KR20140032586A - A pharmaceutical composition for radiation therapy of egfr-tki-resistant lung cancer caused by pten function deficiency - Google Patents

Abstract

The present invention relates to an epithelial growth factor receptor tyrosine kinase inhibitor caused by PTEN dysfunction including a mTOR (mammalian target of rapamycin) inhibitor or a pharmaceutically acceptable salt, solvate, or hydrate thereof as a radiation sensitive agent. Receptor-Tyrosine Kinase Inhibitor (EGFR-TKI) to provide a pharmaceutical composition for the radiation treatment of lung cancer.

Description

A pharmaceutical composition for radiation therapy of EGFR-TKI-resistant lung cancer caused by PTEN function deficiency}

The present invention is for the radiation therapy of epithelial growth factor receptor tyrosine kinase inhibitor (hereinafter, abbreviated as "EGFR-TKI") resistant lung cancer due to PTEN (phosphatase and tensin homolog) function The present invention relates to a pharmaceutical composition for radiation therapy, comprising a drug that can act as a radiosensitizer for the treatment of EGFR-TKI resistant lung cancer due to PTEN hypofunction, more specifically to treat EGFR-TKI resistant lung cancer. It is about.

Gefitinib is an anticancer agent used for the treatment of breast cancer, lung cancer, and other cancers known under the trade name Iressa. Zephytinib functions as an epithelial growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) and serves to inhibit signaling through EGFR in target cells. Thus, it is effective in the treatment of cancers that have mutated and overactive EGFR (hereinafter also referred to as "activated mutant EGFR"). As such, EGFR-TKI drugs that act as EGFR-TKI and are known to be effective in the treatment of cancers with EGFR activation mutations include zefitinib, erlotinib, and the like.

EGFR-TKIs, including Zephytinib, are known to be effective in the treatment of lung cancers, particularly non-small cell lung cancers, with activating variant EGFR. Lung cancers with EGFR activation mutations show more than 70% response to the EGFR-TKI, while lung cancers without EGFR activation mutations show less than 1% response rate. Lung cancer patients with EGFR activating mutations in the lung cancer group are predominantly female, non-smokers and Asians, while lung cancer patients without EGFR activating mutations are known in males, smokers and Westerners.

Therefore, EGFR-TKI, including gefitinib, is a very important drug for the treatment of lung cancer. However, the use of EGFR-TKI in lung cancer patients with EGFR activating mutations is highly effective but results in acquired resistance after an average of 6-8 months in almost all patients. Patients with acquired resistance to EGFR-activated mutant lung cancer do not have any other appropriate chemotherapy and radiation therapy is ineffective, so there is little effective treatment to date. To date, the mechanisms of acquisition tolerance have been identified, including EGFR T790 gatekeeper mutation and MET oncogene amplification. Recently, it is known that drug resistance can occur even by PTEN loss mutations (Non-Patent Documents 1 and 2).

Since the acquisition of this resistance in lung cancer with EGFR activating mutations is very fatal, methods of treating EGFR-TKI resistant acquisition lung cancer are very urgent.

On the other hand, mTOR (mammalian target of rapamycin) inhibitor is a drug that stops the cell cycle while inhibiting mTOR, which is a lower target of PI3K / Akt signaling system that mediates drug resistance and migration, and rapamycin is a representative drug. It is used to suppress rejection, but also works on several tumors. Rapamycin forms a conjugate with the intracellular protein FKBP12 (FK-binding protein 12) and then directly binds to mTOR Complex 1 (mTORC1) to inhibit mTOR activity. Everolimus (RAD001) is a rapamycin derivative that is an mTOR inhibitor and also forms a conjugate with FKBP12, an intracellular protein, which inhibits mTOR activity. Inhibition of mTOR activity by rapamycin or everolimus treatment inhibits phosphorylation of p70 S6 ribosomal protein kinase. Subsequently, phosphorylation of S6 is inhibited, resulting in inhibition of protein synthesis and cell proliferation.

