TW201400810A - Chemotherapy selection method for stomach cancer patients - Google Patents

Abstract

The purpose of the invention is to provide chemotherapy having a strong life-prolonging effect and few adverse effects for stomach cancer patients. Provided to achieve this purpose is a method for predicting the therapeutic effect of chemotherapy using tegafur-gimeracil-oteracil potassium compounding agent in stomach cancer patients, including the following steps (1)-(3): (1) a step for measuring the HER2 mRNA expression level contained in a biological sample collected from the patient; (2) a step for comparing the HER2 mRNA expression level obtained in step (1) with a cut-off point that is the pre-set median value of the HER2 mRNA expression level contained in biological samples from stomach cancer patients; and (3) a step for predicting that there is a high likelihood that selection of chemotherapy combining tegafur-gimeracil-oteracil potassium compounding agent and an HER2 inhibitor will have an adequate therapeutic effect in the patient when the expression level of HER2 mRNA is at or above the cut-off point in the result of the comparison in step (2), and to predict that there is a high likelihood that selection of chemotherapy using tegafur-gimeracil-oteracil potassium compounding agent alone will have an adequate therapeutic effect when the HER2 mRNA expression level is below the cut-off point.

Description

Method of selecting chemotherapy for patients with gastric cancer Field of invention Cross-references to related applications

The present application claims priority to the specification of Japanese Patent Application No. 2012-125554, the entire disclosure of which is hereby incorporated by reference.

The present invention relates to a method for predicting the therapeutic effect of chemotherapy using at least a tegafur-gimerazine-oteracil potassium complex, and an antitumor agent for predicting the use of the complexing agent. Chemotherapy will show that patients with high likelihood of adequate therapeutic effects are administered.

Background of the invention

For the chemotherapy of progressive gastric cancer, 5-Flurouracil, Cisplatin, Irinotecan, Docetaxel, Tegafur-uracil complex are clinically applied. (trade name: UFT (registered trademark)), tegafur-giramin-otarazide potassium complex (trade name: TS-1 (registered trademark); below, will also be tegafur, gemcitabine, oloti A preparation in which Laxi potassium is compounded with a molar ratio of 1:0.4:1 is called an antitumor agent such as TS-1).

On the other hand, in the post-operative adjuvant chemotherapy for gastric cancer for the purpose of preventing recurrence/metastasis after resecting gastric cancer tumor tissue, the third phase clinical trial ACTS-GC was accumulated by accumulating cases with more than 1000 cases. In the study, significant survival prolongation was achieved in either the overall survival time and the recurrence-free survival time, as compared with the single surgery group. There is no problem with its safety, and it is currently recognized as a standard treatment in Japan (Non-Patent Document 1).

However, there is a problem in that since the adjuvant chemotherapy after surgery will work largely depending on the genetic factors of the patient, it is not known if the antitumor agent is not actually administered. Thus, biomarkers for predicting whether the antitumor agent will work before administration of the antitumor agent are being actively pursued. In the case of post-operative adjuvant chemotherapy using TS-1 alone in gastric cancer patients, it is suggested that TS, DPD (Non-Patent Document 2), ERCC1 (Non-Patent Document 3), and EGFR (Non-Patent Document 4) are biomarkers. possibility. On the other hand, HER2 was reported, and the amount of expression of the protein was not observed to be related to the therapeutic effect (Non-Patent Documents 4 and 5).

As described above, although post-operative adjuvant chemotherapy for gastric cancer is being actively developed, the therapeutic effect is not satisfactory at present.

Prior technical literature

Non-patent literature

Non-Patent Document 1 N Engl J Med. 2007;357(18):1810-20.

Non-Patent Document 2 J Clin Oncol. 2012;30 (Suppl. 4; abstr 52)

Non-Patent Document 3 J Clin Oncol. 2012;30 (Suppl. 4; abstr 53),

Non-Patent Document 4 J Clin Oncol. 2011;29(Suppl;abstr 4013)

Non-Patent Document 5 J Clin Oncol. 2011;29(Suppl;abstr e14501)

Summary of invention

The object of the present invention is to provide a chemotherapy for a gastric cancer patient that achieves a powerful prolongation effect with few side effects.

The inventors of the present invention have found that chemotherapeutic treatment of tegafur-giramin-otimrazide potassium alone is used when the expression of HER2 mRNA is lower than the median for the repeated study of chemotherapy for patients with gastric cancer. It is easy to work; and when the amount of expression of HER2 mRNA is higher than the median, the chemotherapy with tegafur-gibimerine-oteracil potassium complex and HER2 inhibitor is easy to work, and the present invention is completed.

Furthermore, in the ToGA test for patients with progressive/recurrent gastric cancer who cannot be surgically removed, patients with high HER2 expression (3+ of the HER2 protein by the IHC method) The patient, or the patient who scored the HER2 protein by the IHC method, and the patient with the HER2 gene by the FISH method, was positive, suggesting trastuzumab (trastuzumab) An additional effect on the combination of 5-FU or Capecitabine with cisplatin (Lancet. 2010; 376 (9742): 687-97.). However, as described in Non-Patent Documents 4 and 5, In the postoperative adjuvant chemotherapy for patients with gastric cancer using TS-1, there was no significant difference in therapeutic effect between the group with high performance of HER2 protein and the group with low performance, suggesting trastuzumab (trastuzumab) The extra effect is low.

In addition, in the case of postoperative adjuvant chemotherapy for patients with gastric cancer using TS-1, it is not known that patients with a higher than HER2 mRNA expression have trastuzumab (trastuzumab). Additional effects, especially in patients with low performance of HER2 in the determination of the ToGA test for trastuzumab, which cannot be expected to have additional effects (Identification score for HER2 protein by IHC method) 0 or 1+ patients, or the HER2 protein judged by the IHC method is classified as 2+ and the HER2 gene is judged by the ISH method to be negative, and the expression of HER2 mRNA is For patients above the median, there is an additional effect of trastuzumab.

