KR20200097042A - Composition for preventing or treating of cancer comprising manassantin A and EGFR inhibitor - Google Patents

Abstract

The present invention relates to a composition for preventing and/or treating cancer comprising manassantin A and an epidermal growth factor receptor inhibitor as active ingredients. The composition of the present invention can be expected to be effectively used to suppress resistance to chemotherapy, recurrence, metastasis or proliferation of various solid cancers through combined administration.

Description

Composition for preventing or treating cancer comprising manassantin A and an epithelial growth factor receptor inhibitor {Composition for preventing or treating of cancer comprising manassantin A and EGFR inhibitor}

The present invention relates to a pharmaceutical composition for preventing or treating cancer, including manasanthin A and an epidermal growth factor receptor inhibitor (EGFR inhibitor) as an active ingredient.

Manassantin A is a type of lignan isolated from Saururus chinensis (Lour.) Baill, one of the medicinal plants, and has the effect of preventing or treating non-alcoholic fatty liver, anti-inflammatory, and anti-human immunodeficiency. It has been found that there is a therapeutic effect on various diseases such as viral effect and hyperpigmentation disease improvement effect, and research on this has been actively conducted recently, but the relationship between manasanthin A and cancer is still insufficient.

On the other hand, since malignant advanced tumor cells proliferate faster than endothelial cells forming blood vessels, the newly formed blood vessels are not normally distributed inside the cancer tissue, so sufficient blood cannot be supplied, resulting in a lack of nutrients and acidification. , Oxygen deficiency, etc. are induced, and such tumor microenvironment is known to affect the therapeutic effect of cancer. In practice, the median value of the partial pressure of oxygen in normal tissues is 40-60 mmHg, but in the case of solid cancer, the median value of the partial pressure of oxygen is less than 10 mmHg in most cases. It is known that cancer cells are adapted to such hypoxic conditions, and thus cancer cells become more malignant under hypoxic conditions, and become resistant to various cancer treatment therapies such as anticancer drug therapy and radiation therapy.

The present inventors have studied the relationship between manasanthin A and cancer. As a result, manasanthin A relieves the microenvironment inside solid cancer and at the same time inhibits the proliferation of cancer cells, thereby showing the effect of preventing and/or treating cancer. , It was confirmed that the therapeutic effect was significantly increased when the combination administration of manasanthin A and an epithelial growth factor receptor inhibitor, a therapeutic agent for lung cancer, was significantly increased, and the present invention was completed.

Domestic patent application 10-2015-0106051

The present invention was conceived to solve the problems of the prior art as described above, and comprises manasanthin A and an EGFR inhibitor capable of alleviating hypoxic conditions inside cancer tissues and inhibiting the proliferation of cancer cells as active ingredients. An object thereof is to provide a composition for preventing or treating cancer.

However, the technical task to be achieved by the present invention is not limited to the above-mentioned tasks, and other tasks that are not mentioned can be clearly understood by those of ordinary skill in the technical field to which the present invention belongs from the following description. will be.

The present invention provides a pharmaceutical composition for preventing or treating cancer comprising manassantin A and an epidermal growth factor receptor inhibitor (EGFR inhibitor) as an active ingredient.

In an embodiment of the present invention, the epidermal growth factor receptor inhibitor is preferably gefitinib, erlotinib, afatinib, brigatinib, and icotinib. ) May be any one or more selected from the group consisting of, but is not limited thereto as long as it is an epithelial growth factor receptor inhibitor used as a lung cancer treatment.

In another embodiment of the present invention, manasanthin A may exhibit a therapeutic effect on cancer by inhibiting the expression of a hypoxia-inducible factor 1α (HIF-1α; Hypoxia-inducible factor-1 alpha) protein in cancer cells.

In another embodiment of the present invention, the pharmaceutical composition is characterized in that it suppresses the proliferation, metastasis, recurrence, etc. of cancer, or suppresses resistance to anticancer therapy, but is part of a generally used cancer treatment method. If yes, it is not limited thereto.