1.Reduction of PTEN protein and loss of epidermal growth factor receptor gene mutation in lung cancer with natural resistance to gefitinib (IRESSA) Br J Cancer. 2005 May 9; 92 (9): 1711-9. 2.Loss of PTEN expression by blocking nuclear translocation of EGR1 in gefitinib-resistant lung cancer cells harboring epidermal growth factor receptor-activating mutations. Cancer Res. 2010 Nov 1; 70 (21): 8715-25.

It is an object of the present invention to provide a pharmaceutical composition capable of treating EGFR-TKI resistant lung cancer caused by various mechanisms, in particular EGFR-TKI resistant lung cancer caused by PTEN hypofunction.

In order to achieve the above object, an aspect of the present invention provides a pharmaceutical composition for radiotherapy of EGFR-TKI resistant lung cancer by PTEN hypofunction, including a mTOR (mammalian target of rapamycin) inhibitor as a radiosensitizer.

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

All technical terms used in the present invention are used in the sense that they are generally understood by those of ordinary skill in the relevant field of the present invention unless otherwise defined. Also, preferred methods or samples are described in this specification, but similar or equivalent ones are also included in the scope of the present invention. The contents of all publications cited herein are hereby incorporated by reference in their entirety.

The present inventors studied a method for treating lung cancer (hereinafter referred to as "EGFR-TKI resistant lung cancer") that acquired resistance to EGFR-TKI in lung cancer with EGFR activating mutations.

Specifically, after constructing a cell-level model for patients with zephytinib-resistant lung cancer induced by PTEN expression degradation, the model was used to test the effect of radiation therapy on patients with zephytinib-resistant lung cancer. In order to construct a PTEN hypothesis-inducing zephytinib resistance model, in this study, the loss of 746-750 amino acids of EGFR protein by EGFR partial loss mutation resulted in abnormal activation of EGFR, thereby making it sensitive to EGFR-TKI. HCC827, a lung cancer cell line known to be a cell line, was used. HCC 827 cells carry normal PTEN genes. In this study, the lentiviral into which short hairpin RNA (shRNA) into which PTEN mRNA was targeted was infected with HCC827 to decrease PTEN expression of HCC827 cells (FIG. 1A). HCC827 cells with reduced PTEN expression were irradiated with zefitinib at various concentrations. After 72 hours, the cell survival was examined by the MTT method. As a result, it was confirmed that zephytinib resistance was obtained by PTEN expression reduction (FIG. 1B). Thus, a cell-level model for gefitinib resistance acquired lung cancer patients was established. Using the established zephytinib resistance model, we investigated the effects of radiation therapy on patients with zephytinib-resistant lung cancer caused by PTEN expression. As a result, PTEN expression was not only resistant to zephytinib (FIG. 1B) but also to radiation therapy. It has also been shown to impart resistance to (FIG. 1C). As a result of the administration of mTOR inhibitor rapamycin or everolimus alone to the zephytinib resistance model, resistance to rapamycin and everolimus was shown regardless of the expression of PTEN. However, when the mTOR inhibitor was administered with radiation, it showed a significant increase in radiation treatment efficiency for the zephytinib resistance model (B of FIG. 2 and B of FIG. 3).

Thus, in one aspect, the present invention comprises a mammalian target of rapamycin (mTOR) inhibitor or a pharmaceutically acceptable salt, solvate, or hydrate thereof as a radiosensitizer, EGFR-TKI resistance by PTEN hypofunction Provided is a pharmaceutical composition for radiotherapy of lung cancer.

As used herein, the term "EGFR-TKI resistant lung cancer" refers to lung cancer in which lung cancer, which had a therapeutic effect on EGFR-TKI, obtained resistance to EGFR-TKI as a result of treatment of EGFR-TKI. The EGFR-TKI is not particularly limited, and means any EGFR-TKI that is effective in treating lung cancer having a mutated and overactive EGFR, for example, gefitinib, ertintinib, or Combinations thereof.

