KR20230021988A - Composition for treating, alleviating or preventing non small cell lung cancer comprising cyclophilin A inhibitors and afatinib - Google Patents

Abstract

The present invention relates to a composition for the treatment of non-small cell lung cancer exhibiting chemical synthetic lethality. Since the combined administration of a cyclosporin A inhibitor (cyclosporin A or compound 9) and afatinib has a synergistic synthetic lethal interaction for non-small cell lung cancer stem-like cells compared to individual components, it is possible to be usefully used for treating, preventing, or alleviating non-small cell lung cancer.

Description

Composition for treating, alleviating or preventing non small cell lung cancer comprising cyclophilin A inhibitors and afatinib

The present invention relates to a composition for treating non-small cell lung cancer exhibiting chemical synthetic lethality, and the like.

Cancer is one of the most common causes of death worldwide. Approximately 10 million new cases occur each year, accounting for approximately 12% of all deaths, making it the third leading cause of death. Among them, lung cancer refers to malignant tumors that occur in the lungs, and the types of primary lung cancers that occur in the lungs themselves are classified into non-small cell lung cancer and small cell lung cancer based on the size and shape of cancer cells. 80-85% of lung cancer is non-small cell lung cancer, which is further divided into adenocarcinoma (adenocarcinoma), squamous cell carcinoma, and large cell carcinoma.

Despite advances in the field of cancer treatment, the current leading treatments are surgery, radiation and chemotherapy. Chemotherapy approaches are used primarily to treat metastatic or particularly aggressive cancers. Most of the cancer chemotherapy drugs currently used clinically are cytotoxins. Cytotoxic agents work by injuring or killing rapidly growing cells.

On the other hand, cancer stem cells (CSC or Tumor Initiating Cells: TICs) refer to cancer cells (found in tumors or blood cancers) that have the ability to generate tumors. Cancer stem cells have the same characteristics as normal stem cells, specifically the ability to give rise to all cell types found in a specific cancer sample. That is, cancer stem cells are tumorigenic, unlike cancer cells that do not form tumors. Cancer stem cells generate tumors through self-renewal and differentiation capabilities, which are characteristics of stem cells, in various cell types.

In addition, it is differentiated from other groups in the tumor and causes recurrence and metastasis by generating new tumors. Therefore, the development of a specific treatment method targeting cancer stem cells can be a hope for increasing the survival rate and improving the quality of life of cancer patients, especially in patients with metastatic disease.

Accordingly, the inventor of the present invention completed the present invention as a result of studying a treatment target differentiated from existing targets for selectively controlling the self-renewal, proliferative, and differentiation-inducing abilities of cancer stem cells.

Republic of Korea Patent Registration No. 10-1419816

An object of the present invention is a cyclophilin A inhibitor or a pharmaceutically acceptable salt thereof; And to provide a pharmaceutical composition for preventing or treating non-small cell lung cancer comprising afatinib or a pharmaceutically acceptable salt thereof as an active ingredient.

Another object of the present invention is a cyclophilin A inhibitor or a salt thereof; And to provide a food composition for preventing or improving non-small cell lung cancer comprising afatinib or a salt thereof as an active ingredient.

In order to achieve the object of the present invention, a cyclophilin A inhibitor or a pharmaceutically acceptable salt thereof; And it provides a pharmaceutical composition for preventing or treating non-small cell lung cancer comprising afatinib or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention relates to a cyclophilin A inhibitor or a salt thereof; And it provides a food composition for preventing or improving non-small cell lung cancer comprising afatinib or a salt thereof as an active ingredient.

In one embodiment of the present invention, the cyclophilin A inhibitor may be cyclosporin A or compound 9 represented by Formula 2 below.

[Formula 2]

In one embodiment of the present invention, the molar concentration ratio of the cyclophilin A inhibitor and afatinib may be 1:0.08 to 1.5.

In one embodiment of the present invention, the composition can kill non-small cell lung cancer stem-like cells.

In one embodiment of the present invention, the non-small cell lung cancer stem-like cells may be of an epidermal growth factor receptor mutant type (EGFR mutant type).

