WO2019112243A1 - Composition containing interferon-gamma as active ingredient for prevention or treatment of neurofibroma - Google Patents

Abstract

The present invention relates to the treatment of neurofibromatosis type 1 malignancies. According to the present invention, combined use of interferon gamma with cisplatin or gemcitabine, on neurofibromatosis type 1 malignancies exhibits a proliferation-inhibitor effect, and thus can be utilized as an optimum anti-cancer composition which exhibits a synergistic effect with the anti-cancer action of cisplatin or gemcitabine when used for treatment of neurofibromatosis type 1 malignancies. Furthermore, interferon gamma can be used as an ancillary treatment agent for reducing side effects by decreasing the dose of the anti-cancer agents cisplatin and gemcitabine.

Description

Composition for preventing or treating nerve fibrosis containing interferon gamma as an active ingredient

The present invention relates to the treatment of malignant tumors of type 1 neurofibromatosis, and more particularly, to a method of treating malignant tumors of type 1 neurofibromatosis, .

Neurofibromatosis Type 1 (NF1) was first described by Fredrich von Recklin-ghausen in 1882 and is an autosomal dominant genetic disorder that forms tumors along the peripheral nerves. It occurs in about 1 in 3,500 patients. The tumor is called neurofibromatosis because it can occur in any part of the nervous body, and the number is more than one. The main symptoms are neurofibroma and cafe-au-lait spots, Lisch nodules, optic pathway glioma, scoliosis, Tibial dysplasia, plexiform neurofibroma, pontine glioma, etc. (see Fig. 1 of the present invention).

The most common type 1 neurofibromatosis patient is a neurofibroma, a benign tumor that occurs in the skin. About 30-40% of patients with type 1 neurofibromatosis have gunshot nerve fibromatosis, a benign tumor that occurs under the skin or in the deep part of the body. In particular, gunshot neurofibromas show a massive lump (lump) shape that grows sideways and spreads sideways in various body parts (see FIG. 2 of the present invention), and a large number of multiple gunshot neurofibromas develop in the deep part 3 of the present invention). Although plexiform neurofibroma (PN) is a benign tumor, it develops very seriously in patients, and in about 10% of patients with type 1 neurofibromatosis, neurofibromatosis is a malignant neurofibroma neurofibrosarcoma, malignant peripheral nerve sheath tumor (MPNST) (see FIG. 4 of the present invention). In patients with type I neurofibromatosis, the malignant progression of benign tumors develops rapidly, resulting in a very high mortality rate.

(PN) stimulates the growth of bone and surrounding tissues and grows to a large size, but it may also appear to be a skewed appearance, and about 8 to 13% of such patients are malignant with neurofibromatosis (MPNSTs) 5). MPNST is a highly malignant metastatic cancer that is known to have a high mortality rate and low response to chemotherapy and radiation.

Type 1 neurofibromatosis is caused by a mutation in the NF1 gene, and about 50% of the patients are caused by naturally occurring mutations that are not hereditary. The NF1 gene is located on chromosome 17q11.2, and consists of 350 kb genomic DNA and 57 exons. The mRNA is about 11 to 13 kb. The tumor suppressor gene NF1 encodes a neurofibromin protein in the cytoplasm of ~ 320 kDa that encodes a GAP-related protein that negatively regulates the Ras signal at amino acid 1125-1537 Domain (GAP-related domain). GTPase-activating protein (GAP) modulates the activity of Ras by stimulating intrinsic GTPase of Ras and converting it into an inactive form of Ras-GDP in the activated form, Ras-GTP. Oncogene The Ras protein, which is encoded by Ras, is expressed in both neuronal and endothelial cells, and the signal is transmitted downstream (downstream). Increased GTP-Ras induces signal transduction through a representative downstream pathway, Raf kinase, and Raf induces signaling through kinase kinase including Erk1 and Erk2 isoforms of MEK kinase and mitogen-activated protein (MAP) kinase (See FIG. 6 of the present invention) by activating the cell.