In another embodiment of the present invention, the cancer is lung cancer, breast cancer, glioma cancer, colon cancer, uterine cancer, ovarian cancer, prostate cancer, gastric cancer, brain tumor, multiple myeloma, pediatric cancer, rectal cancer, colon cancer, thyroid cancer, oral cancer, pharyngeal cancer , Laryngeal cancer, bladder cancer, kidney cancer, liver cancer, pancreatic cancer, bone cancer, skin cancer, etc., or if the type of solid cancer is not limited thereto.

In addition, the present invention provides a method for treating cancer, comprising administering the composition to an individual.

In addition, the present invention provides the use of the composition for cancer treatment.

Manasanthin A according to the present invention inhibits the expression of HIF-1α inside the solid cancer and, through this, not only alleviates microenvironmental requirements such as hypoxic conditions, but also suppresses the proliferation of cancer cells. / Or it can be used for treatment, and its effect can be remarkably improved through co-administration with an epithelial growth factor receptor inhibitor such as gefitinib, so it is expected that it can be effectively used for various cancer treatments.

1 is a view showing a result of confirming the effect of manasanthin A on a tumor according to an embodiment of the present invention.
2 is a view showing the results of confirming the effect of co-administration of manasanthin A and an epithelial growth factor receptor inhibitor according to an embodiment of the present invention on a tumor.

The present inventors confirmed that manasanthin A can be used for the treatment of cancer by effectively inhibiting the expression of HIF-1α, thereby alleviating the microenvironmental requirements of solid cancer and inhibiting the proliferation of cancer cells. The present invention was completed by confirming that the cancer treatment effect can be remarkably improved by using it with a growth factor receptor inhibitor.

In the present specification, "manassantin A" refers to a compound having a structure such as the following formula (1), and has a non-alcoholic liver disease treatment effect, anti-inflammatory, anti-human immunodeficiency virus effect, hyperpigmentation Various biological activities have been reported such as disease improvement effect.

[Formula 1]

In the present specification, the term "epidermal growth factor receptor inhibitor (EGFR inhibitor)" is an anticancer agent targeting an epidermal growth factor receptor, and by binding a transforming growth factor, a ligand, etc. to the epidermal growth factor receptor, As a kind of anticancer agent that treats cancer by inhibiting the mechanism by which a number of intracellular mechanisms are activated, and through which the proliferation of tumor cells occurs, preferably gefitinib, erlotinib, afatinib, brigatinib, ico It is not limited thereto as long as it is a substance capable of inhibiting activation of an epidermal growth factor receptor, such as tinib.

In the present specification, “prevention” refers to any action that suppresses or delays onset of diseases such as cancer by administration of the composition according to the present invention.

In the present specification, "treatment" refers to any action in which symptoms such as cancer are improved or advantageously changed by administration of the composition according to the present invention.

In the present specification, “individual” refers to a subject to which the composition of the present invention can be administered, and the subject is not limited.

In the present specification, the term "pharmaceutical composition" may be characterized in that it is in the form of capsules, tablets, granules, injections, ointments, powders or beverages, and the pharmaceutical composition is intended for humans. I can. The pharmaceutical compositions are not limited thereto, but may be formulated and used in the form of oral dosage forms such as powders, granules, capsules, tablets, aqueous suspensions, etc., external preparations, suppositories, and sterile injectable solutions according to conventional methods. . The pharmaceutical composition of the present invention may contain a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers can be used as binders, lubricants, disintegrants, excipients, solubilizers, dispersants, stabilizers, suspending agents, colors, flavors, etc. for oral administration, and buffers, preservatives, painlessness, etc. for injections. Agents, solubilizers, isotonic agents, stabilizers, etc. can be mixed and used, and in the case of topical administration, base agents, excipients, lubricants, preservatives, and the like can be used. The formulation of the pharmaceutical composition of the present invention can be variously prepared by mixing with a pharmaceutically acceptable carrier as described above. For example, for oral administration, it can be prepared in the form of tablets, troches, capsules, elixir, suspension, syrup, wafers, etc.In the case of injections, it can be prepared in the form of unit dosage ampoules or multiple dosage forms. have. Others, solutions, suspensions, tablets, capsules, can be formulated as sustained-release preparations.