As used herein, the term "PTEN deterioration" refers to a case in which the expression of PTEN is reduced or the dephosphorylation function of PTEN is inhibited by obtaining a mutation that induces impairment.

As used herein, an "mTOR inhibitor" refers to a drug that stops the cell cycle while inhibiting mTOR, which is a lower target of the PI3K / Akt signaling system that mediates drug resistance and migration, including rapamuycin, everolimus, Or combinations thereof.

As used herein, “pharmaceutically acceptable salts” may be acid addition salts formed by free acid. The acid addition salts can be prepared by conventional methods, for example, by dissolving the compound in an excess aqueous acid solution and precipitating the salt using a water miscible organic solvent such as methanol, ethanol, acetone, or acetonitrile. An equimolar amount of the compound and an acid or alcohol in water such as glycol monomethyl ether can be heated and then the mixture is evaporated to dryness or the precipitated salts can be suction filtered.

The free acid may be an organic acid or an inorganic acid. For the inorganic acid, for example, hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, and the like may be used. For the organic acid, for example, methanesulfonic acid, p-toluenesulfonic acid, acetic acid, Trifluoroacetic acid, citric acid, meleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, manderic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, Aspartic acid, ascorbic acid, carbonic acid, vaninyl acid, hydroiodic acid and the like can be used.

Bases may also be used to make pharmaceutically acceptable metal salts. An alkali metal or alkaline earth metal salt can be obtained, for example, by dissolving a compound in an excess of alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and then evaporating and drying the filtrate. Examples of the metal salts include sodium, potassium, or calcium salts. The preparation of such salts may be appropriately selected by those skilled in the art according to the specific type of mTOR inhibitor.

When the mTOR inhibitor and radiation alone were administered alone for the EGFR-TKI resistant lung cancer model, the apoptotic effect was very weak. However, the combination showed a synergistic killer effect on the EGFR-TKI resistant lung cancer model. Excellent radiation treatment effect was shown. Therefore, the mTOR inhibitor acts as a radiosensitizer for EGFR-TKI resistant lung cancer and can be used as a composition for radiotherapy of EGFR-TKI resistant lung cancer.

According to one embodiment, the EGFR-TKI resistant lung cancer is lung cancer obtained resistance by PTEN loss, in particular small cell lung cancer.

The pharmaceutical composition for radiation therapy according to the present invention may be administered at the same time as the radiation treatment, and as long as the effects of the radiation therapy and the radiation therapy composition according to the present invention are within a range having interaction with each other, administration of the radiation therapy composition And time gaps in radiation therapy.

The pharmaceutical composition for radiotherapy according to the present invention may further be formulated in a conventional pharmaceutical formulation known in the art, further comprising a pharmaceutically acceptable carrier. The pharmaceutical formulation may be formulated and administered in any formulation including, but not limited to, oral administration, injection, suppository, transdermal administration, and non-administration, but preferably liquid, suspension, powder Oral dosage forms such as granules, tablets, capsules, pills, or extracts.

The term "pharmaceutically acceptable carrier" is used herein to refer to any component except for an mTOR inhibitor which is included as the radiation sensitive agent. "Pharmaceutically acceptable" means a property that does not cause pharmaceutically undesirable changes by interacting with other constituents present in the composition (e.g., interaction between carriers or radiation-sensitive agent and carrier). Means. The choice of such pharmaceutically acceptable carrier will depend on such factors as the nature of the particular dosage formulation, the manner of administration, the solubility and the effect of the carrier on stability.

In one embodiment, when the composition is formulated into a tablet, the pharmaceutically acceptable carrier is 1 selected from diluents, binders, lubricants (or lubricants), disintegrants, stabilizers, solubilizers, sweeteners, colorants, flavors. It may be more than one species, but is not limited thereto.