There are three types of Ras genes: K-Ras, H-Ras, and N-Ras. Mutations in the Ras gene are found in about 50% of human cancers. Mutation of the Ras gene and excessive activity of the Ras protein are involved in the carcinogenesis of the cell, and thus research on the cancer treatment through suppression of the intracellular expression of the Ras protein has been actively carried out. It is known that all three Ras proteins, K-Ras, H-Ras and N-Ras, are overactivated by the mutation of the NF1 gene. However, recent studies have shown that astrocytes in patients with type I neurofibromatosis have H- Ras and N-Ras are not activated, only K-Ras is specifically activated, and hematopoietic cells are activated only by H-Ras.

20% of NF1 patients with overactive and accumulating RAS due to neurofibromin loss or dysfunction cause severe life-threatening complications and about 8 to 13% And the clinical symptoms are severe, but inherited, so it is impossible to treat it. In the case of malignant neurofibrosarcoma, only mass therapy such as resection of the tumor is the only treatment, or surgery is impossible according to the location of the tumor, and it is directly related to the life of the patient regardless of the timing of the cancer. The exact mechanism of the variability and malignancy of the clinical phenotype of type 1 neurofibromatosis remains unclear. Traditional therapies include radiotherapy, surgical resection, and cytotoxic drugs, but no long-term effects were found.

It is an object of the present invention to provide an adjuvant for enhancing the nerve fiber sarcoma anticancer effect.

Another object of the present invention is to provide an anticancer composition for preventing or treating nerve fibrosis.

In order to achieve the above object, the present invention provides an adjuvant for enhancing neurofibrosarcoma (MPNST) anti-cancer effect comprising interferon-gamma as an active ingredient.

In order to accomplish the above object, the present invention provides a neurofibrosarcoma (hereinafter referred to as " neurofibrosarcoma ", hereinafter referred to as " malignant peripheral nerve sheath tumor ") comprising an interferon-gamma and / or cisplatin or gemcitabine as an active ingredient. MPNST) for the prevention or treatment of cancer.

The present invention relates to a composition for preventing or treating neurofilament sarcoma comprising interferon gamma and an adjuvant for enhancing anticancer efficacy which reduces the dose of an anticancer agent when used in combination with cisplatin and gemcitabine anticancer agent and interferon gamma. Since the above-mentioned interferon-gamma is included, the anticancer effect of cisplatin and gemcitabine in the type 1 neurofibromatous malignant tumor cells is maximized and the anticancer effect is more remarkable than that in the cases where each of them has a synergistic effect, The composition can effectively prevent, ameliorate, or treat malignant tumors of type 1 neurofibromatosis.

Figure 1 shows representative symptoms of type 1 neurofibromatosis (A) milk-cough-au-lait spots, (B) neurofibroma, (C) Lisch nodules, (D) optic pathway glioma, (E) scoliosis, (F) tibial dysplasia, (G) plexiform neurofibroma, (H) glioma).

Figure 2 shows gunshot neurofibroma occurring in various body parts.

FIG. 3 shows an example of a gunshot neurofibroma occurring in the deep part.

Fig. 4 shows an example of nerve fibrosarcoma developed malignantly in benign bullet neurofibroma.

Figure 5 shows the process of tumor formation and malignancy of type 1 neurofibromatosis.

Fig. 6 schematically shows the role of neurofibromin and the mechanism of Ras activation.

FIG. 7 shows the results of confirming the synergistic effect of interferon gamma on NF1 malignant tumor Schwannoma cell line (S462) when cell antigens were treated with 5 kinds of anticancer drugs alone or in combination with interferon gamma.

Fig. 8 shows the results of confirming the apoptosis effect of NF1 malignant tumor Schwann cell line (S462) when treated with cisplatin and gemcitabine alone and with interferon gamma.

