Composition for enhancing anticancer effect of EGFR-targeted anticancer agent, comprising aripiprazole as active ingredient

Abstract

The present invention provides a novel therapeutic means for cancer cells resistant to epidermal growth factor receptor (EGFR)-targeted anticancer agents. Almost no cell death was observed when cancer cells having an exon 19 deletion mutation in the EGFR gene (EGFR Del19), cancer cells having a substitution mutation in which the 790th amino acid threonine in the EGFR protein is substituted with methionine (EGFR T790M), and cancer cells having a substitution mutation in which the 858th amino acid leucine is substituted with arginine (EGFR L858R) were treated solely with aripiprazole or solely with an EGFR-targeted anticancer agent. However, cell viability decreased and cleavage of the apoptosis marker PARP was observed when the cancer cells were treated with aripiprazole in combination with an EGFR-targeted anticancer agent. Therefore, aripiprazole is provided as an enhancer of the anticancer effect of EGFR-targeted anticancer agents.

Description

Composition for enhancing the anticancer effect of an EGFR-targeted anticancer agent containing aripiprazole as an active ingredient

[0001] The present invention relates to a composition for enhancing an anticancer agent targeting EGFR (epidermal growth factor receptor).

[0002] Epidermal growth factor receptor (EGFR) targeted therapies inhibit the growth and spread of cancer cells by blocking EGFR activity. EGFR is a tyrosine kinase receptor that plays a crucial role in cell growth and proliferation and is overexpressed or mutated in many cancers. Overexpression of EGFR promotes tumor growth and progression by inducing angiogenesis, cancer invasion, and metastasis. EGFR-targeted therapies bind to EGFR and block this signaling pathway, thereby inhibiting cancer cell growth and inducing cell death. EGFR-targeted therapies can be used to treat a variety of cancers, including non-small cell lung cancer and colorectal cancer, which primarily harbor EGFR mutations. However, despite the use of EGFR-targeted therapies, many cancer patients develop resistance to EGFR therapy. Resistance to EGFR-targeted therapies is caused by various genetic and non-genetic mechanisms, with EGFR mutations being the most common type of resistance.

[0003] EGFR mutations occur in the tyrosine kinase domain of EGFR, increasing the activity of EGFR kinase, thereby continuously activating the cell signaling system and ensuring continued cell differentiation and growth. The most representative EGFR mutations are the exon 19 deletion mutation (Del 19) in the EGFR gene and the mutation in which leucine, the 858th amino acid of the EGFR protein, is substituted with arginine (L858R). These EGFR mutations are sensitive to first- and second-generation EGFR targeted therapies. However, a mutation in which threonine, the 790th amino acid of the EGFR protein, is substituted with methionine (T790M) has been identified in cancers that exhibit acquired resistance mechanisms to first- and second-generation EGFR targeted therapies. While EGFR-targeted therapies demonstrate excellent anticancer effects in the early stages of cancer treatment, treatment resistance develops in a vast majority of patients, necessitating new approaches to addressing this issue. A representative strategy for overcoming the problem of targeted drug resistance caused by intratumor heterogeneity is combination therapy, which involves prescribing two or more different drugs simultaneously to reduce the risk of developing resistant cell populations.

[0004] The purpose of the present invention is to provide a pharmaceutical composition for enhancing the anticancer effect of an EGFR (epidermal growth factor receptor)-targeting anticancer agent, and to provide a composition capable of enhancing the sensitivity and anticancer effect of an EGFR-targeting anticancer agent to cancer cells by confirming that when aripiprazole is co-treated with EGFR-targeting cancer cells, the survival rate of cancer cells decreases, a synergistic effect is demonstrated, and apoptosis is induced.

[0005] The present invention provides a composition for enhancing the anticancer effect of an EGFR target anticancer agent, comprising aripiprazole as an active ingredient.

[0006] According to the present invention, when aripiprazole was treated alone or with an EGFR-targeted anticancer drug alone, cell death was hardly observed in cancer cells with an exon 19 deletion mutation (EGFRDel19) in the EGFR gene, a leucine at position 858 in the EGFR protein was substituted with an arginine mutation (EGFRL858R), or a threonine at position 790 was substituted with a methionine mutation (EGFRT790M). However, when aripiprazole and an EGFR-targeted anticancer drug were combined, cell viability decreased, and PARP cleavage, a cell death marker, was confirmed. Therefore, aripiprazole can be provided as an agent that enhances the anticancer effect of an EGFR-targeted anticancer drug.