On the other hand, examples of carriers, excipients and diluents suitable for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, malditol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate or mineral oil may be used. In addition, fillers, anti-aggregating agents, lubricants, wetting agents, flavoring agents, emulsifying agents, preservatives, and the like may additionally be included.

The route of administration of the pharmaceutical composition according to the present invention is not limited thereto, but oral, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, local , Sublingual or rectal. Oral or parenteral administration is preferred. The term “parenteral” as used herein includes subcutaneous, intradermal, intravenous, intramuscular, intraarticular, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection or infusion techniques. The pharmaceutical composition of the present invention can also be administered in the form of suppositories for rectal administration.

The pharmaceutical composition of the present invention depends on a number of factors including the activity of the specific compound used, age, weight, general health, sex, formulation, time of administration, route of administration, excretion rate, drug formulation and the severity of the specific disease to be prevented or treated. It may vary in various ways, and the dosage of the pharmaceutical composition varies depending on the patient's condition, weight, degree of disease, drug form, administration route and duration, but may be appropriately selected by those skilled in the art, and 0.0001 to 50 mg per day /kg or 0.001 to 50 mg/kg. Administration may be administered once a day or may be divided several times. The above dosage does not limit the scope of the present invention in any way. The pharmaceutical composition according to the present invention may be formulated as a pill, dragee, capsule, liquid, gel, syrup, slurry, or suspension.

Hereinafter, preferred embodiments are presented to aid in understanding the present invention. However, the following examples are provided for easier understanding of the present invention, and the contents of the present invention are not limited by the following examples.

[ Example ]

Example One: Manasanthin Confirmation of anticancer effect of A

To confirm the effect of manassantin A on cancer, Lewis lung carcinoma (LLC) 4 x 10 ⁵ cells, a mouse lung cancer cell line, were allografted subcutaneously to the flank of 6-7 weeks old C57BL/6J mice. After preparing the tumor animal model, 2 mg/kg of manasanthin A was injected intraperitoneal on days 2, 4, 6, 8, and 10. As a control, an equal amount of phosphate buffered saline (PBS) was injected. In order to measure the hypoxic state inside the tumor on the 17th day, 60 mg/kg of Hypoxyprobe-1™ (pimonidazole hydrochloride; PIMO, HPI) was administered by intraperitoneal injection, and 90 minutes later, the mice were sacrificed to extract the tumor. And IHC technique (Immunohistochemistry analysis) was performed using the obtained tumor tissue. In more detail, the obtained tumor was fixed with 3.7% paraformaldehyde (PFA) and then dehydrated by stepwise treatment with a 10-40% sucrose solution. And, after treatment with 0.3% Triton X-100 to increase the osmoticity of cells, and after blocking process using 5% bovine serum albumin (BSA), rat anti-CD31 (BD Biosciences) or Alexa Fluor 647 -Conjugated anti-HIF-1 alpha antibody (Abcam) was treated and reacted at 4°C for 16 hours. Mice treated with HIF-1 alpha antibody were mice not treated with Hypoxyprobe-1™. And after washing unbound antibody with phosphate buffer solution, treatment with Alexa Fluor 488-conjugated anti-rat IgG (Invitrogen) antibody and mouse anti-hypoxyprobe antibody (HPI), which are secondary antibodies to which fluorescent labels are bound. And reacted at room temperature for 1 hour. And finally, after staining the cell nucleus by treatment with DAPI (Sigma-Aldrich), it was observed using a confocal microscope, and fluorescence values were quantified using ImageJ. All experimental results were at least three, and ANOVA verification was performed to indicate meaningful differences between the groups, and only values with statistical significance with P-values <0.05 were expressed as mean values and ± standard deviations (SD). The results are shown in FIG. 1.

As shown in FIG. 1, the hypoxic state generally observed inside of solid cancer was significantly reduced in the experimental group administered with manasanthin A, and the expression level of HIF-1α (Hypoxia-inducible factor-1 alpha) was also significantly reduced. I could confirm. In addition, it was confirmed that the microvessel density (MVD) of CD31-positive vascular endothelial cells was significantly reduced in the experimental group administered with manasanthin A. Through the above results, it was confirmed that manasanthin A can effectively inhibit the growth and metastasis of solid cancer by inhibiting the expression of HIF-1α, which is known as an important transcription factor related to angiogenesis and anticancer drug resistance in solid cancer.