Neurofibromatosis Type 1 (NF1) was first described by Fredrich von Recklin-ghausen in 1882 and is an autosomal dominant genetic disorder that forms tumors along the peripheral nerves. It occurs in about 1 in 3,500 cases. Malignant tumors present in about 8 to 13% of patients are unpredictable, have severe clinical symptoms and high mortality rates, but are difficult to treat due to the inability to perform surgery depending on the location of the tumor. The efficacy of anticancer drugs is low and recurrence rates are high.

Accordingly, the inventors of the present invention have confirmed that the combination therapy of interferon gamma and an anticancer agent is effective for inhibiting the proliferation and treatment of malignant tumors of type 1 neurofibromatosis, and completed the present invention.

Accordingly, the present invention provides an adjuvant for enhancing neurofibrosarcoma (MPNST) anti-cancer effect containing interferon-gamma as an active ingredient.

That is, when the interferon gamma is administered in combination with an anticancer agent, the anticancer effect can be maximized or the administration dose of the anticancer agent can be minimized.

Preferably, the interferon gamma is most effective in enhancing the anticancer effect when the cisplatin or gemcitabine is coadministered with the anticancer agent.

Further, the present invention provides a method for preventing or treating neurofibrosarcoma (MPNST), which comprises a) interferon-gamma and b) an anticancer agent selected from cisplatin or gemcitabine as an active ingredient To provide a pharmaceutical composition.

Preferably, the composition may contain interferon gamma, cisplatin and gemcitabine as an active ingredient.

It is known that the neurofilament sarcoma is caused by malignant transformation of the bulbar neurofibroma, and the neurofibrosarcoma is caused by malignant development in about 8-13% of the patients with bulbar neurofibroma.

The composition inhibits cell proliferation of neurofilament sarcoma, increases cell death, does not cause cytotoxicity to normal cells, has few side effects, and has excellent therapeutic effect on nerve fibrosarcoma.

Further, by using interferon gamma in combination with a specific combination of anticancer agents, the effective dose of the anticancer agent can be remarkably reduced to thereby reduce adverse effects in the treatment of cancer, and when used in combination with conventional anticancer agents of the same amount, Lt; / RTI >

When the composition of the present invention is a pharmaceutical composition, for administration, it may contain a pharmaceutically acceptable carrier, excipient or diluent in addition to the above-mentioned effective ingredient. Examples of the carrier, excipient and diluent include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, Polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

The pharmaceutical composition of the present invention may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols or the like, oral preparations, suppositories or sterilized injection solutions according to conventional methods . In detail, when formulating, it can be prepared using diluents or excipients such as fillers, weights, binders, humectants, disintegrants, surfactants and the like which are usually used. Solid form preparations for oral administration include, but are not limited to, tablets, pills, powders, granules, capsules and the like. Such a solid preparation may be prepared by mixing at least one excipient such as starch, calcium carbonate, sucrose, lactose, gelatin and the like in addition to the active ingredient. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid preparations for oral administration, liquid paraffin, and various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations and tasks. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, and the like. As a base for suppositories, it is possible to use witepsol, macrosole, tween 61, cacao paper, laurin, glycerogelatin and the like.

The appropriate dose of the pharmaceutical composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the type of drug, and the time, but can be appropriately selected by the person skilled in the art. 0.001 mg / kg to 50 mg / kg, and may be administered once to several times per day as needed.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail with reference to the following examples. It is to be understood that both the foregoing description and the following detailed description are exemplary and explanatory and are intended to provide further explanation of the invention as claimed. no.

Example 1: Cell culture

Human cell (HSC) (Scientific Research Laboratories) and S462 (American Type Culture Collection, USA), which is a neurofibrosarcoma (malignant peripheral nerve sheath tumor, MPNST) cell, were cultured in 10% fetal bovine serum (Dulbecco's modified Eagle's medium (Hyclone Laboratories, Logan, USA) supplemented with FBS, Hyclone Laboratories, Logan, USA), penicillin (100 U / ml) and streptomycin (100 μg / ml) And cultured in a CO ₂ incubator at 37 ° C.