[0007] Figure 1 shows the results of evaluating the effects of aripiprazole alone, osimertinib alone, or aripiprazole and osimertinib in combination on cell viability in HCC827 cells with an exon 19 deletion mutation (EGFRDel19).

[0008] Figure 2 shows the results of analyzing the synergistic effect of drugs as a score in the Bliss model based on the effect of combined treatment of aripiprazole and osimertinib on cell viability in HCC827 cells with exon 19 deletion mutation (EGFRDel19).

[0009] Figure 3 shows the results of analyzing the effects of aripiprazole alone, EGFR-targeted anticancer drugs alone, or aripiprazole and EGFR-targeted anticancer drugs in combination on PARP cleavage in HCC827 cells with an exon 19 deletion mutation (EGFRDel19).

[0010] Figure 4 shows the results of analyzing the effects of aripiprazole alone, EGFR-targeted anticancer drugs alone, or aripiprazole and EGFR-targeted anticancer drugs in combination on PARP cleavage in NCI-H1650 cells with an exon 19 deletion mutation (EGFRDel19).

[0011] Figure 5 shows the results of analyzing the effects of aripiprazole alone, EGFR-targeted anticancer drugs alone, or aripiprazole and EGFR-targeted anticancer drugs in combination on PARP cleavage in NCI-H1975 cells, which are double mutants (EGFRL858R / T790M) in which threonine, the 790th amino acid in the EGFR protein, is substituted with methionine, and leucine, the 858th amino acid, is substituted with arginine.

[0012] The terms used in this specification have been selected from widely used, current terms, taking into account the functions of the present invention. However, these terms may vary depending on the intentions of those skilled in the art, precedents, the emergence of new technologies, etc. Furthermore, in certain cases, terms may be arbitrarily selected by the applicant, and in such cases, their meanings will be described in detail in the relevant description of the invention. Therefore, the terms used in this invention should not be defined simply as names, but rather based on their inherent meanings and the overall content of the present invention.

[0013] Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with their meaning in the context of the relevant technology, and shall not be interpreted in an idealized or overly formal sense unless explicitly defined herein.

[0014] Hereinafter, the present invention will be described in more detail.

[0015] The present invention provides a composition for enhancing the anticancer effect of an EGFR target anticancer agent, comprising aripiprazole as an active ingredient.

[0016] The above EGFR targeting anticancer agent is at least one selected from osimertinib, lazertinib, afatinib, gefitinib, and erlotinib.

[0017] The above cancer is at least one selected from melanoma, breast cancer, lymphoma, brain tumor, brain cancer, colon cancer, prostate cancer, lung cancer, osteosarcoma, bone cancer, pancreatic cancer, skin cancer, oral cancer, oropharyngeal cancer, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, endometrial cancer, cervical cancer, vaginal cancer, small intestine cancer, thyroid cancer, parathyroid cancer, chronic or acute leukemia, bladder cancer, kidney cancer, and liver cancer.

[0018] The above composition may be administered concurrently, separately, or sequentially with an EGFR targeting anticancer agent.

[0019] The composition may enhance the anticancer effect of an EGFR-targeted anticancer agent against cancer cells resistant to such agents. Alternatively, the composition may enhance the anticancer effect of an EGFR-targeted anticancer agent by inducing apoptosis within cancer cells. The composition may enhance the cancer cell growth inhibition effect of an EGFR-targeted anticancer agent.

[0020] The cancer cells above are EGFR (epidermal growth factor receptor) mutant cancer cells, and the mutation is at least one selected from among an exon 19 deletion mutation in the EGFR gene (EGFRDel19), a mutation in which threonine at the 790th amino acid in the EGFR protein is substituted with methionine (EGFRT790M), and a mutation in which leucine at the 858th amino acid in the EGFR protein is substituted with arginine (EGFRL858R).

[0021] The above composition may be a pharmaceutical composition administered in combination with an EGFR targeting anticancer agent for the prevention or treatment of cancer.

[0022] The pharmaceutical composition of the present invention can be manufactured in a unit dose form or can be manufactured by placing it in a multi-dose container by formulating it using a pharmaceutically acceptable carrier according to a method that can be easily performed by a person having ordinary skill in the art to which the present invention pertains.