That is, the present invention encompasses the following aspects.

Item 1, a method for predicting the therapeutic effect of chemotherapy using tegafur-giramin-oteracil potassium in a gastric cancer patient, comprising the following steps (1) to (3): (1) Taking the step of expressing the amount of HER2 mRNA contained in the living sample of the patient; (2) comparing the expression amount of the HER2 mRNA obtained in the above step (1) with a preset cut-off point In the comparison step, the corresponding cut-off point is the median amount of HER2 mRNA expression contained in the living sample of the gastric cancer patient; (3) a prediction step, which is based on the comparison result of the above step (2), when the expression amount of the HER2 mRNA is higher than or equal to the corresponding cut-off point, it is predicted that the patient is selected to use tegafur-giramin- Chemotherapy with oltipraz potassium complex and HER2 inhibitors may show a high degree of therapeutic benefit; and when the HER2 mRNA performance is below the corresponding cut-off point, it is predicted that the patient should be used alone. Chemotherapy of the tegafur-giramin-oteracil potassium complex may show a high degree of therapeutic benefit.

Item 2. The method according to Item 1, wherein the molar ratio of each active ingredient of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4 :1.

Item 3. The method of Item 1 or 2, wherein the HER2 inhibitor is trastuzumab.

The method of any one of items 1 to 3, wherein the chemotherapy is postoperative adjuvant chemotherapy.

Item 5. The method according to any one of items 1 to 4, wherein the gastric cancer patient is a patient whose score is 0 or 1+ by using the IHC method, or is the HER2 using the IHC method. The protein was scored as 2+ and the patients who were negative by the interpretation of the HER2 gene by the ISH method were classified.

Item 6, an anti-tumor agent consisting of tegafur-giramin-oteracil potassium complex and HER2 inhibitor, which is used for treating gastric cancer patients, and the gastric cancer patient has been as in the first It is predicted in any of the five methods that chemotherapy with a tegafur-giramin-oteracil potassium complex and a HER2 inhibitor may show a high therapeutic effect.

Item 7. The antitumor agent according to Item 6, wherein the molar ratio of each active ingredient of the tegafur-gimerazine-otarazide potassium complex is tegafur: gemcitabine: oltipraz potassium = 1 :0.4:1.

Item 8. The antitumor agent according to Item 6 or 7, wherein the HER2 inhibitor is trastuzumab.

Item 9, an antitumor agent consisting of a tegafur-giramin-oteracil potassium complex for treating a gastric cancer patient, and the gastric cancer patient has been in any of items 1 to 5 In one method, it was predicted that chemotherapy with tegafur-giramin-oteracil potassium complex alone would show a high probability of adequate therapeutic effects.

Item 10. The antitumor agent according to Item 9, wherein the molar ratio of each active ingredient of the tegafur-gimerazine-otarazide potassium complex is tegafur: gemcitabine: oltipraz potassium = 1 :0.4:1.

Item 11. The antitumor agent according to any one of items 6 to 10, which is for postoperative adjuvant chemotherapy.

Item 12, a method for treating gastric cancer, comprising the following steps (1) to (3): (1) a step of measuring the amount of expression of HER2 mRNA contained in a living sample of a gastric cancer patient; (2) The step of comparing the expression amount of the HER2 mRNA obtained in the step (1) with a preset cut-off point, which is the median of the expression amount of the HER2 mRNA contained in the living body sample of the gastric cancer patient. And (3) the step of performing chemotherapy, which is a comparison of the above steps (2) If the expression level of HER2 mRNA is higher than or equal to the corresponding cut-off point, then chemotherapy with tegafur-gimex-otaprazole potassium complex and HER2 inhibitor is carried out.

Item 13. The method according to Item 12, wherein the molar ratio of each active ingredient of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4 :1.

Item 14, a method for treating gastric cancer, comprising the following steps (1) to (3): (1) a step of measuring the amount of expression of HER2 mRNA contained in a living body sample of a gastric cancer patient; (2) The step of comparing the expression amount of the HER2 mRNA obtained in the step (1) with a preset cut-off point, which is the median of the expression amount of the HER2 mRNA contained in the living body sample of the gastric cancer patient. And (3) a step of performing chemotherapy according to the comparison result of the above step (2), when the expression amount of the HER2 mRNA is lower than the corresponding cut-off point, the patient is separately administered tegafur-jimi Chemotherapy of pyrimidine-oteracil potassium complex.

Item 15. The method according to Item 14, wherein the molar ratio of each active ingredient of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4 :1.

Item 16. The method of Item 14 or 15, wherein the HER2 inhibitor is trastuzumab.

Item 17, the method of any one of the items 12 to 16, wherein Learning therapy is postoperative adjuvant chemotherapy.

The method of any one of the items 12 to 17, wherein the gastric cancer patient is a patient whose score is 0 or 1+ by using the IHC method, or is the HER2 using the IHC method. The protein was scored as 2+ and the patients who were negative by the interpretation of the HER2 gene by the ISH method were classified.

Item 19, a use of a tegafur-jimeicilin-oteracil potassium complex and a HER2 inhibitor for the manufacture of an antitumor agent for treating a gastric cancer patient, and the gastric cancer patient has been as in the first It is predicted in any of the five methods that chemotherapy with tegafur-giramin-oteracil potassium complex and HER2 inhibitor may show a high therapeutic effect.

Item 20. The use of the item 19, wherein the molar ratio of the active ingredients of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4 :1.

Item 21. The use of item 19 or 20, wherein the HER2 inhibitor is trastuzumab.

Item 22, a use of a tegafur-giramin-oteracil potassium complex for the manufacture of an antitumor agent for treating a gastric cancer patient, and the gastric cancer patient has been in any of items 1 to 5 In one method, it was predicted that chemotherapy with tegafur-giramin-oteracil potassium complex alone would show a high probability of adequate therapeutic effects.