Diluent refers to any excipient added to increase the volume of the composition to make it the appropriate size according to the formulation. The diluent may be a starch (eg potato starch, corn starch, wheat starch, pregelatinized starch), microcrystalline cellulose (eg low water microcrystalline cellulose), lactose (eg lactose monohydrate, Anhydrous lactose, spray lactose), glucose, sorbitol, mannitol, sucrose, alginate, alkaline earth metal salts, clay, polyethylene glycol and dicalcium phosphate, anhydrous calcium hydrogen phosphate, silicon dioxide and the like may be used alone or as a mixture thereof. It is not limited.

Binder refers to a material used to impart adhesion to powdery materials to facilitate compaction and improve flowability. The binder is starch, microcrystalline cellulose, highly dispersible silica, mannitol, lactose, polyethylene glycol, polyvinylpyrrolidone, cellulose derivatives (e.g., hydroxypropylmethylcellulose, hydroxypropylcellulose, low substituted hydroxy) Propylcellulose), natural gums, synthetic gums, povidone, copovidone and gelatin may be one or more selected from, but is not limited thereto.

Disintegrant refers to a substance that is added to facilitate the collapse or disintegration of a solid formulation after in vivo administration. The disintegrant is starch or modified starch such as sodium starch glycolate, corn starch, potato starch or starch gelatinized starch, clay such as bentonite, montmorillonite, veegum, microcrystalline cellulose, hydroxypropyl cellulose or Celluloses such as carboxymethylcellulose, alginates such as sodium alginate or alginic acid, crosslinked celluloses such as croscarmellose sodium, gums such as guar gum, xanthan gum, crosslinked polymers such as crosslinked polyvinylpyrrolidone Effervescent agents such as sodium bicarbonate and citric acid may be used alone or in combination, but is not limited thereto.

Glidant or lubricant refers to a material that performs the function of preventing the adhesion of powder to the compaction equipment and improving the flow of granules. The lubricant is hard silicic anhydride, talc, stearic acid, metal salt of stearic acid (such as magnesium salt or calcium salt), sodium lauryl sulfate, hydrogenated vegetable oil, sodium benzoate, sodium stearyl fumarate, glyceryl bihenate, glyceryl mono Stearate or polyethylene glycol may be used alone or in combination, but is not limited thereto.

Adsorbents can be used alone or in combination with hydrous silicon dioxide, light silicic anhydride, colloidal silicon dioxide (trade name: Aerosil, degussa), magnesium metasilicate aluminate, microcrystalline cellulose, lactose or crosslinked polyvinylpyrrolidone. It is not limited to this.

Stabilizers include antioxidants such as butylhydroxyanisole, butylhydroxytoluene, carotene, retinol, ascorbic acid, tocopherol, tocopherol polyethylene glycol succinic acid or propylgallate, cyclodextrin, carboxyethyl cyclodextrin, hydroxy Cyclic compounds of saccharides such as propyl cyclodextrin, sulfobutyl ether or cyclodextrin, organic acids such as phosphoric acid, lactic acid, acetic acid, citric acid, tartaric acid, succinic acid, maleic acid, fumaric acid, glycolic acid, propionic acid, gluconic acid or glucuronic acid It may be more than one species, but is not limited thereto.

Optionally, known additives may be included to enhance the palate to enhance palatability of the drug. For example, sweeteners such as sucralose, sucrose, fructose, erythritol, acesulfame potassium, sugar alcohols, honey, sorbitol or aspartame can be added. In addition, acidulant such as citric acid, sodium citrate, natural flavors such as plum flavor, lemon flavor, pineapple flavor, herbal flavor, natural pigments such as natural fruit juice, chlorophyllin, flavonoids can be used.

According to one embodiment, the tablet may be compressed tablets, multi-compressed tablets, internal tablets, dragees, chewed tablets, troche tablets, sublingual tablets, thin knife tablets, effervescent tablets, fast tablets, diffusion tablets or orally disintegrating tablets.

Techniques required for such formulations and pharmaceutically suitable carriers, additives and the like are well known to those of ordinary skill in the art of pharmaceutical formulations and reference may be made to Remington's Pharmaceutical Sciences (19th ed., 1995). .

