treatment with niraparib, and grayscale analysis was used for statistical analysis (DE).

[0018] Figure 4 The present invention is that lapatinib combined with niraparib inhibits ES-2 derived xenograft tumors in vivo; Figure 4 Middle: (A) 1x10 7 Cells were transplanted subcutaneously into mice, and when the tumor volume was ≥100 mm 3Drug administration was started at 1:00 pm, and the tumor volume was measured every 3 days; (B) The tumor was euthanized and weighed at the end of drug administration; (C) Protein was extracted from tumor tissues of each group, and the protein expression of cell proliferation-related proteins was detected by immunoblotting; (D) IHC staining of Ki67. Scale bar, 50 μm; (E) The weight of mice was measured every 3 days; (F) HE staining was performed on mice in each group; F The drug toxicity of mice in each group was detected by HE staining.

[0019] Figure 5 The results of immunoblotting and statistical analysis are shown in the present invention; Figure 5 Middle: (AC) Immunoblotting was used to detect the expression changes of proteins related to EGFR / AKT and ERK signaling pathways in SKOV3 and ES-2 cells after co-treatment with niraparib and lapatinib, and grayscale analysis was used for statistical analysis; (DE) Immunoblotting was used to detect the expression changes of PARP andγH2AX related proteins in SKOV3 and ES-2 cells with niraparib, and grayscale analysis was used for statistical analysis; (F) Immunofluorescence staining of H2AX in cells treated with niraparib and lapatinib; scale bar, 50 μm. DETAILED DESCRIPTION

[0020] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be clearly and completely described below in conjunction with the drawings of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0021] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in the present invention can be purchased from the market or prepared by existing methods.

[0022] In a first aspect, the present invention provides a use of an EGFR inhibitor combined with a PARP inhibitor in the preparation of a drug for treating ovarian cancer.

[0023] The present invention provides an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer. The EGFR inhibitor Lapatinib and the PARP inhibitor Niraparib are obtained by screening the FDA / CFDA approved compound library. The present invention finds that Lapatinib and Niraparib have a good synergistic anti-ovarian cancer effect. In order to further study the molecular mechanism of the synergistic effect of the two, the present invention also detects the cell death pattern after the combined action of the two by apoptosis and protein immunoblotting. Lapatinib combined with Niraparib effectively inhibits the growth of ovarian cancer in an ovarian cancer animal model, showing good clinical research and application value, and is expected to become a new tool for the treatment of ovarian cancer.

[0024] The EGFR inhibitors in the present invention are also called EGFR tyrosine kinase inhibitors (TKIs), which are a class of drugs that can inhibit the activity of EGFR, thereby preventing excessive proliferation of cancer cells. Currently, a number of EGFR inhibitors have been approved for marketing in China, including first-generation, second-generation and third-generation EGFR inhibitors, such as gefitinib, erlotinib, icotinib, afatinib, dacomitinib, osimertinib, ametinib, vometinib, befortinib and lapatinib.

[0025] The PARP inhibitor in the present invention is a medical agent that can affect the self-replication mode of cancer cells, including niraparib and the like.

[0026] In some specific embodiments, the EGFR inhibitor includes lapatinib, or a tautomer, stereoisomer, prodrug, a pharmaceutically acceptable salt, hydrate or solvate thereof; the PARP inhibitor includes niraparib, or a tautomer, stereoisomer, prodrug, a pharmaceutically acceptable salt, hydrate or solvate thereof.

[0027] In the present invention, "tautomers" refer to a functional group in certain compounds that changes its structure to become another functional group isomer, and can rapidly convert to each other, forming two isomers in dynamic equilibrium, and these two isomers are called tautomers.

[0028] The compounds of the present invention may include one or more asymmetric centers, and therefore may exist in a variety of stereoisomeric forms, for example, enantiomers and / or diastereoisomeric forms. For example, the compounds of the present invention may be a single enantiomer, diastereomer or geometric isomer (e.g., cis and trans isomers), or may be in the form of a mixture of stereoisomers, including a racemic mixture and a mixture rich in one or more stereoisomers. Isomers may be separated from the mixture by methods known to those skilled in the art, including: chiral high pressure liquid chromatography (HPLC) and the formation and crystallization of chiral salts; or preferred isomers may be prepared by asymmetric synthesis.