Example 2: Measurement of cell viability

Cell viability was analyzed using the D-Plus CCK cell viability assay kit (Donglin LS, Korea). Cells were seeded at a cell density of 7 x 10 < 3 > cells per well in 96 well plates and cultured for 48 hours after drug treatment. Subsequently, the culture medium was replaced with 100 μl of a culture medium supplemented with 10 μl of CCK reagent and further cultured for 2 hours. Cell viability was determined by measuring the absorbance at 450 nm using an ELISA plate reader (Bio-Rad Model 680).

Example 3: Statistical analysis

Student's t-test was used for significance test of in vitro graph (significance level: <0.05).

Experimental Example 1: Combination treatment of interferon gamma and anticancer agent

Cisplatin (2 μg / ml), gemstabine (1 μM), methotrexate (1 μM), and cisplatin (2 μg / ml), which are commonly used for malignant sarcoma in NF1 malignant tumor Schwann cell line (S462) ), Paclitaxel (50 μM), Vinblastine (100 nM) alone or in combination with interferon gamma (at a concentration of 1,000 U / ml) and analyzing cell viability according to Example 2 And the synergy effect of cancer was observed.

As a result, as shown in Fig. 7, when cisplatin or gemcitabine was used in combination with interferon-gamma, the synergistic effect of promoting apoptosis of NF1 malignant tumor Schwann cell line (S462) was shown, while mesotrexate, paclitaxel, In the case of the blastin anticancer agent, there was no synergistic effect due to the interferon-gamma treatment. In the treatment of normal schwann cell line (HSC), cytotoxicity of cisplatin, gemcitabine, methotrexate, and vinblastine anticancer was not observed.

Accordingly, the interferon-gamma according to the present invention appears to be synergistic only when it is administered together with a specific anticancer drug such as cisplatin or gemcitabine. In the subsequent experiments, synergistic effects with cisplatin and gemcitabine were also tested.

Experimental Example 2: Confirmatory effect of interferon gamma on cisplatin and gemcitabine anticancer agent

Cefplatin and gemcitabine alone or in combination with interferon-gamma were administered to NF1 malignant tumor Schwann cell line (S462) and normal Schwann cell line (HSC), and cell viability of the cells was confirmed according to Example 2 above. At this time, cisplatin and gemcitabine were treated at a concentration of 0.5 μg / ml and 500 nM, respectively, and interferon gamma was treated at a concentration of 1,000 U / ml. In addition, cisplatin and gemcitabine were treated at a concentration of 0.25 μg / ml and 250 nM, respectively.

As a result, as shown in FIG. 8, when cisplatin or gemcitabine was used in combination with interferon-gamma, there was synergistic effect of promoting apoptosis of NF1 malignant tumor Schwann cell line (S462), and cisplatin, gemcitabine and interferon gamma The same anticancer effect was obtained even when the amount of cisplatin and gemcitabine was reduced to 1/2. On the other hand, normal cisplatin (HSC) did not show cytotoxic toxicity when treated with cisplatin, gemcitabine or interferon gamma.

Therefore, interferon gamma, when used with cisplatin and gemcitabine, has a synergistic effect on the anticancer effect, and it is expected to have a good effect in reducing the side effects of anticancer drugs by reducing the use of cisplatin and gemcitabine.

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the present invention is not limited thereby. Do. That is, the practical scope of the present invention is defined by the appended claims and their equivalents.

Claims (2)

1. A pharmaceutical composition for the prophylaxis or treatment of neurofibrosarcoma (MPNST) comprising a) interferon-gamma and b) cisplatin or gemcitabine as an active ingredient.
2. The pharmaceutical composition for preventing or treating neurofibromatosis according to claim 1, wherein the neurofilament sarcoma is caused by malignant transformation of neurofibromatosis.

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