Item 23. The use of the item 22, wherein the molar ratio of each active ingredient of the tegafur-giramin-oteracil potassium complex is tegafur: Meipyrimidine: oltipraz potassium = 1:0.4:1.

Item 24. The use of any one of items 19 to 23 for postoperative adjuvant chemotherapy.

Item 25, an agent for predicting a therapeutic effect on chemotherapy for a gastric cancer patient, wherein the chemotherapy uses a tegafur-giramin-oteracil potassium complex, the reagent comprising: the sequence number 3 A primer consisting of a base sequence, a primer consisting of the nucleotide sequence shown by SEQ ID NO: 4, and a probe consisting of the nucleotide sequence shown by SEQ ID NO: 5.

The predictive method of the present invention makes it possible for gastric cancer patients to select chemotherapy with a higher prolongation effect. That is to say, in the case of gastric cancer, a patient with a relatively low fatal effect on chemotherapy using a single dose of tegafur-gimerazine-oteracil potassium compound may provide a strong prolongation effect. Combination chemotherapy with tegafur-giramin-oteracil potassium complex and HER2 inhibitor.

In addition, chemotherapeutic treatment with a single dose of tegafur-gimidin-oteracil potassium combination showed higher chemotherapeutic than the combination of tegafur-jimeicil-otarazide potassium complex with HER2 inhibitor. In the case of gastric cancer patients with therapeutic effects, since unnecessary HER2 inhibitors can be omitted, the side effects caused by the HER2 inhibitor can be reduced, and the medical economy is also ideal.

Figure 1 shows the IHC score of HER2 and the amount of HER2 mRNA expression (HER2/ACTB, log2 conversion value (ratio)). For each group, the median (median, HER2/ACTB) of the number of cases and the amount of HER2 mRNA expression was combined.

Fig. 2 shows the results of the HER2 protein expression amount of human gastric cancer-derived cell line GCIY measured by immunohistochemical staining (IHC method).

Figure 3 shows TS-1 and Hepatoma by the mean (mean) of the tumor volume of the administration group relative to the tumor volume of the non-administered control group (Relative Tumor Volume (RTV)). (Herceptin) combined effect. In the figure, the ** line indicates a significant difference with respect to the non-administration control group (Control) (P < 0.01, Aspin-Welch's t test). The ## line indicates a significant difference with respect to all comparisons (the maximum value of the significant level obtained at all comparisons is P<0.01 (Overall maximal P<0.01), intersection-joint assay ( Intersection Union test)).

Form for implementing the invention

(i) Prediction method of the present invention

The prediction method of the present invention is based on the amount of HER2 expression in a gastric cancer patient to predict whether the chemotherapy for the patient using the tegafur-gimerazine-oteracil potassium complex will show sufficient therapeutic effect.

HER2, which is an indicator in the present invention, is also referred to as the ERBB2, and is a receptor protein of the tyrosine kinase type belonging to the epidermal growth factor receptor family, which exists through the cell membrane. It transmits the signal of epidermal growth factor and controls the proliferation and growth of cells. It is known that many cancers such as breast cancer and gastric cancer will over-examine. Now. The base sequence and amino acid sequence of human HER2 are registered in GenBank under accession numbers NM004448.2 and NP004439.2, respectively, and such sequence information can be utilized in the present invention. Preferably, in the present invention, the base sequence of human HER2 and the amino acid sequence are shown in SEQ ID NO: 1 and SEQ ID NO: 2, respectively.

The patient who is the subject of the present invention is a gastric cancer patient. In the present invention, "stomach cancer" includes gastric cancer with local recurrence or metastatic gastric cancer that has metastasized to other tissues (for example, lymph nodes) in addition to primary gastric cancer, and is preferably primary gastric cancer. In addition, the patients with gastric cancer here are not only patients with gastric cancer tissue, but also patients who have undergone resection of gastric cancer tissue.

Further, the patient who is the subject of the present invention is preferably a gastric cancer patient whose score is 0 or 1+ by the IHC method, or a judgment score of 2 for the HER2 protein by the IHC method. + Patients with gastric cancer who have a negative score in the HER2 gene by the ISH method, and more preferably use the IHC method to calculate the HER2 protein in 0 or 1+ patients, or use the IHC method. The HER2 protein was counted as 2+ and the HER2 gene was negatively diagnosed by the FISH method. The patient was diagnosed with the HER2 protein by the IHC method. patient.

The interpretation score of the HER2 protein by the IHC method can be determined by the HER2 test by the IHC method and by evaluating the intensity of expression of the HER2 protein. Here, the IHC method (Immunohistochemistry) is an immunohistochemical staining method in which a protein of interest is detected by an antigen-antibody reaction, and the intensity of protein expression is judged. The interpretation score of the IHC method is expressed in four stages of 0, 1+, 2+, and 3+ in response to the HER2 protein expression intensity (Histopathology 2008; 52: 797-805.). The interpretation score of the HER2 protein by the IHC method can be determined by a commonly known method using an examination using a general anti-HER2 antibody. This inspection check may also be used a commercially available kit (e.g., HercepTest ^TM kit (manufactured by DACO)) is performed.

Furthermore, the HER2 gene interpretation score by the ISH method can be performed by the HER2 test by the ISH method and evaluated by evaluating the copy number of the HER2 gene. The ISH method (in situ hybridization) is a method in which a probe is used to hybridize with a gene of interest to detect amplification of a gene, and a FISH method using fluorescent probe (fluorescence in situ hybridization) can be used. Fluorescence in situ hybridization)), DISH method using two types of labeled probes (Dual color in situ hybridization), CISH method using a probe labeled with a dye such as DAB Chromogen in situ hybridization or the like is preferably a FISH method and a DISH method. The interpretation score of the ISH method is expressed in two stages of positive and negative in response to the copy number of the HER2 gene (Histopathology 2008; 52: 797-805.).