[0029] In the present invention, "prodrug" is also called prodrug, drug precursor, prodrug, etc., which refers to a compound obtained after chemical structure modification of a drug, which is inactive or less active in vitro, and releases active drugs in vivo through enzymatic or non-enzymatic conversion to exert drug effects. Prodrugs include, for example, compounds of the present invention in which hydroxyl, amino or sulfhydryl groups are bonded to any group, and when administered to a patient, they can be cleaved to form hydroxyl, amino or sulfhydryl groups. Therefore, representative examples of prodrugs include (but are not limited to) acetate / amide, formate / amide and benzoate / amide derivatives of hydroxyl, sulfhydryl and amino functional groups of compounds of formula (I). In addition, in the case of formic acid (-COOH), esters such as methyl esters, ethyl esters, etc. can be used. Esters themselves can be active and / or can be hydrolyzed under human body conditions. Suitable pharmaceutically acceptable in vivo hydrolyzable ester groups include those groups that are easily decomposed in the human body to release the parent acid or its salt.

[0030] Those skilled in the art will appreciate that organic compounds can form complexes with solvents in which they react or from which they precipitate or crystallize. These complexes are referred to as "solvates". When the solvent is water, the complex is referred to as a "hydrate". The present invention encompasses all solvates of the compounds of the present invention.

[0031] In the present invention, "pharmaceutically acceptable salts" refer to those salts that are suitable for contact with human and lower animal tissues without excessive toxicity, irritation, allergic reaction, etc., and are commensurate with a reasonable benefit / risk ratio within the scope of reliable medical judgment. Pharmaceutically acceptable salts are well known in the art. For example, Berge et al. describe pharmaceutically acceptable salts in detail in J.Pharmaccutical Sciences (1977) 66: 1-19. Pharmaceutically acceptable salts of the compounds of the present invention include salts derived from suitable inorganic and organic acids and inorganic and organic bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are salts formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid, or salts formed with organic acids, such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid. Also included are salts formed using conventional methods in the art, such as ion exchange methods. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, gluconate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthosulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, dihydroxynaphthoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, and the like. Pharmaceutically acceptable salts derived from suitable bases include alkali metal, alkaline earth metal, ammonium and N (C4 alkyl): salts. Representative alkali metal or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium salts, and the like. Other pharmaceutically acceptable salts include non-toxic ammonium salts, quaternary ammonium salts and amine cations formed with counterions, such as halides, hydroxides, formates, sulfates, phosphates, nitrates, lower alkyl sulfonates and aryl sulfonates, if appropriate.

[0032] In the present invention, "solvate" refers to a form of a compound or a salt thereof that is combined with a solvent and is usually formed by a solvent decomposition reaction. Conventional solvents include water, methanol, ethanol, acetic acid, DMSO, THF, ether, etc. The compounds described herein can be prepared, for example, in a crystalline form and can be solvated. Suitable solvates include pharmaceutically acceptable solvates and further include stoichiometric solvates and non-stoichiometric solvates. In some cases, the solvate will be able to separate, for example, when one or more solvent molecules are incorporated into the crystal lattice of the crystalline solid. "Solvate" includes solvates in solution and separable solvates. Representative solvates include hydrates, ethanolates, and methanolates.

[0033] In a second aspect, based on the same inventive concept, the present invention provides a drug for treating ovarian cancer, wherein the drug for treating ovarian cancer comprises an EGFR inhibitor and a PARP inhibitor.

[0034] In some specific embodiments, the EGFR inhibitor includes lapatinib, or a tautomer, stereoisomer, prodrug, a pharmaceutically acceptable salt, hydrate or solvate thereof; the PARP inhibitor includes niraparib, or a tautomer, stereoisomer, prodrug, a pharmaceutically acceptable salt, hydrate or solvate thereof.