The combined administration of a cyclophilin A inhibitor (cyclosporine A or compound 9) and afatinib shows synergistic synergistic lethal interactions on cancer stem-like cells compared to the individual components, thereby treating, preventing or improving non-small cell lung cancer. It can be used for useful purposes.

1 shows the degree of apoptosis of A549 (EGFR wild-type) non-small cell lung cancer stem-like cells when a cyclophilin A inhibitor (cyclosporine A or compound 9) and afatinib were co-administered.
Figure 2 is a graph showing the results of Figure 1.
Figure 3 shows the degree of apoptosis of NCI-H1650 (EGFR mutant) non-small cell lung cancer stem-like cells when a cyclophilin A inhibitor (cyclosporine A or compound 9) and afatinib were co-administered.
Figure 4 is a graph showing the results of Figure 3.

The inventor of the present invention discovers a treatment target differentiated from existing targets for selectively controlling the self-renewal ability, proliferation ability, and differentiation induction ability of cancer stem cells, and various types of “customized anticancer agents for cancer stem cells”. With the goal of “suggesting a new strategy for development”, “screening for anti-cancer drugs that exhibit chemical synthetic lethality specific to cancer stem cells” without affecting non-cancer stem cells will lead to new cancer lines. The target cell therapy was discovered, and the present invention was completed by discovering that there was a synergistic proliferation inhibitory effect on non-small cell lung cancer stem-like cells of EGFR mutant type. .

Accordingly, the present invention provides a cyclophilin A inhibitor or a pharmaceutically acceptable salt thereof; and a pharmaceutical composition for preventing or treating non-small cell lung cancer comprising afatinib or a pharmaceutically acceptable salt thereof as an active ingredient.

In the present invention, the cyclosporin A inhibitor, cyclosporin A, is an immunosuppressive agent that inhibits the activity of calcineurin phosphatase by combining calcineurin and cyclosporin. has the structure of

[Formula 1]

In the present invention, compound 9 (23-demethyl 8,13-deoxynargenicin), the nargenisin A1 derivative, is a novel cyclophilin A inhibitor recently discovered by the present inventors (Patent Application No. 10-2020-0102555; Int J Mol Sci. 2021 Mar 1;22(5):2473. doi: 10.3390/ijms22052473), which has the structure of Formula 2 below,

[Formula 2]

In the present invention, afatinib is a target anticancer agent used for the treatment of specific non-small cell lung cancer, and since it inhibits the proliferation of cancer cells by selectively inhibiting the tyrosine kinase activity of EGFR, non-small cell lung cancer patients with EGFR genetic mutations It is used for the treatment of, and has the structure of Formula 3 below,

[Formula 3]

In the present invention, "non-small cell lung cancer" means that the size of cancer cells confirmed under a microscope is small compared to small cell lung cancer, and the size of cancer cells confirmed under a microscope is not small. It includes, but is not limited to, adenosquamous cell carcinoma, sarcoma carcinoma, carcinoid tumor, salivary gland carcinoma, unclassified carcinoma, and the like.

In the present invention, "pharmaceutically acceptable salt" means a salt formed with any inorganic or organic acid or base that does not cause serious irritation to the organism to which it is administered and does not impair the biological activity and physical properties of the compound. As the salt, a salt commonly used in the art, such as an acid addition salt formed by a pharmaceutically acceptable free acid, may be used.

As used herein, the term "prevention" refers to any activity that inhibits or delays the onset of brain cancer by administration of the pharmaceutical composition according to the present invention.

As used herein, the term "treatment" refers to all activities that improve or beneficially change symptoms caused by brain cancer by administration of the pharmaceutical composition according to the present invention.

The pharmaceutical composition according to the present invention comprises a cyclophilin A inhibitor or a salt thereof; and afatinib or a salt thereof as an active ingredient, and may include a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is one commonly used in formulation and includes, but is not limited to, saline solution, sterile water, Ringer's solution, buffered saline solution, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol, liposome, etc. It is not, and if necessary, other conventional additives such as antioxidants and buffers may be further included.

In addition, diluents, dispersants, surfactants, binders, lubricants, and the like may be additionally added to prepare formulations for injections such as aqueous solutions, suspensions, emulsions, etc., pills, capsules, granules, or tablets. Regarding a suitable pharmaceutically acceptable carrier and formulation, it can be preferably formulated according to each component using the method disclosed in Remington's literature. The pharmaceutical composition of the present invention is not particularly limited in dosage form, but may be formulated as an injection or oral intake.