The interpretation score of the HER2 gene by the FISH method can be determined by a commonly known method using a fluorescent probe having a hybridization with the versatile HER2 gene. This inspection check may also be used a commercially available kit (e.g., pharmDx ^TM kit (manufactured by DACO)) is performed.

The interpretation score of the HER2 gene by the DISH method can be hybridized with the universal HER2 gene by using a commonly known method. The examination of the two types of probes was obtained. This inspection can also be carried out using a commercially available inspection kit (for example, a Ventana INFORM DUAL ISH HER2 kit (manufactured by Roche Diagnostics Co., Ltd.)).

The "tegafloxacin-gimidazole-oteracil potassium complex" in the present invention means a complex of three doses of tegafur, gemcitabine and oltipraz. "Tegafur" (general name; chemical name: 5-fluoro-1-(2-tetrahydrofuranyl)-2,4-(1H,3H)-pyrimidinedione (5-fluoro-1-(2-tetrahydrofuryl) -2,4-(1H,3H)-pyrimidinedione)) is a compound known to be 5-fluorouracil which is activated in vivo and released as an antitumor activity. The Teflon-based system can be produced by a known method, for example, a method described in JP-A-49-10510.

In addition, "Gemexyrimidine" (general name; chemical name: 2,4-dihydroxy-5-chloropyridine) is also a known compound, which itself does not have antitumor activity. However, it inhibits the metabolism of 5-fluorouracil which is metabolized and deactivated in vivo, and enhances the antitumor effect.

Also, "Oturacil potassium" (general name; chemical name: 1,2,3,4-tetrahydro-2,4-dioxo-1,3,5-three Monopotassium 1,2,3,4-tetrahydro-2,4-dioxo-1,3,5-triazine-6-carboxylate) is also a known compound which itself does not have an anti- Tumor activity, but mainly distributed in the digestive tract and inhibits gastrointestinal disorders by inhibiting the activation of 5-fluorouracil at this site.

In the present invention, the ratio of each of the active ingredients of the "Tegafur-gibberel-otarazide potassium complex" is not particularly limited as long as the respective blending purposes can be achieved, for example, as described in Japan. Patent The same range of known compounding agents of the publication No. 2614164 may be the same: relative to tegafur 1 molar, the gemcitabine is about 0.1 to 5 moles, preferably about 0.2 to 1.5 moles, so that Ottilasi Potassium is about 0.1 to 5 moles, preferably about 0.2 to 2 moles. Tejia, tegafur: gemcitabine: oltipraz potassium (Morby) = 1:0.4:1. In addition, tegafur: gemcitabine: oxytetracycline potassium (mole ratio) = 1:0.4:1 tegafur-gimezepine-otarazide potassium complex, can be "TS-1" (commodity Name, Dapeng Pharmaceutical Industry Co., Ltd.).

In the present invention, the "HER2 inhibitor" is not particularly limited as long as it is an agent which hinders the expression or activity of HER2, and may be a low molecular compound such as Lapatinib which is targeted for HER2, trastuzumab (trastuzumab). Any one of anti-HER2 antibodies such as pertuzumab, anti-infective oligonucleotides against HER2, siRNA, shRNA, miRNA, aptamer, etc., among which anti-HER2 antibodies are preferred. , especially good trastuzumab (trastuzumab).

The so-called trastuzumab specifically recognizes HER2 humanized monoclonal antibodies and exerts anti-tumor effects by antibody-dependent cell-mediated Cytotoxicity (ADCC). . It is known that breast cancer and gastric cancer, which are overexpressed by HER2, will work. Further, trastuzumab can be produced by a known method, for example, a method described in Japanese Patent No. 3040121 and Japanese Patent No. 4124480. Commercially available products ("Herceptin" (trade name, Chinese and foreign pharmaceuticals)) can also be used.

Tegafur-Gimperidin-Oturacil Potassium Complex and HER2 The preparation may be prepared by using tegafur, gemcitabine, oltipraz potassium and a HER2 inhibitor as a complexing agent (a preparation containing a plurality of active ingredients) as a single dosage form (1 dosage form), or may be effective as described above. The components are formulated into a single dose and formulated into a plurality of dosage forms (multi-dosage form). Among these, it is preferred to use a formulation of tegafur, gemcitabine and oxylacazine as a complexing agent, and a preparation of a HER2 inhibitor as a single agent, which is used in a multi-dose form.

The dosage form of the antitumor agent is not particularly limited, and may be appropriately selected according to the purpose of treatment, and specifically, an oral agent (a lozenge, a coated lozenge, a powder, a granule, a capsule, a liquid, etc.) may be exemplified. Injections, suppositories, patches, ointments, and the like. Among them, the tegafur-giramin-oteracil potassium complex is preferably in the form of an oral preparation. Each of the antitumor agents can be prepared by a generally known method by using a pharmaceutically acceptable carrier in accordance with the respective administration forms. As such a carrier, various general agents such as excipients, binders, disintegrators, lubricants, diluents, solubilizers, suspending agents, tonicity agents, pH adjusters, buffers can be exemplified. , stabilizers, colorants, flavoring agents, flavoring agents, and the like.

In the present invention, the "chemotherapy using tegafur-giramin-oteracil potassium compound" means at least the chemotherapy of tegafur-giramin-oteracil potassium complex, and not only The chemotherapeutic treatment of tegafur-jimeizide-oteracil potassium complex alone, and the chemotherapy of tegafur-giramin-oteracil potassium complex with other antitumor agents.