[0035] In some specific embodiments, the dosage form of the drug for treating ovarian cancer includes oral solution, capsule, oil drop, powder, tablet or injection.

[0036] In some specific embodiments, the drug for treating ovarian cancer further comprises a pharmaceutically acceptable carrier or excipient.

[0037] In the present invention, "pharmaceutical" can also be referred to as "pharmacologically acceptable", which refers to a substance that is not substantially undesirable biologically or otherwise, that is, the substance can be administered to an individual without causing any undesirable biological effect or interacting in a harmful manner with any other components of the composition containing such a substance.

[0038] In the present invention, "carrier" may also be referred to as "drug carrier", which refers to a system that can change the way drugs enter the human body and their distribution in the body, control the release rate of drugs and deliver drugs to targeted organs.

[0039] In the present invention, "excipients" can also be called "auxiliary materials", such as physiological saline, glucose, vitamin C, amino acids, etc., which refer to additives other than the main drug in pharmaceutical preparations. For example, adhesives, fillers, disintegrants, lubricants in tablets; wine, vinegar, medicinal juice, etc. in Chinese medicine pills; matrix parts in semi-solid preparations such as ointments and creams; preservatives, antioxidants, flavoring agents, aromatics, cosolvents, emulsifiers, solubilizers, osmotic pressure regulators, colorants, etc. in liquid preparations can all be called excipients.

[0040] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended only to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples that do not specify specific conditions are usually measured according to national standards. If there is no corresponding national standard, then the conditions recommended by the manufacturer are followed.

[0041] Example

[0042] This example provides an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer. The specific process is as follows:

[0043] Step 1. Cell lines and culture conditions: Ovarian cell lines (SKOV3, ES-2, OVCA420, and HEYA8) were purchased from the American Type Cell Culture Center (ATCC). SKOV3, ES-2, OVCA420, and HEYA8 cells were cultured in DMEM medium containing 10% fetal bovine serum, 100 U / ml penicillin, and 100 μg / ml streptomycin. The cells were cultured in a completely humidified environment with 5% CO. 2 / 95% air, 37℃.

[0044] Step 2: Cell viability assay and combination index (CI): SKOV3, ES-2, OVCA420 and HEYA8 cells were plated at 8×10 3 The cells were seeded in a 96-well plate at a density of 100 μg / mL and then treated with the compound for 72 h. 20 μL of MTS reagent was added to each well, the cells were incubated for 1-2 h, and then the absorbance was measured at 490 nm. The results of the MTS test were analyzed using GraphPad Prism8 software and were consistent with the half-inhibition rate (half-inhibitory concentration) of cell proliferation. The results are expressed as the average of three repeated measurements. Calcusyn software was used to calculate the CI value and affected fraction (FA) of the combined experiment. FA refers to the proportion of cells whose cell viability is affected. Survival plots and scatter plots of CI values ​​were drawn using GraphPad Prism8.

[0045] Step 3: Colony formation rate: SKOV3, ES-2, OVCA420 and HEYA8 cells were plated at 4×10 per well.3 The concentration of cells was inoculated into a 12-well plate. 1 mL of culture medium containing 10% fetal bovine serum was added to each well. After the cells attached, the drugs were added to treat the cells. An equal volume of DMSO was added as a control. After 1 week, the cells formed colonies, which were washed with PBS, fixed with 4% paraformaldehyde for 30 minutes, removed with 4% paraformaldehyde, and stained with 2% crystal violet solution for 15 minutes. Finally, the floating color was washed in water and air-dried. The number of cell colonies in each well was counted, and the colony formation rate was calculated: colony formation rate (%) / colony formation rate (control) (%).

[0046] Step 4: Western blot analysis: First, lyse the drug-treated cells or tumor tissues with RIPA lysis buffer, and quantify the BCA protein for Western blot experiments. Use Image J software for optical density analysis.

[0047] Step 5, flow cytometry. Add lapatinib, niraparib and combination drugs to ES-2 cells respectively. After 48 hours of treatment with the corresponding drugs, collect the digested cells with the supernatant. Add 100 μL binding buffer, 1 μL RNase (Sigma, USA), 2 μL Annexin V-FITC (BD, USA) and 2 μL propidium iodide (PI) (Sigma, USA) to each test tube and keep away from light at room temperature. Incubate for 15 minutes under this condition, detect by flow cytometry, and analyze by FACS Calibur (BD).

