WO2007086605A1

WO2007086605A1 - Method for prediction of the effect of sulfonamide compound

Info

Publication number: WO2007086605A1
Application number: PCT/JP2007/051747
Authority: WO
Inventors: Yoichi Ozawa
Original assignee: Eisai R & D Management Co., Ltd.
Priority date: 2006-01-27
Filing date: 2007-01-26
Publication date: 2007-08-02

Abstract

Disclosed is a method for prediction of the anti-tumor effect of a sulfonamide compound. The anti-tumor effect of a sulfonamide compound can be predicted by measuring the amount of neuron-specific enolase and determining the anti-tumor effect based on the amount of neuron-specific enolase.

Description

Method for predicting the effects of sulfonamide compounds

Technical field

The present invention relates to a novel method for predicting the antitumor effect of a sulfonamide compound.

Background art

In recent years, sulfonamide compounds have been reported as useful antitumor agents ( ¹⁴ ⁾ .

Food

In particular, N— (3-Chloro-1H-Indanol _7—yl) 1,1,4-Dibenzenedisulfonamide (hereinafter sometimes referred to as “E7070”), N— (3-Cyanol 4-methinole 1 H— 1-Cyanbenzenesolephoneamide (hereinafter sometimes referred to as “E7820”), N — [[((4-Chroophenyl) amino] carbonyl] 1,2,3-Dich Draw 1 H—Indene 1-5—Sulfonamide (hereinafter sometimes referred to as “: LY186641”), N— [[(3, 4 1 'Dicrophine fenenole) amino] Canoleponyl] 1, 2, 3—Dighi Drobenzofuran 5—Snorephone amide (hereinafter sometimes referred to as “LY295501”), N— (2,4-dichloro benzoyl) one 4—Black sulfonyl sulfone amide (hereinafter “LY-ASAP”) ), N— ( 2, 4-diclonal benzoyl) 1-5-promotiofen _ 2-sulfonamide (hereinafter sometimes referred to as “LY573636”), 2-sulfanilamide ₅ —black mouth quinoxaline (hereinafter referred to as rCQS l) Are very useful, showing activity against various types of tumors.

By the way, determining the effect of the sulfonamide compound, determining the effective concentration of the sulfonamide compound, and predicting the effect of the sulfonamide compound before administration are effective in promoting treatment with the sulfonamide compound. It is very useful to contribute to improving patient QOL. There are many studies on the former two. Specifically, sulfonamide is used with various gene expression changes as indicators. Method of assaying compound sensitivity ( ⁵⁾ , Method of determining the effect of a compound on integrin expression in cells other than platelets by measuring the expression level of integrin in platelets of patients receiving the compound ( ⁶ ) has been reported. On the other hand, no effective method has yet been found for predicting the effects of sulfonamide compounds before administration or compound treatment. Predicting the effects of a sulfonamide compound prior to administration is very beneficial and important for avoiding ineffective drug administration and reducing side effects for patients undergoing treatment. .

References

(1) Japanese Patent Laid-Open No. 7-165 708

(2) International publication 00/50395 pamphlet

(3) European Patent Application Publication No. 0222475

(4) International Publication No. 02/098848 Pamphlet

(5) International Publication No. 02/42493 Pamphlet

(6) International Publication No. 02Z066073 Pamphlet Disclosure of Invention

The present invention has been made in view of such circumstances, and the problem to be solved is to find a method for predicting the antitumor effect of a sulfonamide compound.

As a result of intensive investigations to solve the above problems, the present inventors have found that the antitumor effect of E7070, a sulfonamide compound, is neuron specific enolase (hereinafter referred to as `` NSE '') in serum opioma. It was found for the first time that it correlates with the quantity. It was found that the antitumor effect of E7070 can be predicted by measuring the NSE amount and using the NSE amount as an index.

In addition, in experiments with DNA microarrays and cancer cell line panels, E7070, E7820, LY186641, LY295501, LY573636 or CQS, or combinations thereof, showed high correlation between gene variation pattern and cell growth inhibitory activity. And found. In addition, in an assay for measuring cytostatic activity, it was found that cancer cell lines resistant to E7070 showed cross resistance to E7820, LY186641, LY295501, LY-ASAP, LY573636, or CQS. Based on these results, the present inventor has found that E7070, E7820, LY186641, LY295501, LY-ASAP, LY573636 or CQS have the same or similar mechanism of action, and have the same or similar genetic changes and The knowledge that it brings about an effect was acquired.

Therefore, the antitumor effect of a sulfonamide compound, preferably E7070, E7820, LY186641, LY295501, LY-ASAP, LY573636, or CQS, or a combination thereof is predicted by measuring the NSE amount and using the NSE amount as an index. I found out that In other words, in cancer patients, when the amount of NSE was high, it was possible to judge that cancer patients were highly sensitive to sulfonamide compounds.

That is, the present invention relates to the following.

(1) A method for predicting the antitumor effect of a sulfonamide compound, comprising:

Measuring NSE amount;

Determining whether a cancer patient is highly sensitive to a sulfonamide compound using the amount of NSE as an index;

Including the method,

Sulfonamide compounds are

Formula (I)

[Where

A ring is an optionally substituted monocyclic or bicyclic aromatic ring, Ring B is an optionally substituted 6-membered cyclic unsaturated hydrocarbon or an unsaturated 6-membered heterocycle containing one nitrogen atom as a heteroatom,

Ring C may have a substituent, a 5-membered heterocycle containing 1 or 2 nitrogen atoms,

W is a single bond or —CH═CH—,

X is one N (Ri) —or oxygen atom,

Y — C I 3

(R ³ ) —or I—

Z means one N (R2) —

Here, Ri, R ² and R 3 are each independently the same or different and represent a hydrogen atom or a lower alkyl group. ]

A compound represented by

Formula (II)

[Where E is one O—, _N (CH ₃ ) one, one CH ₂ —, _CH2CH ₂ — or _CH ₂ O1, D is one CH ₂ — or one O—, R ^la is , A hydrogen atom or a halogen atom, and R ^2a means a halogen atom or a trifluoromethyl group, respectively. ]

A compound represented by

General formula (III)