Furthermore, the term "the combination of tegafur-gibberel-otarapix potassium complex and HER2 inhibitor" means combination and administration of tegafur-jibiridine-otarazide potassium complexing agent. And the chemistry of HER2 inhibitors Therapy, preferably comprising a combination and administering a molar ratio of tegafur: gemcitabine: oxytetracycline potassium = 1:0.4:1 of tegafur-gimexidine-oteracil potassium complex, and HER2 inhibition Chemotherapy of the agent, the combination of the combination and the administration of the molar ratio to tegafur: gemcitabine: oxytetracycline potassium = 1:0.4:1 of tegafur-gimezepine-otarazide potassium complex, And chemotherapy with trastuzumab. The above agents may be administered simultaneously as long as they have a combined effect, and may be administered at different dates and times.

In the "chemotherapy of tegafur-gimidin-oteracil potassium compound alone" in the present invention, the administration schedule is appropriately selected depending on the age, sex, disease duration, the presence or absence of the transfer, and the medical history of the patient. For example, it can be exemplified by administering a tegafur-giramin-otarazide potassium complex at a dose of 80 mg/m ² (in terms of tegafur, per body surface area) per day (Tegafur: gemcitabine: Ou Tetraxi potassium (Morby) = 1:0.4:1) 4 weeks, after which the drug was discontinued for 2 weeks, so that the total of 6 weeks was a course of treatment, and the course of treatment was repeated once or repeatedly.

In the present invention, the administration schedule in the "combination of tegafur-gibberel-otarazide potassium complex and HER2 inhibitor" may depend on the age, sex, disease duration, metastasis, medical history, etc. of the patient. The conditions are appropriately selected. For example, it can be exemplified by administering Tegafur-Gempyrimidine-Oturacil potassium complex (Tegafur) at 80 mg/m ² (in terms of tegafur, per body surface area)/day. : Gemcitabine: oltipraz potassium (Morby) = 1:0.4:1) 4 weeks, after which the drug was discontinued for 2 weeks, and trastol was administered at 2 mg/kg every 1 week (total 6 times). Monobendum (trastuzumab), for a total of 6 weeks for a course of treatment, the course of treatment is repeated 1 or more times (only, the dose of trastuzumab on the first day of each course is ordered as 4mg/kg).

The chemotherapy of the present invention may be a preoperative adjuvant chemotherapy for tumor removal after the chemotherapy, or a post-operative adjuvant chemotherapy for performing the chemotherapy after the tumor is removed.

In the present invention, the "therapeutic effect" can be evaluated by the tumor shrinking effect and the prolongation effect of survival, and the survival period can be expressed by the overall survival period and the median of progression-free survival. . The so-called "selection and use of tegafur-gimex-otarazide potassium complex and HER2 inhibitor chemotherapy will have a sufficient therapeutic effect" is better than using tegafur-giramin-otelasi alone. The therapeutic effect of the therapeutic effect of the potassium complexing agent on the degree of therapeutic effect, and the so-called "selective use of tegafur-gimidin-oteracil potassium compound chemotherapy alone will have sufficient therapeutic effect", which is better than The therapeutic effect of the degree of therapeutic effect caused by the combined use of tegafur-giramin-oteracil potassium complex and HER2 inhibitor.

The prediction method of the present invention includes the steps (1) to (3) described later.

Step (1) is a step of measuring the amount of expression of HER2 mRNA contained in the living sample of the patient.

The biological sample is a sample of a cancer patient and includes a cancer cell, and is not particularly limited, and examples thereof include a body fluid (blood, urine, etc.), a tissue, a sample thereof, and a culture of the tissue taken out to organize the sample. It is better. Furthermore, the method of taking the living sample can be appropriately selected in accordance with the type of the living sample. From the point of view of ease of use and the point of view of many cancer cells, from cancer Gastric cancer tumor tissue removed by surgery in patients is suitable. For the biological sample, as long as the expression amount of the HER2 mRNA can be measured, a pathological examination specimen subjected to antiseptic treatment such as a formaldehyde-fixed paraffin-embedded pathological specimen may be used.

The measurement method of the present invention is not particularly limited as long as it is a method for measuring the amount of mRNA, and a known measurement method can be used. Examples of such a measurement method include a PCR method, an RT-PCR method, a Northern blot method, a FISH method, a microarray analysis, and a DTP method (Danenberg Tumor Profile method). Among these, the DTP method is preferred from the viewpoints of simplicity and the like. DTP law efficient tumor site embedded pathological specimens from the general pathology specimen formaldehyde fixed, paraffin-extracted mRNA, and by TaqMan ^TM PCR method to determine expression levels of the process (U.S. Pat. No. 6,248,535).

The biological sample is prepared by appropriate treatment in accordance with these measurement methods. Further, as a reagent containing a primer or a probe which can be used for measurement, a reagent related to the present invention described later can be used.

The step (2) is a step of comparing the expression amount of the HER2 obtained in the above step (1) with a preset cut-off point. The corresponding cut-off point is the median of the amount of expression of HER2 mRNA contained in the biological sample of the gastric cancer patient set in advance.

More specifically, it can be exemplified that when the primers using SEQ ID NOs: 3 and 4 and the probe of SEQ ID NO: 5 are determined by the DTP method, it is 0.0503 (the value normalized by the internal standard gene ACTB), and when using Hs00170433_m1 (Applied Biosystems) as a primer / probe and by TaqMan ^TM Low Density Array determined as 0.0437 (by internal standard gene (ACTB, GAPDH, RPLP0) corrects the geometric mean value). Separately, "ACTB" indicates the β-actin gene, "GAPDH" indicates the Glyceraldehyde 3-phosphate dehydrogenase gene, and "RPLP0" indicates the Ribosomal protein LP0 gene.

Step (3) is a prediction step, which is based on the comparison result of the above step (2), when the expression amount of HER2 mRNA is higher than the corresponding cut-off point, it is predicted that the patient is selected and used with tegafur-giramin- Chemotherapy with oltipraz potassium complexes and HER2 inhibitors may show a high likelihood of adequate therapeutic effects; and when HER2 mRNA expression is below the corresponding cut-off point, it is predicted that the patient is selected separately Chemotherapy using a tegafur-jimeizine-oteracil potassium complex will show a high likelihood of adequate therapeutic effects.