The pharmaceutical composition of the present invention can be administered orally or parenterally (for example, intravenously or subcutaneously) depending on the desired method, and the dosage is the condition and weight of the patient, the severity of the disease, the form of the drug, Depending on the route and time of administration, it can be appropriately selected by those skilled in the art.

The composition according to the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level is based on the type, severity, and activity of the drug in the patient. , sensitivity to the drug, time of administration, route of administration and excretion rate, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field.

The composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.

Specifically, the effective amount of the composition according to the present invention may vary depending on the patient's age, sex, and weight, and may increase or decrease depending on the route of administration, severity of disease, gender, weight, age, and the like.

In the present invention, the molar concentration ratio of the cyclophilin A inhibitor and afatinib may be 1:0.08 to 1.5. Specifically, the molar concentration ratio of the cyclophilin A inhibitor and afatinib may be 12.5:1 and 1:1.

In the present invention, the composition can kill non-small cell lung cancer stem-like cells.

In the present invention, the non-small cell lung cancer stem-like cells may be epidermal growth factor receptor (EGFR) mutants.

In another aspect, the present invention provides a cyclophilin A inhibitor or a salt thereof; And it provides a food composition for preventing or improving non-small cell lung cancer comprising afatinib or a salt thereof as an active ingredient.

The food composition is a concept including health functional food.

As used herein, the term "improvement" refers to all actions that at least reduce the parameters related to the condition to be treated, for example, the severity of symptoms.

In the food composition of the present invention, a cyclophilin A inhibitor or a salt thereof; And afatinib or a salt thereof can be added to food as it is or used together with other foods or food ingredients, and can be appropriately used according to conventional methods. Cyclophilin A inhibitor or salt thereof; And the mixing amount of afatinib or a salt thereof may be appropriately determined depending on the purpose of use (for prevention or improvement).

In general, when preparing food or beverage, the composition of the present invention is added in an amount of 15% by weight or less, preferably 10% by weight or less, based on the raw material. However, in the case of long-term intake for the purpose of health and hygiene or health control, the amount may be less than the above range.

The health functional food composition of the present invention is not particularly limited in other components other than containing the above components as essential components in the indicated proportions, and may contain various flavoring agents or natural carbohydrates as additional components like conventional beverages. there is.

Examples of the aforementioned natural carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose and the like; and polysaccharides such as conventional sugars such as dextrins, cyclodextrins, and the like, and sugar alcohols such as xylitol, sorbitol, and erythritol.

As flavoring agents other than those mentioned above, natural flavoring agents (thaumatin, stevia extract (eg rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can advantageously be used. The ratio of the natural carbohydrates can be appropriately determined by a person skilled in the art.

In addition to the above, the health functional food composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and its salts, Alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, and the like may be contained. These components may be used independently or in combination. The ratio of these additives can also be appropriately selected by those skilled in the art.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are intended to illustrate the present invention, and the scope of the present invention is not limited thereto.

<Example 1: Materials and Preparation>

1.1. experimental material

Cyclosporin A used in the experiment was purchased from Sigma-Aldrich, and afatinib was purchased from Tocris. Preparation and separation of Compound 9 are as described in the prior patent of the present inventor (Patent Application No. 10-2020-0102555). All drugs were dissolved at a concentration of 100 mM using DMSO, stored at -20 ° C, and used in experiments after thawing at room temperature during experiments. CellTiter-Glo® luminescent cell viability assay kit was purchased from Promega (Madison, WI, USA).

1.2. Non-small cell lung cancer stem-like cell culture

Non-small cell lung cancer cell lines A549 (EGFR Wild Type) and NCI-H1650 (EGFR Mutant Type) were purchased from Korea Cell Line Bank. The A549 and NCI-H1650 non-small cell lung cancer stem-like cells were supplemented with 1× B-27 serum-free supplement (Gibco), 5μg/mL heparin (Sigma-Aldrich), 2mM L-glutamine (Gibco), 20ng/mL epidermal growth factor (EGF; Prospecbio, East Brunswick, NJ, USA), 20ng/mL basic fibroblast growth factor (bFGF; Prospecbio, East Brunswick, NJ, USA), Dulbecco modified Eagle medium/nutrient with 1% penicillin/streptomycin (Gibco) It was cultured through spheroid culture using mixture F-12 (DMEM/F12; HyClone, Marlborough, MA, USA) medium. The cells were cultured at 37°C in a 5% CO2 cell incubator where appropriate humidity was maintained.