[Wherein J represents one O— or _NH—, Ri ^b represents a hydrogen atom, a halogen atom, May have a substituent CI- C ₆ alkyl group, but it may also have a substituent CI- C ₄ alkoxy group, which may have a substituent CI- C ₄ alkylthio O group, 1 CF ₃ , 1 OCF ₃ , 1 S CF ₃ , Ci 1 C ₄ alkoxycarbonyl group optionally substituted, nitro group, azide group, —O (S 0 2) CH ₃ , — N (CH ₃ ) 2, hydroxyl group, phenyl group, substituted phenyl group, pyridinyl group, chenyl group, furyl group, quinolinyl group or triazole group

R ² is a hydrogen atom, a halogen atom, a cyano group, — CF ₃ , an optionally substituted Ci-C ₆ alkyl group, an optionally substituted Ci_C4 alkoxycarbonyl group, or a substituted group. Ci—C4 alkoxy group which may have a group, a phenyl group which may have a substituent or a quinolinyl group which may have a substituent, R ^3b is a hydrogen atom or a substituent D—C ₄ alkoxy group which may have a hydrogen atom or a C 1 C ₆ alkyl group which may have a substituent (provided that at least one of R ^3b and is a hydrogen atom R ^5b is a hydrogen atom, a halogen atom, an optionally substituted Ci 1 C ₆ alkyl group, 1 CF ₃ or a nitro group, R ^6b is a hydrogen atom, a halogen atom or substituent has good Ci one also be C ₆ alkyl group (provided that the R6b substituent When you may CI- C ₆ alkyl group, R ⁵ b is water atom, a R ^7b is a halogen atom), R ^7b represents a halogen atom, which may have a substituent Ci 1 C ₆ alkyl group or 1 CF ₃ (wherein either R ^5b or R ^{7b is} an optionally substituted Ci—C ₆ alkyl group, or R? B force halogen) In the case of a d—C ₆ alkyl group which may have an atom or a substituent, either R ^5b or R ^6¾ is a hydrogen atom. ]

A compound represented by

Formula (IV)

And compounds represented by

Formula (V)

The method as described above, which is at least one compound selected from the group consisting of the compounds represented by: or a pharmacologically acceptable salt thereof, or a solvate thereof.

) Sulfonamide compounds

Ν— (3-Chloro 1 H—Indanol 7-Inole)-1, 4, 1 Benzene disulfonamide,

Ν— (3-Cyanol 4-methyl-1 1-indole 7-yl) 1 3-Cyanobenzene sulphonone amide,

Ν— [[(4-Cyclophenyl) amino] carbonyl]-2, 3-Dihydr draw 1 Η-Indene 1-5-sulfonamide,

Ν— [[(3,4-diphenyl) amino] carbonyl] 1,2,3-dihydrobenzofuran 5 —snolehonamide,

Ν— (2, 4-dichrome mouth Benzoinole)

Ν— (2,4-Dichlorobenzene) 1-Bromothiophene 2-Sonole Honamide

and

2-sulfanilamide 1-black mouth quinoxaline

At least one compound selected from the group consisting of: The method according to (1), wherein the salt is a pharmaceutically acceptable salt or a solvate thereof. (3) The sulfonamide compound is N— (3-chloro-1 H-indole-17-yl) 1,4 monobenzenedisulfonamide, or a pharmaceutically acceptable salt thereof, or a solvate thereof. The method according to (1), wherein the method is a product.

(4) The sulfonamide compound is N— (3-cyan-4-methyl-1H-india 1-ro 7-yl) 1-3-cyanobenzenesulfonamide, or a pharmaceutically acceptable salt thereof, or The method according to (1), which is a solvate thereof.

(5) The sulfonamide compound is N — [[(3,4-dichlorophenol) amino] canolepol] 1,2,3-dihydrobenzofuran 5-sulfone amide and N— (2,4-dichloro mouth (Benzoyl) at least one compound selected from the group consisting of 1-bromothiophene 1-2-sulfonamide, or a pharmacologically acceptable salt thereof, or a solvate thereof. The method described.

(6) The method according to (1), wherein the sulfonamide compound is a sodium salt of N— (2,4-dichlorodibenzo) -5-bupomochiofen-2-sulfonamide.

(7) The method according to (1), wherein the amount of NSE is the amount of NSE in serum.

(8) The method according to (1), wherein the NSE amount is the amount of NSE in the tumor.

(9) The method according to (1), wherein the NSE amount is measured by an immunochemical method.

(10) A kit for use in the method according to any one of (1) to (9),

The kit comprising an anti-NSE antibody. The present invention provides a method for predicting the antitumor effect of a sulfonamide compound.

More specifically, the antitumor effect of a sulfonamide compound is determined by measuring the amount of NSE. It was possible to predict by using the NSE amount as an index. In other words, in cancer patients, when the amount of NSE was high, it was possible to judge that cancer patients were addictive to sulfonamide compounds.

The method according to the present invention makes it possible to predict an antitumor effect without administering a sulfonamide compound to a patient, so that a patient who can expect a more antitumor effect can be selected and treated. It has become possible to contribute to the patient's QOL. Brief Description of Drawings

Figure 1 shows the correlation between the antitumor effect of E7070 and the serum NSE level in a human cancer cell line subcutaneous transplantation model.

Figure 2 shows the correlation between the antitumor effect of E7070 and the amount of NSE in the tumor in a human cancer cell line subcutaneous transplantation model.

Figure 3 shows the results of hierarchical clustering analysis on the DNA microarray in Reference Example 1.

FIG. 4 shows the correlation coefficient in the DNA microarray in Reference Example 2. Figure 5 shows the results of hierarchical clustering analysis on the DNA microarray in Reference Example 2.

FIG. 6 shows the correlation coefficient in the DNA microarray in Reference Example 2. Figure 7 shows the results of hierarchical clustering analysis on the DNA microarray in Reference Example 2.

Figure 8 shows the growth inhibitory effects of E7070, E7820, CQS, LY186641, LY295501 and LY-ASAP on HCT116-C9, HCT116-C9-C1 and HCT116-C9-C4 in the assay to measure cytostatic activity. Is.