[0023] The pharmaceutically acceptable carriers mentioned above are those commonly used in formulations, and include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition to the above components, the pharmaceutical composition of the present invention may further include a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifier, a suspending agent, a preservative, and the like.

[0024] In the present invention, the content of the additive included in the pharmaceutical composition is not particularly limited and can be appropriately adjusted within the content range used in conventional formulations.

[0025] The above pharmaceutical composition may be formulated in the form of one or more external preparations selected from the group consisting of injectable formulations such as aqueous solutions, suspensions, emulsions, pills, capsules, granules, tablets, creams, gels, patches, sprays, ointments, ointments, lotions, liniments, pastes, and cataplasmas.

[0026] The pharmaceutical composition of the present invention may further comprise pharmaceutically acceptable carriers and diluents for formulation. The pharmaceutically acceptable carriers and diluents include, but are not limited to, excipients such as starches, sugars, and mannitol; fillers and extenders such as calcium phosphate; cellulose derivatives such as carboxymethylcellulose and hydroxypropylcellulose; binders such as gelatin, alginates, and polyvinyl pyrrolidone; lubricants such as talc, calcium stearate, hydrogenated castor oil, and polyethylene glycol; disintegrants such as povidone and crospovidone; and surfactants such as polysorbates, cetyl alcohol, and glycerol. The pharmaceutically acceptable carriers and diluents may be biologically and physiologically compatible with the subject. Examples of diluents include, but are not limited to, saline, aqueous buffers, solvents, and / or dispersion media.

[0027] The pharmaceutical composition of the present invention may be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically) depending on the intended method. In the case of oral administration, it may be formulated as tablets, troches, lozenges, aqueous suspensions, oily suspensions, prepared powders, granules, emulsions, hard capsules, soft capsules, syrups, or elixirs. In the case of parenteral administration, it may be formulated as injections, suppositories, powders for respiratory inhalation, aerosols for sprays, ointments, powders for application, oils, creams, etc.

[0028] The dosage of the pharmaceutical composition of the present invention may vary depending on the patient's condition and weight, age, sex, health status, dietary constitution, nature of the formulation, severity of the disease, administration time of the composition, administration method, administration period or interval, excretion rate, and drug form, and may be appropriately selected by a person skilled in the art. For example, the dosage may range from about 0.1 to 10,000 mg / kg, but is not limited thereto, and may be administered once or several times a day in divided doses.

[0029] The pharmaceutical composition may be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically) depending on the intended method. The pharmaceutically effective amount and effective dosage of the pharmaceutical composition of the present invention may vary depending on the formulation method, administration method, administration time, and / or administration route of the pharmaceutical composition, and a person skilled in the art can easily determine and prescribe an effective dosage for the intended treatment. The pharmaceutical composition of the present invention may be administered once a day or divided into several doses.

[0030] The above composition may be a health functional food composition administered in combination with an EGFR target anticancer agent for the prevention or improvement of cancer.

[0031] The present invention can be generally used as a commonly used food.

[0032] The food composition of the present invention can be used as a health functional food. The term "health functional food" refers to a food manufactured and processed using raw materials or ingredients with functional properties beneficial to the human body, as defined by the Health Functional Food Act. "Functionality" refers to ingestion for the purpose of obtaining beneficial effects for health purposes, such as regulating nutrients for the structure and functions of the human body or physiological effects.

[0033] The food composition of the present invention may include conventional food additives, and its suitability as the "food additive" is determined by the specifications and standards for the relevant item in accordance with the general provisions and general test methods of the Food Additive Code approved by the Ministry of Food and Drug Safety, unless otherwise specified.

[0034] Items listed in the above "Food Additives Code" include, for example, chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamic acid; natural additives such as persimmon pigment, licorice extract, crystalline cellulose, high-molecular-weight pigment, and guar gum; and mixed preparations such as sodium L-glutamate preparations, alkaline agents for noodles, preservative preparations, and tar color preparations.

[0035] The food composition of the present invention can be manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc.

[0036] For example, among health functional foods in capsule form, hard capsules can be manufactured by mixing and filling a composition according to the present invention with additives such as excipients into a conventional hard capsule, and soft capsules can be manufactured by mixing the composition according to the present invention with additives such as excipients and filling it into a capsule base such as gelatin. The soft capsules may contain a plasticizer such as glycerin or sorbitol, a coloring agent, a preservative, etc., as needed.