Example 2: Confirmation of the effect of co-administration of manasanthin A and gefitinib

In order to confirm whether an anticancer effect was exhibited without side effects when the combination of epithelial growth factor receptor inhibitor (EGFR inhibitor) gefitinib and manasanthin A was administered, a tumor animal model was prepared in the same manner as in Example 1, 5 mg/kg of manasanthin A and 5 mg/kg of gefitinib were injected once every 3 days for a total of 5 times (intraperitoneal injection). As a control, an equal amount of phosphate buffered saline (PBS) was injected. In addition, tumor growth and toxicity were observed at intervals of 2 days from the 8th day when the tumor was touched on the skin of the mouse. The volume of the tumor was measured by measuring the width (mm) X length (mm) X height (mm) with a Caliper, and multiplying it to calculate the volume (mm ³ ), and toxicity was confirmed by measuring the weight of the mouse. And on the 17th day, the tumor was excised, weighed, and a picture was obtained. Animal breeding and all experimental procedures were conducted in accordance with the laws and regulations for animal experiments. All experimental results were at least three, and ANOVA verification was performed to indicate meaningful differences between the groups, and only values with statistical significance with P-values <0.05 were expressed as mean values and ± standard deviations (SD). The results are shown in FIG. 2.

As shown in FIG. 2, in the case of the experimental group administered with manasanthin A and gefitinib, it was confirmed that the size of the tumor was significantly reduced compared to the experimental group administered with each of them alone, and there was no change in weight, so low toxicity. It was confirmed that the treatment effect increased by more than 3 times. Through the above results, it was confirmed that a high therapeutic effect can be exhibited with low side effects through the co-administration of manasanthin A and gefitinib.

Through the above results, the composition containing manasanthin A and an epithelial growth factor receptor inhibitor of the present invention inhibits the expression of HIF-1α in solid cancer by manasanthin A, thereby reducing microenvironmental requirements such as hypoxic conditions. In addition, it was confirmed that by increasing the therapeutic effect of an epithelial growth factor receptor inhibitor such as gefitinib in solid cancer, the therapeutic effect due to co-administration can be significantly increased. Accordingly, it is expected that the composition of the present invention can be effectively used to inhibit the proliferation, metastasis, recurrence, or resistance to anticancer therapy of solid cancer.

The above description of the present invention is for illustrative purposes only, and those of ordinary skill in the art to which the present invention pertains will be able to understand that other specific forms can be easily modified without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative and non-limiting in all respects.

Claims (5)

Manasantin A (manassantin A) and epidermal growth factor receptor inhibitor (epidermal growth factor receptor inhibitor) containing as an active ingredient, cancer prevention or treatment pharmaceutical composition. The method of claim 1,
The epidermal growth factor receptor inhibitor is characterized in that at least one selected from the group consisting of gefitinib, erlotinib, afatinib, brigatinib, and icotinib. To, pharmaceutical composition. The method of claim 1,
The manasanthin A is characterized in that inhibiting the expression of HIF-1α (Hypoxia-inducible factor-1 alpha) protein in cancer cells, pharmaceutical composition. The method of claim 1,
The composition is characterized in that for inhibiting the proliferation, metastasis, recurrence or resistance to anticancer therapy of cancer, pharmaceutical composition. The method of claim 1,
The cancer is lung cancer, breast cancer, glioma cancer, colon cancer, uterine cancer, ovarian cancer, prostate cancer, stomach cancer, brain tumor, multiple myeloma, childhood cancer, rectal cancer, colon cancer, thyroid cancer, oral cancer, pharyngeal cancer, laryngeal cancer, bladder cancer, kidney cancer, liver cancer, A pharmaceutical composition, characterized in that at least one selected from the group consisting of pancreatic cancer, bone cancer, and skin cancer.

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