FIG. 9 shows the growth inhibitory action of E7070 and LY573636 on HCT116-C9, HCT116-C9-C1 and HCT116-C9-C4 in the assay for measuring cytostatic activity. BEST MODE FOR CARRYING OUT THE INVENTION Embodiments of the present invention will be described below. The following embodiment is an example for explaining the present invention, and is not intended to limit the present invention only to this embodiment. The present invention can be implemented in various forms without departing from the gist thereof.

Documents cited in the present specification, as well as patent publications, patent gazettes and other patent documents are incorporated herein by reference. This specification also includes the disclosure content of Japanese Patent Application JP2006-018912, which is the basis of the priority claim of the present application. The present invention relates to a method for predicting the antitumor effect of a sulfonamide compound, comprising a step of measuring the amount of NSE and whether or not a cancer patient is highly sensitive to a sulfonamide compound using the amount of NSE as an index. And providing the method.

1. Process of measuring NSE amount

Neuron Specific Enolase (hereinafter also referred to as “NSE” or “γ-Enolase”) is a glycolytic enzyme enolase that forms a dimeric structure having a γ subunit, In normal tissue, it exists in nerve tissue and neuroendocrine cells. NSE has also been reported to be localized in lung cancer, neuroblastoma, breast cancer, stomach cancer, and colon cancer tissue (Hiroshi Ariyoshi, et al .: Neuron-specific enolase (NSE). Cancer and chemistry. Therapy 10: 1744-: 1753, 1983). Furthermore, the amount of serum NSE is used as a tumor marker for small cell lung cancer (Ariyoshi Y, et al: Evaluarion of serum neuron-specinc enolase as a tumor marker for carcinoma of the lng. Gann 74: 219-225 , 1983).

NSE can include, for example, a polypeptide containing the amino acid sequence represented by SEQ ID NO: 2 (GenBank accession number: NM_001975). In the present invention, NSE has one or more (for example, one or several) amino acids deleted, substituted or added in the amino acid sequence represented by SEQ ID NO: 2, or their Polypeptides containing amino acid sequences mutated by combinations Chido is also included.

In the amino acid sequence represented by SEQ ID NO: 2, an amino acid sequence in which one or more (for example, one or several) amino acids are deleted, substituted or added, or mutated by a combination thereof, Examples of the polypeptide to be included include (i) 1 to 9 (for example, 1 to 5, preferably 1 to 3, more preferably 1 to 2) in the amino acid sequence represented by SEQ ID NO: 2. More preferably 1) amino acid sequence lacking amino acid,

(ii) 1 to 9 (for example, 1 to 5, preferably 1 to 3, more preferably 1 to 2, more preferably 1) of the amino acid sequence represented by SEQ ID NO: 2 Amino acid sequence added with amino acid,

(iii) 1 to 9 amino acids in the amino acid sequence represented by SEQ ID NO: 2 (for example, 1 to 5, preferably 1 to 3, more preferably 1 to 2, more preferably 1) An amino acid sequence in which no acid is substituted with another amino acid,

(iv) an amino acid sequence mutated by the combination of (i) to (iii) above,

And the like. As used herein, an amino acid “deletion” means a mutation in which one or more of the amino acid residues in the sequence has been deleted. In the deletion, an amino acid residue deleted from the end of the amino acid sequence and Those in which an amino acid residue in the middle of the amino acid sequence is deleted are included.

Here, “addition” of an amino acid means a mutation in which one or more amino acid residues are added to the sequence. For addition, an amino acid residue is added to the end of the amino acid sequence and the amino acid sequence Those with amino acid residues added in the middle are included. Anything added along the way can be called “insertion”.

Here, “substitution” of an amino acid means a mutation in which one or more amino acid residues in the sequence are changed to different kinds of amino acid residues.

Furthermore, the NSE to be measured can also be measured using these partial peptides in addition to the polypeptide containing the amino acid sequence represented by SEQ ID NO: 2 or a variant thereof. As a partial peptide, for example, amino acids constituting NSE Among the sequences, peptides containing 6 or more, preferably 8 or more amino acid sequences can be mentioned.

The amount of NSE is the amount of NSE in a living body or in a test sample collected from a living body (for example, in a test sample collected from a cancer patient). For example, the amount of NSE in blood, serum or plasma and in a tumor The amount of NSE is exemplified, and the amount of NSE in the tumor is more preferable.

The blood is preferably blood collected from cancer patients. Blood can be collected from cancer patients according to conventional methods. Serum and plasma can be obtained from blood collected according to conventional methods.

The tumor is preferably a tumor removed from a cancer patient. The tumor can be obtained, for example, by removing it from a cancer patient by a surgical procedure (for example, biopsy).

The size of the tumor collected from cancer patients only needs to be large enough to measure the amount of NSE.

The types of tumors are, for example, brain tumor, cancer, esophageal cancer, tongue cancer, lung cancer, breast cancer, knee cancer, stomach cancer, colon cancer, small intestine or duodenal cancer, colon cancer, rectal cancer, bladder cancer, kidney cancer, liver cancer , Prostate cancer, uterine cancer, ovarian cancer, thyroid cancer, gallbladder cancer, pharyngeal cancer, sarcoma (eg, osteosarcoma, chondrosarcoma, force positive sarcoma, muscle tumor, angiosarcoma, fibrosarcoma, etc.), leukemia (eg, chronic bone marrow) Leukemia (CML), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL) and acute lymphocytic leukemia (ALL), lymphoma, multiple myeloma (MM), etc. and melanoma Preferred are colon cancer, breast cancer, small cell lung cancer and sputum cancer, and more preferred are small cell lung cancer.

The amount of NSE can be analyzed, for example, by measuring the amount of protein.

Methods for measuring NSE protein include, for example, immunochemical methods (eg, immunohistochemical methods, Western blot, ELISA (enzyme-linked immunosorbent assay), EIA enzyme immunoassay), RIA (radioimmunoassay; etc.), mass spectrometry The method is preferably an immunochemical method, particularly preferably Preferred is ELISA. These methods can be carried out according to conventional methods.

The amount of NSE can be analyzed, for example, by measuring the amount of mRNA expressed in tumor cells.

For example, NSE mRNA can be measured by in situ hybridization, Northern blot analysis, DNA microarray, RT "PCR, etc., and preferably: These methods can be performed according to conventional methods. it can.