The pharmaceutical composition for radiation therapy may be administered in several divided doses so that the total daily dose is 1-1000 mg / kg as a compound as an active ingredient in order to obtain a radiotherapy-promoting effect on EGFR-TKI resistant lung cancer. have. The dosage may be appropriately increased or decreased depending on the specific type of mTOR inhibitor, the progression of lung cancer, the route of administration, sex, age, weight, and the like.

The pharmaceutical composition according to the present invention includes a mammalian target of rapamycin (mTOR) inhibitor as a radiation sensitizer, thereby preventing susceptibility to radiotherapy of epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistant lung cancer including zephytinib. It is expected that it can be effectively used in the treatment of EGFR-TKI resistant lung cancer.

FIG. 1A shows Western blotting results for EGFR, PTEN, AKt, mTOR, and S6 of HCC827 cells infected with HCC827 cells with a lentivirus inserted with shPTEN (PTEN shRNA), thereby reducing PTEN expression. The photograph was taken in comparison with the result of the inserted cells.
FIG. 1B shows that HCC827 cells were infected with a lentivirus inserted with shPTEN (PTEN shRNA) and treated with various concentrations of zephytinib for HCC827 cells that decreased PTEN expression. Is a graph shown in comparison with the results of cells inserted with shCON (control shRNA).
Figure 1c shows that HCC827 cells were infected with a lentivirus inserted with shPTEN (PTEN shRNA) and exposed to radiation of 1, 2. and 3 Gy to HCC827 cells that reduced PTEN expression. The result of the measurement is a graph which is compared with the result of the cell which inserted shCON (control shRNA).
FIG. 2A shows HCC827 cells infected with a lentivirus inserted with shPTEN (PTEN shRNA) and treated with various concentrations of rapamycin for HCC827 cells having reduced PTEN expression. It is a graph compared with the result of the cell which inserted shCON (control shRNA).
FIG. 2B shows that HCC827 cells were infected with a lentivirus inserted with shPTEN (PTEN shRNA) and treated with 100 nM of rapamycin for HCC827 cells that degraded PTEN expression and then exposed to radiation of 1, 2., and 3 Gy. It is a graph showing the results of the radiotherapy efficiency measured by the clonal survival analysis, compared with the results of the cells inserted shCON (control shRNA).
FIG. 3A shows HCC827 cells infected with a lentivirus inserted with shPTEN (PTEN shRNA) and treated with various concentrations of RAD001 for HCC827 cells having reduced PTEN expression. It is a graph compared with the result of the cell which inserted (control shRNA).
3B shows HCC827 cells infected with a lentivirus inserted with shPTEN (PTEN shRNA) and treated with RAD001 100 nM for HCC827 cells that degraded PTEN expression, followed by radiation of 1, 2. and 3 Gy for HCC827 cells. After exposure to the radiation treatment efficiency was measured by the clonal survival analysis, a graph showing the results compared with the results of the cells inserted shCON (control shRNA).

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

<Experimental Method>

1. Cell culture

HCC827 cells, which are non-small cell lung cancer cell lines, were purchased from the US Center for Microbial Conservation (ATCC) and cultured in an incubator at 37 ° C. and 5% CO ₂ in RPMI-1640 medium.

2. Reagents and Irradiation Conditions

Gefitinib (Cat # G-4408), RAD001 (Cat # E-4040), and rapamycin (Cat # R-5000) were all purchased from LC Laboratories (Woburn, Mass., USA).

Irradiation was performed at 3.51 Gy / min using a 137Cs γ-ray source (Atomic Energy of Canada). A total of 1, 2, or 3 Gy was investigated.

3. Western Blotting

Where to purchase the primary antibody used for Western blotting is as follows.

anti-phospho EGFR (Cat # 2236), anti-PTEN (Cat # 9552), anti-phospho AKT (Cat # 4060), anti-phospho mTOR (Cat # 2971), anti-phospho S6 (Cat # 4856), anti -S6 (Cat # 2317), and anti-Beclin1 (Cat # 3738) were purchased from Cell Signaling (Danvers, MA, USA), anti-AKT (Cat # 5298) and anti-EGFR (Cat # 03) were Santa It was purchased from Cruz Biotechnology (Santa Cruz, CA, USA), and anti-LC3 (Cat # NB100-2220) was purchased from Novus Biologicals (Littleton, CO, USA).