[0037] The definitions of terms for the above excipients, binders, disintegrants, lubricants, flavoring agents, etc. are described in literature known in the art and include those with identical or similar functions. There are no specific restrictions on the type of food, and all health functional foods in the conventional sense are included.

[0038] As used herein, the term "prevention" refers to any action that suppresses or delays the onset of a disease by administering a composition according to the present invention. The term "treatment" refers to any action that improves or beneficially alters the symptoms of a disease by administering a composition according to the present invention. As used herein, "improvement" refers to any action that improves the worsening condition of a disease by administering or ingesting a composition according to the present invention to a subject.

[0039] Hereinafter, to aid understanding of the present invention, examples will be given in detail. However, the following examples are intended only to illustrate the scope of the present invention and are not intended to limit its scope. These examples are provided to more fully explain the present invention to those of average skill in the art.

[0040]

[0041] <Example 1>

[0042] The effect of aripiprazole on the sensitivity of epidermal growth factor receptor (EGFR) genetic mutation therapy in cancer cells harboring EGFR activating mutations was evaluated. The anticancer effect of osimertinib, an EGFR-targeted anticancer drug, was evaluated in HCC827 cells (EGFRDel19 mutant), a lung cancer cell line harboring EGFR activating mutations, after treatment with aripiprazole. Osimertinib is a targeted anticancer drug used in the treatment of non-small cell lung cancer that selectively inhibits the tyrosine kinase activity of EGFR, thereby inhibiting cancer cell proliferation.

[0043] The anticancer effect was evaluated by measuring the viability of cancer cells using the LDH (lactate dehydrogenase) cytotoxicity assay. HCC827 cells were seeded in 96-well plates at a concentration of 5,000 cells / well and pre-cultured for 24 hours. The medium was then removed, and aripiprazole was treated alone at concentrations of 0 μM, 2.5 μM, 5 μM, 10 μM, 20 μM, or 30 μM, or in combination with osimertinib at concentrations of 1 nM, 2 nM, or 5 nM. After treating the cancer cells with the reagents, the viability of cells was measured 36 hours later using the EZ-LDH cytotoxicity assay kit from Dozen.

[0044] As shown in Figure 1, it was confirmed that cell viability was maintained at 100% when treated with aripiprazole alone or with osimertinib alone. The above results demonstrate that combined treatment with aripiprazole or osimertinib exhibits superior anticancer effects, with a significant decrease in cancer cell viability compared to treatment with aripiprazole or osimertinib alone.

[0045]

[0046] <Example 2>

[0047] Based on the effect of combined treatment of aripiprazole and an EGFR-targeting anticancer drug on the survival rate of cancer cells, the synergistic effect of the drugs was calculated as a score using the Bliss model. HCC827 cells were cultured using the same method as in Example 1, and then treated with aripiprazole alone at a concentration of 0 μM, 2.5 μM, 5 μM, 10 μM, 20 μM, or 30 μM, or in combination with osimertinib at a concentration of 1 nM, 2 nM, or 5 nM. After treating the cancer cells with the reagents, the cell viability was measured 36 hours later using the EZ-LDH cytotoxicity assay kit from Dozen. The measured viability was applied to the Bliss model, and the synergy score was calculated using SynergyFinder (https: / / synergyfinder.fimm.fi / ). A synergy matrix was created using SynergyFinder based on the calculated synergy score.

[0048] As shown in Figure 2, the synergistic effect score for the combined administration of aripiprazole and osimertinib was calculated to be 12.374. The results demonstrate that the combined administration of aripiprazole and osimertinib enhances the anticancer effect.

[0049]

[0050] <Example 3>

[0051] To analyze the effect of aripiprazole on the apoptotic action of EGFR-targeted anticancer drugs in cancer cells, the degree of PARP cleavage, an apoptosis marker, was determined by Western blotting. EGFR-targeted anticancer drugs used were osimertinib, lazertinib, afatinib, gefitinib, and erlotinib. To evaluate the effect of EGFR-targeted anticancer drugs, among lung cancer cell lines, HCC827 cells with an exon 19 deletion mutation (EGFRDel19) in the EGFR gene, NCI-H1650 cells, and NCI-H1975 cells with a double mutation (EGFRL858R / T790M) in which threonine at position 790 is substituted with methionine and leucine at position 858 is substituted with arginine in the EGFR protein were used. Each cancer cell line was treated with aripiprazole alone or in combination with an EGFR-targeting anticancer drug. After 16 hours, cells were washed with PBS, protein was extracted, and PARP cleavage levels (C-PARP) were analyzed by Western blotting.