The amount of NSE can be calculated, for example, as the amount of NSE per unit volume in blood, serum or plasma, the amount of NSE per unit volume in tumor, and the amount of NSE per unit weight. It can also be calculated as the amount of NSE per unit tumor volume, tumor weight or total protein in blood, serum or plasma. In addition, the amount of NSE can be calculated using the expression level of a protein that is constantly expressed in cells as an index. Examples of proteins constitutively expressed in cells include 3-actin and GAPDH. The expression level of the protein that is constitutively expressed in the cells can be calculated in the same manner as the method for measuring the NSE level.

In this case, the NSE amount can be calculated as the NSE amount per expression level of the protein that is constantly expressed in the cells.

Furthermore, the amount of NSE in the test sample can be calculated using the amount of NSE in the control sample as an index. In this case, the amount of NSE in the test sample can be calculated as a relative value to the amount of NSE in the control sample. The control sample to be used is not particularly limited, and any sample can be used.

2. The process of determining whether cancer patients are highly sensitive to sulfonamide compounds.

The present invention predicts the antitumor effect of a sulfonamide compound by measuring the amount of NSE and using the amount of NSE as an index. That is, in this step, it can be determined whether or not it is highly sensitive to the sulfonamide compound using the NSE amount in the living body or in the test sample collected from the living body as an index. The antitumor effect of the sulfonamide compound can be predicted from the sensitivity determination result. The amount of NSE is, for example, the amount of NSE per unit volume in blood, serum or plasma, the amount of NSE per unit volume in the tumor, the amount of NSE per unit weight, and the expression of proteins that are normally expressed in cells. The amount of NSE per unit can be used as an index. In addition, the amount of NSE per unit tumor volume in blood, serum or plasma can be used as an index.

If the amount of NSE is high, it can be judged that the cancer patient is highly sensitive to the sulfonamide compound. On the other hand, if the amount of NSE is small, it can be determined that the cancer patient is not highly sensitive to the sulfonamide compound.

The case where the amount of NSE is large is, for example, 1.2 times or more, preferably 1.5 times or more, more preferably 2 times or more, more preferably compared to the average amount of NSE in the blood, serum or plasma of cancer patients. Is 3 times or more, or 1.2 times or more, preferably 1.5 times or more, more preferably 2 times or more, more preferably 3 times or more, compared to the average amount of NSE in the tumor of a cancer patient. I can say that.

Here, the average NSE amount of cancer patients can be determined by averaging the NSE amounts measured in a plurality of cancer patients. Cancer patients whose NSE amount is measured may be extracted randomly, or may be extracted based on factors such as the type of cancer affected, the degree of progression of symptoms, sex, and age.

In addition, the case where the amount of NSE is large is, for example, 1.5 times or more, preferably 2 times or more, more preferably 3 times or more, compared to the average NSE amount in the blood, serum or plasma of a healthy person. Preferably, the case of 4 times or more can be mentioned.

The average NSE amount of healthy individuals can be determined by averaging the NSE amounts measured in multiple healthy subjects. Healthy individuals whose NSE amount is measured may be extracted randomly or based on factors such as gender and age. In addition, the amount within the standard value of the clinical test can be used as a reference for comparison.

Furthermore, when the amount of NSE is large, for example, the amount of NSE in blood, serum or plasma is 6 ng / ml or more, preferably 10 ng / ml or more, more preferably 20 ng / ml or more, more preferably The case of 50 ng / ml or more can be said.

Also, when there is a large amount of NSE, for example, in the blood, serum or plasma of cancer patients Statistically significant compared to the average amount of NSE in the blood, serum, or plasma of a healthy person if there is a statistically significant difference compared to the average amount of NSE in the medium or tumor It can be said that there are many differences.

In the present invention, the antitumor effect is predicted mainly for the purpose of predicting the antitumor effect of a sulfonamide compound in cancer patients before administration.

When a cancer patient is judged to be highly sensitive to a sulfonamide compound, the sulfonamide compound can be predicted to have a more antitumor effect. For example, if you can expect a higher antitumor effect than the average antitumor effect in patients with similar symptoms, you can expect a higher antitumor effect than other patients with the same cancer type. Can be expected, or can be expected to have a higher antitumor effect than patients suffering from other cancer types.

In addition, as described later, since the sulfonamide compound in the present invention has an antitumor effect, even when it is determined that a cancer patient is not highly sensitive to the sulfonamide compound, the sulfonamide compound is used. Is not expected to have no anti-tumor effect.

3. Sulfonamide compounds

In the present invention, the sulfonamide compound includes a compound represented by the following general formula (I).

In the above general formula (I),

W is a single bond or one CH = CH—

X is one N (Ri) one or oxygen atom,

Y is C I 3

— (R ³ )-or one I——

Z means one N (R2) one.

Here, Ri, R ² and R ³ are independently the same or different and each represents a hydrogen atom or a lower alkyl group.