[0048] Step 6: Immunofluorescence Place the cover glass in a 24-well plate and inoculate 8×10 3 Cells. Cells were treated with different concentrations of lapatinib and niraparib and incubated at 37°C, 5% CO2 and 95% humidity for 48 h. Fixed cells were permeabilized with 0.2% Triton (Sangon, China) in 1× PBS for 30 min. Cells were incubated in 1% BSA (Sangon) and 0.2% Triton / PBS for 30 min. They were then incubated with rabbit anti-γH2AX antibody (1:400) at 4°C overnight. Cells were then washed three times with 0.2% triton / PBS for 3 min each and incubated with the second anti-rabbit 800 antibody for 1 h in the dark. Cell nuclei were stained with DAPI (D9542, Sigma) for 5 min and washed three times with 0.2% triton / PBS for 5 min each. Images were taken using an Olympus inverted fluorescence microscope.

[0049] Step 7: Xenograft tumor growth A 6-8 week old female nude mouse was subcutaneously injected with 1×10 7 ES-2 cells. When the average tumor volume reaches 100mm 3The mice were randomly divided into groups and injected intraperitoneally for 18 days. The body weight and tumor size of the rats were measured. After administration, the mice were removed and the tumors and main organs were dissected for subsequent experiments.

[0050] The test results are as follows:

[0051] Figure 1 Figure 3 is a diagram showing that lapatinib is identified as a target drug for combination testing with niraparib in the present invention; Figure 1 Middle: (A) Screening scheme; Cell viability of niraparib and non-niraparib drugs in SKOV3 (B) and ES-2 cells (C); (B) and (C) are FDA / CFDA-approved compound libraries used alone in SKOV3 (B) and ES-2 cells (C) as control groups, and combined with the PARP inhibitor niraparib as a control group. Heat maps were drawn based on cell activity to screen for synergistic drug combinations; (DG) After treating ovarian cancer cell lines (SKOV3 / ES-2 / OVCA420 and HEYA8) with gradient niraparib and lapatinib at different concentrations, MTS was added to detect cell proliferation ability; (H) shows the IC of niraparib and lapatinib on ovarian cancer cell lines (SKOV3 / ES-2 / OVCA420 and HEYA8) 50 .

[0052] Figure 2 The effect of lapatinib combined with niraparib on the growth of ovarian cancer cells in the present invention; Figure 2 Middle: (AD) Proliferation graphs of SKOV3, ES-2, HeyA8, and OVCA420 cell lines after treatment with niraparib and lapatinib; "synergy" indicates that the CI is lower than 1, and "antagonism" indicates that the CI is higher than 1. Data are presented as mean ± SD (n = 4 per group); (EH) Niraparib and lapatinib were co-treated with SKOV3, ES-2, HeyA8, and OVCA420 cells to detect the clone formation ability and statistical significance; (I) Niraparib and lapatinib were co-treated with SKOV3 and ES-2 cells to detect their migration ability; (J) Western blot was used to detect the changes in the expression of MMP2 and vimentin in SKOV3 and ES-2 cells after co-treatment with niraparib and lapatinib.

[0053] Figure 3 The effect of lapatinib combined with niraparib on apoptosis of ovarian cancer cells in the present invention; Figure 3 Middle: (A) Cell apoptosis of ES-2 cells after 48 h of treatment with niraparib and lapatinib alone or in combination; (BC) Western blot was used to detect the expression changes of apoptosis-, cell cycle-, or proliferation-related proteins in SKOV3 and ES-2 cells after co-treatment with niraparib, and grayscale analysis was used for statistical analysis (DE).