Western blotting method was as follows.

To prepare total cell lysate, the cells were washed in cold phosphate buffered saline and the cells were lysed [50 mM Tris, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1 mM phenylmethylsulfonyl fluorine]. Ride, 1 μg / ml aprotinin, 1 μg / ml leupetin, 1 μg / ml pepstatin, 1 mM NaF, 1 mM sodium orthovanadate, 0.25% sodium deoxycholate, and 1% Nonidet P-40 Dissolved. After centrifugation at 10,000 × g, supernatants were collected. A 50 ug aliquot of protein sample was electrophoresed and then transferred to nitrocellulose membrane. The nitrocellulose membrane was immunoblotted with primary antibody and horseradish peroxidase-conjugated anti-mouse IgG. Immunoblotting proteins were visualized with a chemiluminescent ECL system (Amersham Biosciences).

4. Lentivirus shRNA  Production and in vitro Transduction

PLKO.1 lentiviral vectors expressing shRNA (Cat # 25638) or sc shRNA (scrambled shRNA) (Cat # 1864) for PTEN were purchased from Addgene (USA). Lentiviral vector stocks were produced in 293T cells by lipofectamine with metastatic transfection of shRNA constructs with pHR8.2R and pCMV-VSV-G helper constructs. The supernatant was collected after 30 hours and filtered through a 0.22 micron filter (Millipore) to remove any unattached 293T cells. HCC827 cells (5 × 10 ⁴ in a 35 mm diameter dish) were then transduced with 1 ml of virus containing supernatant supplemented with 8 μg polybrene. Medium was changed 1 day after infection and fresh medium was fed for 2 days.

5. MTT Assay

Cells in the log growth phase were harvested and plated in 96-well plates. After incubation for 24 hours, gefitinib was added. After 3 days of incubation, MTT assays were performed to determine cell viability: 4 hours after the end of each experiment, 50 μl of PBS containing 5 mg / ml MTT was added to the culture medium. The medium was then gently removed and 50 μl of DMSO was added to each well to dissolve the formazan precipitate. The absorbance of each well was measured at 560 nm.

6. Clonogenic  Survival analysis Clonogenic survival assay )

The cells were trypsinized and then plated into three 60-mm dishes in triplicate 24 hours prior to irradiation. Cells were treated with the indicated concentrations of mTOR inhibitor or vehicle control 2 hours prior to irradiation of the indicated dose. Six hours after irradiation, the medium containing the mTOR inhibitor was removed and the cells were kept in normal culture medium. Two weeks later, the cells were stained with crystal violet to determine the number of colonies consisting of 50 or more cells. Plating efficiency was calculated by dividing the average number of cell colonies per well by the amount of plated cells. Survival rates were calculated as normalization to the smear efficiency of the appropriate control.

<Experimental Results>

1. Susceptibility to Radiation and Zephytinib in PTEN Degraded HCC827 Cells

HCC827 cells have a normal PTEN gene. Lentiviruses inserted with shRNA (short hairpin RNA) targeting PTEN mRNA were infected with HCC827 to reduce PTEN expression of HCC827 cells.

As shown in FIG. 1A, shCON (control shRNA) and shPTEN (PTEN shRNA) were inserted into HCC827 cells using a virus and cells (-) not exposed to the virus were used as negative controls. Decreased PTEN expression by shPTEN was confirmed by Western blotting (FIG. 1A).

Phosphorylation of Akt, mTOR, and S6 was confirmed by Western blotting to determine the effect of lower PTEN expression on downstream signaling. Decreased PTEN expression increased phosphorylation of Akt, mTOR, and S6.