In the above general formula (I), the “monocyclic or bicyclic aromatic ring optionally having a substituent” in the A ring means an aromatic hydrocarbon, a nitrogen atom, an oxygen atom An aromatic heterocyclic ring containing at least one of the sulfur atoms, and having 1 to 3 substituents on the ring. Examples of the main aromatic ring contained in the A ring include pyrrole, pyrazole, imidazole, thiophene, furan, thiazol. Oxazole, benzene, pyridine, pyrimidine, pyrazine, pyridazine, naphthalene, quinoline, isoquinoline, phthalazine, There are naphthyridine, quinoxaline, quinazoline, cinnoline, indole, isoindole, indolizine, indah zonole, benzofuran, benzothai phen, benz aged zonazore, benzimidazole, benzopyrazole, benzothiazole, etc. Is not limited to these. The aromatic ring may have 1 to 3 substituents, and when there are a plurality of substituents, they may be the same or different. Examples of the substituent include an amino group optionally substituted with a lower alkyl group or a lower cycloalkyl group, a lower alkyl group, a lower alkoxy group, a hydroxyl group, a nitro group, a mercapto group, a cyano group, a lower alkylthio group, a halogen atom. Atom, Formula 1 a— b [where a is a single bond, 1 (CH ₂ ) _k —, −0- (CH ₂ ) k, 1 S— (CH ₂ ) k 1 Or N (R3)-(CH ₂ ) k-, k is an integer from 1 to 5, R3 is a hydrogen atom or lower alkyl group, b is one CH ₂ -d (where d is lower Means an amino group optionally substituted with an alkyl group, a halogen atom, a hydroxyl group, a lower alkylthio group, a cyano group or a lower alkoxy group)], a group represented by the formula 1 a-e 1 f [formula Wherein a is as defined above, e is 1 S (O) 1 or 1 S (O) ₂ , f is an amino group optionally substituted by a lower alkyl group or a lower alkoxy group, Alkyl group, trifluoromethyl group, one (CH ₂ ) _m — b or one N (R4) — (CH ₂ ) _m -b (wherein b is as defined above, R 4 is a hydrogen atom or lower An alkyl group, m means an integer of 1 to 5), a group represented by the formula: a—g_h [wherein a is as defined above, and g is _C. O) — or one C (S) one, h is an amino group optionally substituted with a lower alkyl group, a hydroxyl group, a lower alkyl group, a lower alkoxy group, one (CH ₂ ) _n — b or N (R 5)-(CH ₂ ) n- b (wherein b is as defined above, R 5 is a hydrogen atom or a lower alkyl group, and n is an integer of 1 to 5) A group represented by the formula: a—N (R6) -g-i [wherein a and g have the same meaning as described above, R6 represents a hydrogen atom or a lower alkyl group, i represents a hydrogen atom, A lower alkoxy group or a group represented by f (ί has the same meaning as described above), a formula N a— N (R ⁷ )-e-f (where a, e and f are as defined above) Meaning a hydrogen atom or a lower alkyl group), or a group represented by the formula (CH ₂ ) p-j-(CH ₂ ) q_ b (wherein j represents an oxygen atom or a sulfur atom) B means And P and q are the same or different and each represents an integer of 1 to 5).

In the above substituent examples, when the amino group is substituted with two alkyl groups, these alkyl groups may be bonded to form a 5- or 6-membered ring. Further, when the A ring is a nitrogen-containing heterocycle having a hydroxyl group or a mercapto group, these groups may take the form of an oxo group or thixo group by taking a resonance structure.

In general formula (I), the meaning of “ring B” is “optionally substituted, 6-membered ring “Unsaturated 6-membered heterocycle containing one nitrogen atom as an unsaturated hydrocarbon or heteroatom” is, for example, benzene or pyridine, in which part of the unsaturated bond may be hydrogenated, One or two or more substituents may be present on the ring, and when there are two or more substituents, they may be the same or different.

The meaning of the C ring “optionally substituted, 5-membered heterocycle containing 1 or 2 nitrogen atoms” means that a part of the unsaturated bond may be hydrogenated, pyrrole, Pyrazole and imidazole, which may have 1 or 2 substituents on the ring, and when there are 2 substituents, may be the same or different.

Examples of the substituent that the ring B and the ring C may have include, for example, a halogen atom, a cyano group, a lower alkyl group, a lower alkoxy group, a hydroxyl group, an oxo group, Formula 1 C (O) -r (formula Wherein r represents a hydrogen atom, an amino group which may be substituted with a lower alkyl group, a lower alkyl group, a lower alkoxy group or a hydroxyl group), an amino group which may be substituted with a lower alkyl group, Examples thereof include, but are not limited to, an chloromethyl group.

In the general formula (I), Z means 1 N (R ² ) —. R ² is independently the same as or different from R i and represents a hydrogen atom or a lower alkyl group.

In the general formula (I), Y is

—C (R ³ ) — or one —. In the above formula, R ³ represents a hydrogen atom or a lower alkyl group.

In the above general formula (I), “lower alkyl group” in the definition of the substituents that R ¹ , R ² and R ³ and the A ring, B ring and C ring may have has 1 to 6 carbon atoms. A straight chain or branched alkyl group of, for example, but not limited to, a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, sec -Butyl group, tert-butyl group, n-pentyl group (amyl group), isopentyl group, neopentyl group, tert-pentyl group, 1-methylbutyl Group, 2-methylbutyl group, 1,2-dimethylpropyl group, _n- hexyl group, isohexyl group, 1-methylpentyl group, 2-methylpentyl group, 3-methylpentyl group, 1-ethylpropyl group, 1, 1 —Dimethylptyl group, 1,2-dimethylbutyl group, 2,2-dimethylbutyl group, 1,3-dimethylbutyl group, 2,3-dimethylbutyl group, 3,3-dimethylbutyl group, 1-ethylbutyl group, 2 — It means an ethylbutyl group, 1,1,2-trimethylpropyl group, 1,2,2-trimethylpropyl group, 1-ethyl-1-monomethylpropyl group, 1-ethyl-2-methylpropyl group, and the like. Among these, preferred groups include a methyl group, an ethyl group, an 11-propyl group, an isopropyl group, an n-butyl group, an isoptyl group, a sec-butyl group, and a tert-butyl group. Among them, the most preferred group includes a methyl group, an ethyl group, an n-propyl group, and an isopropyl group. The “lower cycloalkyl group” in the definition of the substituent that the A ring may have is a carbon A cycloalkyl group having a number of 3 to 8 means, for example, a cyclopropyl group, a cyclopentinole group, a cyclopentinole group, a cyclohexyl group, a cyclohexynole group, a cyclooctyl group, etc. It is not limited to. Further, the “lower alkylthio group” means an alkylthio group derived from the above lower alkyl group, for example, a methylthio group, an ethylthio group, an n-propylthio group, an isopropylthio group, an n-butylthio group, an isobutylthio group, Examples thereof include, but are not limited to, sec-butylthio group and tert-butylthio group.

The “lower alkoxy group” in the definition of the substituent that the A ring, the B ring, and the C ring may have is not limited to these, but includes, for example, a methoxy group, an ethoxy group, n-propoxy group, isopropoxy group, n-butoxy group, isobutoxy group, sec-butoxy group, tert-butoxy group and the like, which means a lower alkoxy group derived from the above lower alkyl group. Examples thereof include a methoxy group and an ethoxy group. Examples of the “halogen atom” include a fluorine atom, a chlorine atom, a bromine atom, and an iodine atom.