[0052] As shown in Fig. 3, in HCC827 cells (EGFRDel19 mutant), PARP cleavage (C-PARP) was not observed when aripiprazole was treated alone or when an EGFR targeting anticancer drug was treated alone, but PARP cleavage (C-PARP) was observed in all cases when aripiprazole and an EGFR targeting anticancer drug were combined.

[0053] In addition, as shown in Fig. 4, in NCI-H1650 cells (EGFRDel19 mutant), PARP cleavage (C-PARP) was not observed when aripiprazole was treated alone or when an EGFR targeting anticancer drug was treated alone, but PARP cleavage (C-PARP) was observed in all cases when aripiprazole and an EGFR targeting anticancer drug were combined.

[0054] In addition, as shown in Fig. 5, in NCI-H1975 cells (EGFRL858R / T790M double mutant), PARP cleavage (C-PARP) was not observed when aripiprazole was treated alone or when an EGFR target anticancer drug was treated alone, but PARP cleavage (C-PARP) was observed in all cases when aripiprazole and an EGFR target anticancer drug were combined.

[0055] The above results demonstrate that combined treatment with aripiprazole or osimertinib induces PARP cleavage in cancer cells compared to treatment with aripiprazole or osimertinib alone.

[0056]

[0057] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions merely represent preferred embodiments and are not intended to limit the scope of the present invention. In other words, the substantial scope of the present invention is defined by the appended claims and their equivalents.

[0058] Numerical ranges are inclusive of the values ​​defined in the ranges above. Any maximum numerical limitation given throughout this specification includes any lower numerical limitation, as if that lower numerical limitation were explicitly stated. Any minimum numerical limitation given throughout this specification includes any higher numerical limitation, as if that higher numerical limitation were explicitly stated. Any numerical limitation given throughout this specification will include any better numerical range within the broader numerical range, as if that narrower numerical limitation were explicitly stated.

Claims

1. A composition for enhancing the anticancer effect of an EGFR target anticancer agent, comprising aripiprazole as an active ingredient.

2. A composition according to claim 1, characterized in that the EGFR target anticancer agent is at least one selected from osimertinib, lazertinib, afatinib, gefitinib, and erlotinib.

3. A composition according to claim 1, characterized in that the cancer is at least one selected from among melanoma, breast cancer, lymphoma, brain tumor, brain cancer, colon cancer, prostate cancer, lung cancer, osteosarcoma, bone cancer, pancreatic cancer, skin cancer, oral cancer, oropharyngeal cancer, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, endometrial cancer, cervical cancer, vaginal cancer, small intestine cancer, thyroid cancer, parathyroid cancer, chronic or acute leukemia, bladder cancer, kidney cancer, and liver cancer.

4. A composition according to claim 1, characterized in that the composition is administered concurrently, separately, or sequentially with an EGFR targeting anticancer agent.

5. A composition according to claim 1, characterized in that the composition enhances the anticancer effect of an EGFR-targeted anticancer drug against cancer resistant to the EGFR-targeted anticancer drug.

6. A composition according to claim 1, characterized in that the composition induces apoptosis in cancer cells and enhances the anticancer effect of an EGFR-targeting anticancer agent.

7. A composition according to claim 1, characterized in that the composition enhances the cancer cell growth inhibition effect of an EGFR target anticancer agent.

8. A composition according to claim 7, characterized in that the cancer cells are EGFR (epidermal growth factor receptor) mutant cancer cells.

9. A composition according to claim 8, characterized in that the mutation is at least one selected from among an exon 19 deletion mutation in the EGFR gene (EGFRDel19), a mutation in which threonine, the 790th amino acid in the EGFR protein, is substituted with methionine (EGFRT790M), and a mutation in which leucine, the 858th amino acid in the EGFR protein, is substituted with arginine (EGFRL858R).

10. A composition according to claim 1, characterized in that the composition is a pharmaceutical composition administered in combination with an EGFR target anticancer agent for the prevention or treatment of cancer.

11. A composition according to claim 1, characterized in that the composition is a health functional food composition administered in combination with an EGFR target anticancer agent for the prevention or improvement of cancer.

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