As in FIG. 1B, HCC827 cells expressing shCON or shPTEN were treated at various concentrations indicated in the figure. After 72 hours, the effect of iresa on cell survival was investigated by MTT method, and it was confirmed that the acquisition of iresa resistance by PTEN expression reduction was performed (FIG. 1B).

Through the above experiments, a cell-level model was established for patients with IRESA resistance-induced lung cancer due to PTEN expression.

As shown in Fig. 1c, HCC827 cells expressing shCON or shPTEN were exposed to radiation of various intensities indicated in the figure, and the radiation treatment efficiency was measured by clonality survival analysis (Fig. 1c). Surprisingly, PTEN expression was shown to confer not only resistance to iresa (FIG. 1B) but also resistance to radiation therapy (FIG. 1C).

2. PTEN  Lowering HCC827  Radiation of cells and Zephytinib  Sensitivity to

As shown in FIG. 2A, HCC827 cells expressing shCON or shPTEN were treated with rapamycin at various concentrations as shown in the figure, and the effects of rapamycin on HCC827 survival were examined by MTT assay. The results are shown in A of FIG.

As known, rapamycin did not affect the survival of HCC827 cells and showed resistance to rapamycin regardless of the difference in PTEN expression (FIG. 2A).

Rapamycin alone treatment did not show anticancer effects in HCC827 cells (FIG. 2A). However, after treatment with rapamycin to HCC 827 cells, irradiation showed a significant increase in radiotherapy efficiency (FIG. 2B).

Another mTOR inhibitor, RAD001, also showed an increase in radiotherapy efficiency, similar to rapamycin. After HCON827 cells expressing shCON or shPTEN as shown in FIG. 3A were treated with various concentrations of RAD001, the effect of RAD001 on the survival of HCC827 was examined by MTT assay. As a result, similar to rapamycin, RAD001 did not affect the survival of HCC827 cells and showed resistance to RAD001 regardless of the difference in PTEN expression (FIG. 3A).

RAD001 alone treatment did not show anticancer effect in HCC827 cells (FIG. 3A). However, when RAD001 was treated with HCC827 cells and then irradiated with radiation, the radiation treatment efficiency was significantly increased (FIG. 3B).

According to the experimental results, for epithelial growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistant lung cancer, such as zephytinib, radiation alone or mTOR inhibitor alone has little anticancer effect. In addition, it can be seen that the anticancer effect is significantly increased when the mTOR inhibitor is used in combination with irradiation. Thus, it can be seen that the mTOR inhibitor can be used as a radiosensitizer in radiotherapy of EGFR-TKI resistant lung cancer.

Claims (8)

Inhibitor of epidermal growth factor receptor tyrosine kinase due to mTOR (mammalian target of rapamycin) inhibitor or phosphatase and tensin homolog (PTEN) function including pharmaceutically acceptable salts, solvates, or hydrates thereof Epithelial Growth Factor Receptor-Tyrosine Kinase Inhibitor (EGFR-TKI) -resistant pharmaceutical composition for radiotherapy of lung cancer. The pharmaceutical composition of claim 1, further comprising a pharmaceutically acceptable carrier. The pharmaceutical composition of claim 1, wherein the mTOR inhibitor is selected from the group consisting of rapamuycin, everolimus, and combinations thereof. The pharmaceutical composition of claim 1, wherein the epidermal growth factor receptor tyrosine kinase inhibitor is selected from the group consisting of gefitinib, erlotinib, and a combination thereof. The pharmaceutical composition of claim 1, wherein the lung cancer is small cell lung cancer. The pharmaceutical composition according to any one of claims 1 to 5, which is for oral administration. The pharmaceutical composition of claim 6 which is a tablet. The pharmaceutical composition of claim 7, wherein the tablet is a compressed tablet, a multi-compressed tablet, an internal tablet, a dragee tablet, a chewed tablet, a troche tablet, a sublingual tablet, a thin tablet, an effervescent tablet, a fast tablet, a diffusion tablet, or an orally disintegrated tablet.

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