The compound represented by the general formula d) of the present invention can be produced by a known method, for example, It can be produced by the method described in WO 95/07276 pamphlet (WO95 / 07276) and Z or JP-A-7-165708 (JP7-165708).

— In general formula (I), preferred compounds are E7070 or E7820.

E7070 refers to N— (3-Chloro-1H-indole-7-yl) 1-1,4-benzenedisulfonamide, and its structural formula is shown in the following formula (VI).

E7070 can be produced by a known method, and is described in, for example, Example 19 in WO95 / 07276 pamphlet (WO95 / 07276) and / or JP-A-7-165708 (JP7-165708). Can be manufactured by different methods.

Note that E7070 can also be reconstituted with N— (3-Chrome 1 H-indole-7-yl) — 4 1 Sulfone amide.

E7820 refers to N— (3-cyano-4-methyl-1H-indole-7-yl) 1-3-cyanobenzenesulfonamide, and its structural formula is shown in the following formula (VII).

E7820

E7820 can be produced by a known method, for example, by the method described in International Publication No. 00/50395 pamphlet (WO00 / 50395).

In the present invention, the sulfonamide compound includes a compound represented by the following general formula (II).

In the general formula (II), E is 1 O—, —N (CHs), 1 CH ₂ —, 1 C ₂ CH ₂ — or 1 CH ₂ O—, and D is 1 CH ₂ — Or 1 O—, R ^la represents a hydrogen atom or a halogen atom (for example, a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom), and represents a halogen atom or a trifluoromethyl group, respectively.

The compound represented by the general formula (II) of the present invention can be produced by a known method, for example, by the method described in European Patent Application Publication No. 0222475A1 (EP0222475A1).

In general formula (II), a preferred compound is LY186641 or LY295501.

LY186641 refers to N-[[((4-Ciclophenyl) amino] carbonyl] 1 2 3-dihydro 1 H-indene 1-sulfonamide, and its structural formula is shown in the following formula (VIII).

LY186641

LY186641 can be produced by a known method. For example, LY186641 can be produced by the method described in European Patent Application Publication No. 0222475A1 (EP0222475A1). In the present invention, LY295501 refers to N — [[((3,4-diphenyl) phenyl) amino] canoleponi _re] 1,2,3-dihydrobenzofuran-5-snorephoneamide and its structural formula Is shown in the following formula (IX).

LY295501

LY295501 can be produced by a known method, for example, by the methods described in European Patent Application Publication No. 0222475A1 (EP0222475A1) and European Patent Application Publication No. 0555036A2 (EP0555036A2). In the present invention, the sulfonamide compound includes a compound represented by the following general formula (III).

In the general formula (III), J is 1O— or 1 NH—, Rib is a hydrogen atom, a halogen atom, or a d-C ₆ alkyl group which may have a substituent, a substituent. Ci 1 C ₄ alkoxy group which may have a substituent Ci 1 C ₄ alkylthio group which may have a substituent, 1 CF ₃ , 10 CF ₃ , 1 SCF ₃ , which has a substituent Ci 1 C ₄ alkoxycarbonyl group, nitro group, azide group, 10 (so ₂ ) C

H ₃ , —N (CH ₃ ) 2, hydroxyl group, vinyl group, substituted vinyl group, pyridinyl group, chenyl group, furyl group, quinolinyl group or triazole group, R ^2b is hydrogen An atom, a halogen atom, a cyan group, one CF ₃ , an optionally substituted Ci_C ₆ alkyl group, an optionally substituted Ci one C4 alkoxycarbonyl group, and a substituted group An optionally substituted d-Ca alkoxy group, an optionally substituted phenyl group or an optionally substituted quinolinyl group, an optionally substituted hydrogen atom or an optionally substituted group; Good d—C4 alkoxy group, R ^4b may have a hydrogen atom or a substituent d—C ₆ alkyl group (provided that at least one of R ³ b and R ⁴ b is a hydrogen atom) the at it), R ⁵ b represents a hydrogen atom, a halo gen atom, which may have a substituent group d Ce alkyl group, an CF ₃ or two Toro group, R ^6b represents a hydrogen atom, may have a halogen atom or a substituent Ci -C ₆ alkyl group (provided that when the R ^6¾ is optionally may d-Ce Al kills group having a substituent, R ^5b is a hydrogen atom, R? B is a halogen atom) and, R ? b is a halogen atom, which may have a substituent, or a C ₆ alkyl group or a CF ₃ (wherein either R ^5b or R ^7b may have a substituent) d — a C ₆ alkyl group, or R? b force, a halogen atom or a Ci—C ₆ alkyl group optionally having a ^substituent, either R ^5¾ or Each represents a hydrogen atom.

In the general formula (III), the “halogen atom” is preferably a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom.

In the general formula (III), “Ci 1 c ₆ alkyl group” has the same meaning as the above-mentioned “lower alkyl group” and is not particularly limited, but is preferably a methyl group, an ethyl group, n- Examples thereof include propyl group, isopropyl group, n-butyl group, isoptyl group, sec-butyl group, tert-butylene group, n-pentyl group, n-hexyl group and the like.

In the general formula (III), “Ci_C ₄ alkoxy group” means an alkoxy group having 1 to 4 carbon atoms in the above-mentioned “lower alkoxy group” and is not particularly limited, but preferably Examples thereof include a methoxy group, an ethoxy group, an n-propoxy group, an isopropoxy group, an n-butoxy group, an isoptoxy group, a sec-butoxy group, and a tert-butoxy group.

In the general formula (III), in "Ci one C ₄ alkylthio group", although the alkyl group but are not particularly limited, methyl, Echiru, propyl, isopropyl, n- butyl, isobutyl, sec- butyl, tert- butyl Etc. In the general formula (III), examples of “Ci-C4 alkoxycarbonyl group” are not particularly limited, but include methoxycarbonyl group, ethoxycarbonyl group, n-propoxycarbonyl group, isopropoxycarbonyl group. Nore group, n-butoxycarbonyl group, isobutoxycarbonyl group, sec-butoxycarbonyl group, tert-butoxycarbonyl group and the like can be mentioned.