(ii) a reagent of the invention

The reagent of the present invention is a reagent for use in the above-described method for predicting the present invention, and includes a primer and/or a probe which specifically hybridizes to HER2 mRNA.

The primer or probe of the present invention specifically hybridizes to a contiguous base sequence of at least 15 bases in the nucleotide sequence shown in SEQ ID NO: 1 and is composed of a base having a base sequence of 15 bases or longer. A probe or primer consisting of nucleotides. Such a primer or a probe has a sequence length of 15 bases or more and is specifically restricted to hybridize with the mRNA of HER2, and is not particularly limited. The sequence length of the primer and the probe should be 15 to 30 bases long, preferably 16 to 25 bases long. Specifically, the base sequence represented by SEQ ID NO: 3 and SEQ ID NO: 4 is preferably the base sequence of the primer, and the base sequence shown by SEQ ID NO: 5 is preferably the base sequence of the probe. The probe should preferably be a fluorescent probe. Among them, a fluorescent compound such as 6-Carboxy fluorescein (FAM) or tetrachlorofluorescein (TET) is bonded to the 5' end of the polynucleotide, respectively, and is 3 'terminus tetramethyl rhodamine (tetramethyl rhodamine) (TAMRA) and the like matting compound, and the probe can be used for the TaqMan ^TM PCR method is particularly preferred.

Here, the so-called specific hybridization means that under stringent hybridization conditions, a specific hybrid is formed and a non-specific hybrid is not formed. The stringent hybridization conditions can be determined based on the melting temperature (Tm) of a nucleic acid which forms a hybrid according to a conventional method. Specific examples of the washing conditions in which the hybridization state can be maintained include the following conditions: "1 x SSC, 0.1% SDS, and 37 ° C", and more strictly speaking, "0.5 x SSC, 0.1% SDS, and 42 ° C". The conditions of "0.1 × SSC, 0.1% SDS, 65 ° C" are more strictly stated.

Further, the polynucleotide preferably has a base sequence complementary to a contiguous base sequence of at least 15 bases in length of the nucleotide sequence shown in SEQ ID NO: 1, as long as the above-described specific hybridization is possible. It does not have to be completely complementary. As such a polynucleotide, a polynucleotide consisting of a nucleotide sequence of at least 15 bases or more consecutive from the nucleotide sequence shown in SEQ ID NO: 1 or a complementary polynucleotide thereof is preferable. In comparison, the nucleotide sequence has 70% or more, preferably 80% or more, preferably 90% or more, more preferably 95% or more, and particularly preferably 98% or more of the same nucleotide. Here, the identity of the base sequences can be searched for similarity, sequence alignment program, BLAST, FASTA, ClustalW Wait to calculate.

Further, these polynucleotides can be produced by a conventional method, for example, by a commercially available nucleotide synthesizer based on the total base length of the nucleotide sequence shown in SEQ ID NO: 1. Further, the total base length of the nucleotide sequence shown in SEQ ID NO: 1 can also be modulated by a PCR method.

[Examples]

Hereinafter, the present invention will be described in more detail based on the examples, but it should be understood that the present invention is not limited to the examples.

[Example 1] HER2 expression amount in gastric cancer patients

In stage II~III (stage II~III) gastric cancer patients (1059 cases), after resected gastric cancer tumor tissue, they were divided into single surgery group and TS-1 treatment group. In TS-1 treatment group, 80~120mg/day (the body surface area is less than 1.25m ² : 80mg/day; 1.25m ² or more and less than 1.5m ² : 100mg/day; 1.5m ² or more; 120mg/day) TS-1 was administered to the patient for 4 weeks, and then the drug was discontinued for 2 weeks. The course was repeated for 6 weeks, and the course was repeated within 1 year after the operation.

In all cases, 809 cases of formaldehyde-fixed paraffin-embedded pathological specimens were obtained from surgically extracted gastric cancer tumor tissues, and the pathological specimens were obtained from paraffin-embedded pathological specimens using the IHC method to determine the amount of HER2 protein expression by using sequence numbers. 3 probes and primers SEQ ID NO 4 and SEQ ID NO 5 to determine the expression levels of HER2 mRNA (DTP method) (Hs00170433_m1, Applied Biosystems Ltd.) the TaqMan ^TM PCR method. Also, the amplification of the HER2 gene was measured by the DISH method. The whole case was grouped based on the IHC score of HER2 and the presence or absence of HER2 gene amplification determined by the DISH method (0, 1+, 2+/DISH(-), 2+/DISH(+), 3 +).

Figure 1 shows the IHC score of HER2 and the amount of HER2 mRNA expression (HER2/ACTB, x1000, log2 conversion value). Furthermore, the median number of HER2 mRNA expression (HER2/ACTB, x1000) was 0.0503 in all cases (the log2 conversion value was -4.31).

[Example 2] Relationship between therapeutic effect of TS-1 (S-1) and expression amount of HER2 mRNA

Next, the TS-1 (S-1) administered group (413 cases), the extraction of RNA from formalin-fixed paraffin-embedded pathological specimens of the cDNA made by reverse transcription, using ^{TaqMan TM Low-density array (Applied} Biosystems Ltd. ) to determine the amount of HER2 mRNA expression. Further, as the primer and the probe, Hs00170433_m1 (manufactured by Applied Biosystems) was used. The median of the amount of HER2 mRNA expression in the TS-1 (S-1) administration group was calculated to be 0.0439 (the value corrected by the geometric mean of ACTB, GAPDH, RPLP0).