1.3. Cell viability assay (synthetic lethality-luminescence assay)

A549 and NCI-H1650 non-small cell lung cancer stem-like cells were dispensed in a 96-white well culture plate at a concentration of 3×10 ³ cells per well, and then treated with various concentrations of compounds alone or in combination, and cultured for 4 days. To measure cell viability, 20 μL of CellTiter-Glo® luminescent cell viability assay kit was added to each well, and the culture plate was shaken for 2 minutes and reacted for 8 minutes at room temperature in a dark place. Luminescence was measured using a multimode microplate reader (Biotek).

<Example 2: Confirmation of death of cancer stem cells by combined administration of cyclophilin A inhibitor and afatinib>

Cyclosporin A is known as an immunosuppressive agent that inhibits the activity of phosphatase of calcineurin by combining calcineurin and cyclophilin.

Compound 9 (23-demethyl 8,13-deoxynargenicin), a nargenisin A1 derivative, is a novel cyclophilin A inhibitor recently discovered by the present inventors.

The cyclosporin A inhibitor (cyclosporin A, compound 9) was applied to A549 non-small cell lung cancer stem cells and NCI-H1650 non-small cell lung cancer stem cells at 30 μM and 12.5 μM (compound 9) and 0.4 μM and 1 μM (cyclosporin A), respectively. 1 to 4 show the results measured by luminescence spectrometry after 4 days of treatment alone or in combination with afatinib at a concentration of 1 μM.

As shown in Figures 3 and 4, when combined treatment of compound 9 12.5μM and afatinib 1μM and cyclosporin A 1μM and afatinib 1μM in NCI-H1650 non-small cell lung cancer stem-like cells (EGFR mutant) Compared to the single treatment, cell survival was significantly suppressed, indicating a strong synthetic lethal interaction.

On the other hand, as shown in Figures 1 and 2, when combined treatment of compound 9 30μM and afatinib 1μM and cyclosporin A 0.4μM and afatinib 1μM to A549 non-small cell lung cancer stem-like cells (EGFR wild type) It can be seen that there is no significant difference compared to the single treatment.

Claims (10)

a cyclophilin A inhibitor or a pharmaceutically acceptable salt thereof; And a pharmaceutical composition for preventing or treating non-small cell lung cancer comprising afatinib or a pharmaceutically acceptable salt thereof as an active ingredient.
The pharmaceutical composition according to claim 1, wherein the cyclophilin A inhibitor is cyclosporin A or compound 9 represented by Formula 2 below.
[Formula 2]

The pharmaceutical composition according to claim 1, wherein the molar concentration ratio between the cyclophilin A inhibitor and afatinib is 1:0.08 to 1.5.
The pharmaceutical composition according to claim 1, wherein the composition kills non-small cell lung cancer stem-like cells.
The pharmaceutical composition according to claim 1, wherein the non-small cell lung cancer stem-like cells are epidermal growth factor receptor (EGFR) mutants.
a cyclophilin A inhibitor or a pharmaceutically acceptable salt thereof; And a food composition for preventing or improving non-small cell lung cancer comprising afatinib or a pharmaceutically acceptable salt thereof as an active ingredient.
The pharmaceutical composition according to claim 6, wherein the cyclophilin A inhibitor is cyclosporin A or compound 9 represented by Formula 2 below.
[Formula 2]

The food composition according to claim 7, wherein the molar concentration ratio of the cyclophilin A inhibitor and afatinib is 1:0.08 to 1.5.
The food composition according to claim 7, wherein the composition kills non-small cell lung cancer stem-like cells.
The food composition according to claim 7, wherein the non-small cell lung cancer stem-like cells are epidermal growth factor receptor (EGFR) mutants.

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