In the general formula (III), the substituent to be introduced is not particularly limited. Without, for example, C i-C ₆ alkyl group (e.g., methyl group, Echiru group, n- propyl group, an isopropyl group, n- heptyl group, Isopuchiru group, sec- butyl group, tert- heptyl group), Ci-C4 alkoxy group (for example, methoxy group, ethoxy group, n-propoxy group, isopropoxy group, n-butoxy group, isobutoxy group, sec-ptoxy group, tert-butoxy group, etc.), amino group, hydroxyl group And a substituent such as a halogen atom (for example, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom) or a silyl group.

The compound represented by the general formula (III) of the present invention can be produced by a known method. For example, it can be produced by the method described in WO 02/098848, pamphlet (WO02 / 098848). .

In general formula (III), the preferred compound is LY-ASAP.

LY-ASA refers to N— (2,4-dichlorobenzoic) 14-monocyclic phenylsulfonamide, and its structural formula is shown in the following formula (X).

LY-ASAP

LY-ASAP can be produced by a known method. For example, LY-ASAP can be produced by the method described in International Publication No. 02/098848 Pan Fret (WO02 / 098848).

Furthermore, in the present invention, examples of the sulfonamide compound include LY573636. In the present invention, LY573636 refers to N- (2,4-dichlorobenzoyl) 15-bromothiophene-1-sulfonamide, and the structural formula is shown in the following formula (IV).

LY573636

LY573636 is preferably sodium salt. LY573636 can be produced by a known method. For example, in the same manner as described in WO 02/098848 (WO02 / 098848), a commercially available 5-bromothiophene 2-sulfuryl mouthpiece and LY573636 can be produced. It can be produced from 2,4-dichlorodibenzoic acid.

In the present invention, examples of the sulfonamide compound include CQS. In the present invention, CQS refers to 2-sulfanylamide and 5-quinoquinoline, and the structural formula is shown in the following formula (V).

CQS can be produced by a known method. For example, (J. Am. C em. Soc., 1947, 71, 6 ·

10).

Sulfonamide compounds may form pharmacologically acceptable salts with acids or bases. The sulfonamide compound in the present invention includes these pharmacologically acceptable salts. Examples of salts with acids include inorganic acid salts such as hydrochloride, hydrobromide, sulfate, and phosphate, formic acid, acetic acid, lactic acid, succinic acid, fumaric acid, maleic acid, citrate, tartaric acid, Mention may be made of salts with organic acids such as benzoic acid, methanesulfonic acid, benzenesulfonic acid, ρ-toluenesulfonic acid, trifluoroacetic acid. In addition, as salts with bases, alkali metal salts such as sodium salt and potassium salt, alkaline earth metal salts such as calcium salt and magnesium salt, trimethylamine, triethylamine, pyridine, picoline, dicyclohexylamine, Ν:塩, salts with organic bases such as monodibenzylethylenediamine, arginine and lysine (organic amine salts), and ammonium salts.

The sulfonamide compound may be an anhydride or may form a solvate such as a hydrate. The solvate may be either a hydrate or a non-hydrate, but a hydrate is preferred. Solvents are water, alcohol (eg, methanol, eta , N-propanol), dimethylformamide, and the like.

Further, when solvates and / or optical isomers of these compounds exist, the sulfonamide compounds in the present invention include those solvates and z or optical isomers. The sulfonamide compound in the present invention also includes a sulfonamide compound that undergoes metabolism such as oxidation, reduction, hydrolysis, and conjugation in vivo. Furthermore, the sulfonamide compound in the present invention also includes a compound that generates a sulfonamide compound by being subjected to metabolism such as oxidation, reduction, and hydrolysis in vivo.

4. Kit

The present invention provides a kit comprising an anti-NSE antibody for use in a method for predicting the antitumor effect of a sulfonamide compound. The antibody can be prepared according to a conventional method. A commercially available anti-NSE antibody may also be purchased. The kit of the present invention may contain components commonly used in general measurements in addition to the above-described antibodies. Further, the kit of the present invention includes an instruction manual, an attached document, etc. describing that the anti-NSE antibody is used in a method for predicting the antitumor effect of the sulfonamide compound, in addition to the above-described antibody. Also good. Example

Hereinafter, the present invention will be described with specific examples, but the present invention is not limited thereto.

[Example 1] Correlation between antitumor effect of E7070 and serum and tumor NSE levels in various human cancer cell line subcutaneous transplantation models

(1) Preparation of various human cancer cell line subcutaneous transplant models

Human colorectal cancer cell lines HCT116, SW620, HCT15, DLD-l, Colo320D.M. (Purchased from ATCC), WiDr (purchased from Dainippon Pharmaceutical), human non-small cell lung cancer cell line Lu-99 (hi Purchased from the Human Science Foundation), H460, Calu-1 (purchased from ATCC), PC-14 (transferred from RIKEN), human breast cancer cell line MDA-MB468 (purchased from ATCC), BSY-1, HBC- 4 (assigned from Cancer Research Center for Cancer Chemotherapy), human pancreatic cancer cell line PSN-1 (assigned from National Cancer Center), KP-4 (assigned from National Kyushu Cancer Center 1) at 37 ° C Then, the cells were cultured with RPMI1640 (including 10% FBS) in a 5% carbon dioxide incubator until they became about 80% confluent, and the cells were collected by trypsin-EDTA. Prepare a suspension of 8xl0 ⁷ cells / ml for HCT116 and 5xl0 ⁷ cells / ml for other cells using Hanks balanced solution, and 0.1 ml of the resulting cell suspension was transplanted subcutaneously into the body side of nude mice. . In addition, human small cell lung cancer cell line LX-1 (assigned from Cancer Research Center for Cancer Chemotherapy) was aseptically removed from a subcultured tumor subcutaneously in nude mice and cut into 2-3 mm squares. The tumor pieces were transplanted subcutaneously on the nude mouse side by trocar.

The model thus produced was used for the following experiment. (2) Antitumor effect of E7070

For the various human cancer cell line subcutaneous transplantation models prepared in (1) above, when the average tumor volume reached 100-200 mm ³ after transplantation, administration of E7070 (40 mg / kg · once a day · 5 Daily intravenous administration).

E7070 was produced by the method described in Example 19 in WO95 / 07276 pamphlet (WO95 / 07276) or JP-A-7-165708 (JP7-165708).