Next, let this value be the cut off point, classify the patients whose HER2 mRNA expression is lower than the median into the low performance group, and classify the patients whose HER2 mRNA expression is above the median to the high. The performance group was subjected to survival analysis using the overall survival of each group.

As a result, in the TS-1 (S-1) administration group, the low-performance group (5-year survival rate: 77.7%) and the high-performance group (5-year survival rate: 69.5%) had statistically significant prognosis. Good (Log Rank Test (P=0.047)).

Also, the interpretation of the HER2 protein by the IHC method will be used. Patients who scored 0 or 1+, and those who used the IHC method for the HER2 protein to be classified as 2+ and whose HER2 gene was negatively classified by the DISH method were classified into the low-performance group. The HER2 protein judged by the IHC method is divided into 3+ patients, and the HER2 protein by the IHC method is judged to be 2+ and the HER2 gene using the DISH method is positive for the interpretation score. Patients were assigned to the high performance group and survival analysis was performed using the overall survival in each group. As a result, there was no statistically significant difference in overall survival between the low performance group and the high performance group.

Since the above, irrespective of the performance of HER2 mRNA, TS-1 is effective as a postoperative adjuvant chemotherapy for patients with gastric cancer, but suggests that the TS-1 system is particularly effective in the low-expression group of HER2 mRNA, and In the high-performance group of HER2 mRNA, in addition to TS-1, it is preferable to use an agent which interferes with high-performance HER2.

[Example 3] Verification of the effect of combined use of TS-1 and Herceptin of human gastric cancer cell line expressing HER2

(1) Human gastric cancer-derived cell line GCIY

The amount of expression of the HER2 protein of human gastric cancer-derived cell line GCIY was determined by immunohistochemical staining (IHC method). The assay carried out by the IHC method was carried out in the following manner: a thin section of GCIY embedded in formaldehyde-fixed paraffin was dewaxed; and Dako REAL Peroxidase-Blocking Solution was used to block endogenous peroxidase; Primary antibody reaction (anti-human HER2 rabbit monoclonal antibody (4B5)); secondary antibody reaction using biotin-labeled anti-rabbit IgG goat polyclonal antibody, peroxidase-labeled streptavidin; and chromogenic substrate (3 3 '-Diaminobenzidine tetrahydrochloride +CHROMOGEN: made by Dako) to develop color. The results are shown in Figure 2. In the IHC method, it was confirmed that the GCIY cell line corresponds to an IHC score of 1+ and the HER2 protein is a low-to-medium performance strain.

(2) Subcutaneous transplantation model of human gastric cancer cell line in nude mice

In order to verify the usefulness of TS-1 and Herceptin (trastuzumab) in gastric cancer in combination with chemotherapy, an in vivo efficacy test was carried out in a nude mouse subcutaneous transplantation model of a human gastric cancer cell line. The preparation of the nude mouse subcutaneous transplantation model was carried out in accordance with the method described in WO2011/152516, except that the human gastric cancer-derived cell line GCIY was used.

Human gastric cancer-derived GCIY tumors were transplanted subcutaneously in nude mice according to conventional methods. As the TS-1 (S-1) administration group, TS-1 (n=8) was administered orally at 10 mg/kg/day on days 1-28 (Day 1-28). As a Herceptin administration group, intraperitoneal administration of Hepatic on the 1st, 8th, 15th and 22nd day at a dose of 20mg/kg (as trastuzumab) (n=8). In addition, as the drug combination group, TS-1 and Hepatic (n=8) were administered in combination with the same administration amount, administration route, and administration schedule as those administered alone. The tumor volume (major axis x short axis ² ÷ 2) was calculated by measuring the long axis and the short axis of the tumor over the day using a digital vernier caliper, and the relative tumor volume of each group relative to the tumor volume of the grouping day was calculated (Relative Tumor Volume, RTV)). Simultaneously, body weight was measured as an indicator of side effects. By calculation, the reduction ratio of the average relative tumor volume of the administration group relative to the average relative tumor volume of the non-administered control group (n=8) was determined as the in vivo efficacy.

The daily variation of RTV is shown in Figure 3. On the 29th day, the effect of suppressing tumor growth alone was confirmed to be 29% in the case of TS-1 10 mg/kg/day, and 25% in the case of Herceptin alone. By using both, the inhibition effect is enhanced to 54%. Furthermore, for the efficacy of the single administration and the efficacy of the combined administration, a statistical test was performed according to the Intersection-Union test procedure, and the significance was confirmed. That is, it was confirmed that the combination of TS-1 (S-1) and Herceptin has a significant combined effect compared with the treatment of a single agent of TS-1 (S-1) and Herceptin. . Further, based on the synergistic effect interpretation using the fractional product concept, it can be presumed that the combined effect of the two is synergistic. In the discussion of the side effects of the body weight index, it was found that there was no body weight suppression in the combination of Herceptin and TS-1.

The above shows that patients with low expression of HER2 in the judgment method of the ToGA test which did not be administered to Herceptin because of the inability to expect additional effects (especially the judgment score of HER2 protein by the IHC method) For 1+ patients, Herceptin has an excellent additional effect relative to TS-1 (S-1).