[0054] Figure 4 The present invention is that lapatinib combined with niraparib inhibits ES-2 derived xenograft tumors in vivo; Figure 4 Middle: (A) 1x10 7 Cells were transplanted subcutaneously into mice, and when the tumor volume was ≥100 mm 3 Drug administration was started at 1:00 pm, and the tumor volume was measured every 3 days; (B) The tumor was euthanized and weighed at the end of drug administration; (C) Protein was extracted from tumor tissues of each group, and the protein expression of cell proliferation-related proteins was detected by immunoblotting; (D) IHC staining of Ki67. Scale bar, 50 μm; (E) The weight of mice was measured every 3 days; (F) HE staining was performed on mice in each group; F The drug toxicity of mice in each group was detected by HE staining.

[0055] Figure 5 The results of immunoblotting and statistical analysis are shown in the present invention; Figure 5 Middle: (AC) Immunoblotting was used to detect the expression changes of proteins related to EGFR / AKT and ERK signaling pathways in SKOV3 and ES-2 cells after co-treatment with niraparib and lapatinib, and grayscale analysis was used for statistical analysis; (DE) Immunoblotting was used to detect the expression changes of PARP andγH2AX related proteins in SKOV3 and ES-2 cells with niraparib, and grayscale analysis was used for statistical analysis; (F) Immunofluorescence staining of H2AX in cells treated with niraparib and lapatinib; scale bar, 50 μm.

[0056] In the present invention, it is found that the application of niraparib combined with lapatinib can inhibit the activation of EGFR, thereby inhibiting the expression of p-AKT and p-ERK, showing a good synergistic inhibition of apoptosis of ovarian cancer cells, and can induce DNA damage and increase PARP shearing, ultimately inhibiting the progression of ovarian cancer in vitro and in vivo. The results of the present invention provide new ideas for the clinical combination strategy of two different inhibitors, and provide a new treatment approach for alleviating the drug resistance status of ovarian cancer patients.

[0057] In summary, the present invention provides an application of an EGFR inhibitor combined with a PARP inhibitor in the treatment of ovarian cancer. The EGFR inhibitor Lapatinib and the PARP inhibitor Niraparib are obtained by screening the FDA / CFDA approved compound library. The present invention finds that Lapatinib and Niraparib have a good synergistic anti-ovarian cancer effect. In order to further study the molecular mechanism of the synergistic effect of the two, the present invention also detects the cell death pattern after the combined action of the two by apoptosis and protein immunoblotting. The combination of Lapatinib and Niraparib effectively inhibits the growth of ovarian cancer in an ovarian cancer animal model, showing good clinical research and application value, and is expected to become a new tool for the treatment of ovarian cancer.

[0058] The foregoing is merely a specific embodiment of the present invention, which enables those skilled in the art to understand or implement the present invention. Various modifications to these embodiments will be apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to the embodiments shown herein, but rather to the widest scope consistent with the principles and novel features claimed herein.

Claims

1. Use of an EGFR inhibitor combined with a PARP inhibitor in the preparation of a drug for treating ovarian cancer.

2. The use according to claim 1, characterized in that: The EGFR inhibitor includes lapatinib, or a tautomer, a stereoisomer, a prodrug, a pharmaceutically acceptable salt, a hydrate or a solvate thereof.

3. The use according to claim 1, characterized in that: The PARP inhibitor includes niraparib, or a tautomer, a stereoisomer, a prodrug, a pharmaceutically acceptable salt, a hydrate or a solvate thereof.

4. A drug for treating ovarian cancer, characterized in that: The drugs for treating ovarian cancer include EGFR inhibitors and PARP inhibitors.

5. The drug for treating ovarian cancer according to claim 4, characterized in that: The EGFR inhibitor includes lapatinib, or its tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate; the PARP inhibitor includes niraparib, or its tautomer, stereoisomer, prodrug, pharmaceutically acceptable salt, hydrate or solvate.

6. The drug for treating ovarian cancer according to claim 4, characterized in that: The preparation dosage form of the drug for treating ovarian cancer includes oral liquid, capsule, oil drop, powder, tablet or injection.

7. The drug for treating ovarian cancer according to claim 4, characterized in that: The drug for treating ovarian cancer also includes a pharmaceutically acceptable carrier or excipient.

Images