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<120> Method of selecting chemotherapy for patients with gastric cancer

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Claims (25)

A method for predicting the therapeutic effect of chemotherapeutic therapy using a tegafur-giramin-oteracil potassium complex for gastric cancer patients, comprising the following steps (1) to (3): (1) the measurement is taken from the patient a step of expressing the amount of HER2 mRNA contained in the living sample; (2) a step of comparing the expression amount of the HER2 mRNA obtained in the above step (1) with a preset cut-off point, The corresponding cut-off point is the median of the expression amount of HER2 mRNA contained in the living body sample of the gastric cancer patient; and (3) the prediction step, which is the result of the comparison of the above step (2), when the HER2 mRNA is expressed When the amount is higher than or equal to the corresponding cut-off point, it is predicted that the chemotherapeutic treatment of the patient with the tegafur-giramin-oteracil potassium complex and the HER2 inhibitor will show the possibility of adequate therapeutic effect. High; and when the expression level of HER2 mRNA is lower than the corresponding cut-off point, it is predicted that chemotherapy for the patient who chooses to use tegafur-giramin-oteracil potassium alone will show sufficient therapeutic effect. The possibility is high. For example, in the method of claim 1, wherein the molar ratio of each active ingredient of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: otaceta potassium = 1:0.4: 1. The method of claim 1 or 2, wherein the HER2 inhibitor is trastuzumab. The method of any one of claims 1 to 3 wherein the chemotherapy is postoperative adjuvant chemotherapy. The method of any one of claims 1 to 4, wherein the gastric cancer patient is a patient whose score is 0 or 1+ by using the IHC method, or the HER2 protein by the IHC method. The interpretation was divided into 2+ and the patients who were negative by the interpretation of the HER2 gene by the ISH method were classified. An antitumor agent consisting of a tegafur-giramin-oteracil potassium complex and a HER2 inhibitor for treating a gastric cancer patient, and the gastric cancer patient has been in the patent scopes 1 to 5 It is predicted in any of the methods that the chemotherapy with the tegafur-giramin-oteracil potassium complex and the HER2 inhibitor may show a high therapeutic effect. For example, in the anti-tumor agent of claim 6, wherein the molar ratio of the active ingredients of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: oltipraz potassium = 1: 0.4:1. An antitumor agent according to claim 6 or 7, wherein the HER2 inhibitor is trastuzumab. An antitumor agent consisting of a tegafur-jimeicilin-oteracil potassium complex for treating a gastric cancer patient, and the gastric cancer patient has been in any one of claims 1 to 5 It is predicted in the method of the method that chemotherapeutic treatment with tegafur-giramin-oteracil potassium complex alone may show a high possibility of sufficient therapeutic effect. Such as the anti-tumor agent of claim 9 of the patent scope, wherein tegafur-jimei The molar ratio of each active ingredient of the pyridine-otarazide potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4:1. An antitumor agent according to any one of claims 6 to 10 for use in postoperative adjuvant chemotherapy. A method for treating gastric cancer, comprising the following steps (1) to (3): (1) a step of measuring the expression amount of HER2 mRNA contained in a living body sample of a gastric cancer patient; (2) the above step (1) a step of comparing the amount of expression of the obtained HER2 mRNA with a predetermined cut-off point, which is a median amount of expression of HER2 mRNA contained in a living sample of a gastric cancer patient; and (3) a step of performing chemotherapy, which is based on the comparison of the above step (2), when the expression amount of HER2 mRNA is higher than or equal to the corresponding cut-off point, then tegafur-giramin-otelasi is used in combination Chemotherapy of potassium complexes with HER2 inhibitors. For example, in the method of claim 12, wherein the molar ratio of the active ingredients of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: otacetat potassium = 1:0.4: 1. A method for treating gastric cancer, comprising the following steps (1) to (3): (1) a step of measuring the expression amount of HER2 mRNA contained in a living body sample of a gastric cancer patient; (2) the above step (1) The step of comparing the amount of expression of the obtained HER2 mRNA with a preset cut-off point, which is the expression of HER2 mRNA contained in a living sample of a gastric cancer patient The median of the amount; and (3) the step of performing chemotherapy, which is based on the comparison of the above step (2), when the amount of HER2 mRNA expression is lower than the corresponding cut-off point, the patient is used alone. Chemotherapy of tegafur-gibberel-otarazide potassium complex. For example, in the method of claim 14, wherein the molar ratio of the active ingredients of the tegafur-giramin-oteracil potassium complex is tegafur: gemcitabine: otacetat potassium = 1:0.4: 1. The method of claim 14 or 15, wherein the HER2 inhibitor is trastuzumab. The method of any one of claims 12 to 16, wherein the chemotherapy is postoperative adjuvant chemotherapy. The method according to any one of claims 12 to 17, wherein the gastric cancer patient is a patient whose score is 0 or 1+ by using the IHC method, or the HER2 protein by the IHC method. The interpretation was divided into 2+ and the patients who were negative by the interpretation of the HER2 gene by the ISH method were classified. The use of a tegafur-jimeicilin-oteracil potassium complex and a HER2 inhibitor for the manufacture of an antitumor agent for treating gastric cancer patients, and the gastric cancer patient has been in the patent scopes 1 to 5 It is predicted in any of the methods that the chemotherapeutic treatment with the tegafur-giramin-oteracil potassium complex and the HER2 inhibitor may show a high therapeutic effect. For example, the application of the scope of claim 19, wherein tegafur-giramin- The molar ratio of the active ingredients of the oltipraz potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4:1. The use of claim 19 or 20, wherein the HER2 inhibitor is trastuzumab. A use of a tegafur-giramin-oteracil potassium complex for the manufacture of an antitumor agent for treating gastric cancer patients, and the gastric cancer patient has been in any of claims 1 to 5 It is predicted in the method of the method that chemotherapeutic treatment with tegafur-giramin-oteracil potassium complex alone may show a high possibility of sufficient therapeutic effect. For example, in the application of the scope of claim 22, wherein the molar ratio of the active ingredients of the tegafur-giramin-otarazide potassium complex is tegafur: gemcitabine: oltipraz potassium = 1:0.4: 1. The use of any one of claims 19 to 23 for postoperative adjuvant chemotherapy. An agent for predicting a therapeutic effect on chemotherapy for a gastric cancer patient, wherein the chemotherapy uses a tegafur-giramin-oteracil potassium complex, the reagent comprising: consisting of the base sequence represented by SEQ ID NO: The primer, the primer consisting of the nucleotide sequence shown in SEQ ID NO: 4, and the probe consisting of the nucleotide sequence shown by SEQ ID NO: 5.