The tumor major axis and minor axis were measured with Digimatic Caliper (Mitsutoyo) at a frequency of twice / week from the start of administration, and the tumor volume was calculated using the following formula.

Tumor volume (mm ³ ) = tumor major axis (mm) x tumor minor axis ² (mm ² ) / 2

LoglO cell kill was adopted as an index of the antitumor effect and was calculated from the tumor growth curve using the following formula.

Logl0 cell kill = (Τχ 4 in E7070 administration group — Τ _{χ 4 in} control group) Tumor doubling time /3.32 T _{x 4} : Time (days) required for tumor to grow to 4 times the initial tumor volume

Table 1 shows the antitumor effects of Ε7070 against various human cancer cell lines. table 1

(3) For various human cancer cell lines subcutaneously various human cancer cell lines implanted subcutaneously model prepared in NSE amount of measuring the serum in transplant model (1), after transplantation, tumor瘍体product becomes 500 mm ³ or more Blood was collected at the time point (tumor volume 601-1865 mm ³ ), and serum was separated according to a conventional method. Thereafter, the amount of NSE of the serum was measured by ELISA using a bead solid phase method (requested from Koto Institute for Microbial Research). Tables 2 and 3 show the serum NSE levels in various human cancer cell line subcutaneous transplantation models. The value obtained by dividing the amount of NSE in serum by the tumor volume was obtained as the normalized amount of NSE in serum (Table 2). In addition, the average value of NSE content in the standardized serum of various human cancer cell lines was determined (Table 3).

Table 2

Table 3

The antitumor effect of E7070 was compared with the serum NSE level in six human cancer cell line subcutaneous transplantation models (Fig. 1). As a result, it was clarified that the antitumor effect of E7070 in various human cancer cell line subcutaneous transplantation models correlates with the amount of serum NSE (Fig. 1). Therefore, it was clarified that the antitumor effect of E7070 can be predicted by measuring the amount of NSE in serum and using that NSE amount as an index.

(4) Measurement of the amount of NSE in the tumor

Regarding the various human cancer cell line subcutaneous transplant models prepared in (1) above, the tumor was removed, the connective tissue was removed, and the sample was frozen in a tube at −80 ° C. to prepare a cryopreserved sample.

Tumor pieces were excised from cryopreserved samples of various tumors, and 6 L of Lysis Buffer (1M Tris-HCl pH 7.5, 10% Glycerol, 2 mM EDTA, ImM Sodium Orthovanadate, lx Complete EDTA-free (Roche )) Inside, solubilize the protein in the tumor using a homogenizer, Ruffer (trade name: Tris SDS β ME sample treatment solution (Daiichi Kagaku)) was added, treated at 95 ° C for 5 minutes, and used as a sample for SDS-PAGE. SDS-PAGE each lysate with A ^ u L / Lane, and then detect NSE by Western blotting using anti-NSE antibody (Neuron-Specific Enolase Clone BBS / NC / VI-H14, DAKO) did.

The NSE amount in each tumor was compared by comparing the intensity of each band with the intensity of the NSE panda in SW620 and taking the intensity of the NSE panda in SW620 as 1.0 (Table 4). Table 4

1 We compared the antitumor effect of E7070 with the amount of NSE in the tumor in four human cancer cell line subcutaneous transplantation models (Fig. 2). As a result, it was clarified that the antitumor effect of E7070 in various human cancer cell line subcutaneous transplantation models correlates with the amount of NSE in the tumor (Fig. 2). Therefore, it was clarified that the antitumor effect of E7070 can be predicted by measuring the amount of NSE in the tumor and using that NSE amount as an index.

[Reference Example 1] DNA microarray analysis

(1) Cell culture, compound treatment, and RNA extraction

For the purpose of examining gene expression changes induced by compounds by DNA microarray analysis, human colon cancer-derived cell line HCT116 (American Type Culture Collection, Manassas, VA, USA) and human leukemia-derived cell line MOLT-4 (American Tvpe Culture Collection, Manassas, VA, USA), 10% fetus The cells were cultured in RPMI-1640 medium supplemented with bovine serum, 100 units / ml penicillin, and lOO g / ml streptomycin. Hereinafter, the culture Contact Yopi compound treatment was performed in 5% C0 _2, 37 incubator adjusted to ° C. HCT116 cells and MOLT-4 cells were seeded at a rate of 2.0 × 10 ⁶ cells / dish in a 10 cm diameter cell culture dish and cultured for 24 hours, and then treated with the following compounds.

For HCT116 cells, E7820 (0.8μΜ), Ε7070 (0 · 8μΜ), LY295501 (30 zM), CQS (8Μ), adriamycin (0.2, uM), daunomycin (0.2 ··), ICRF154 (80Μ ), ICRF159 (80 Μ), kenpaullone (10 / ζΜ), alsterpuHone (10 Μ), trichostatin A (θ.Ι, αΜ), rapamycin (80 / M) were evaluated. On the other hand, E7070 (0.8 M) was evaluated for MOLT-4 cells. Where adriamycin and daunomycin are DNA tooisomerase II inhibitors that interfer with DNA, ICRF154 and ICRF159 are catalytic types of DNA topoisomerase II P and harmful agents, kenpaullone and alsterpullone are cyclin- de endent kinases (CDKs inhibitor, trichostatin A is a histone deacetyiase | fi harmful agent, and rapamycin is a known compound as an mTOR / FRAP inhibitor. The compound treatment concentration depends on HCT116 cells of each compound. Based on the 50% growth inhibition concentration (based on the 3-day cell growth inhibitory activity using WST-8), the concentration is set to 3 to 5 times the concentration, and the concentration shown in parentheses following the above compound name is set. Cells were collected after treatment for 24 hours, and cells cultured for 24 hours without addition of compounds were also collected.

Total RNA was extracted from the collected cells using TRIZOL reagent (Invitrogene) according to the attached operation method.

(2) Gene expression analysis by DNA microarray

100 RNA obtained

Was dissolved in diethylpyrocarbonate (DEPC) treated with the sterile water, purified using an additional RNeasy column (QIAGEN), 2-strand using Superscript Choice System (Invitrogen) and T7-d (T) ₂ 4 primers CDNA was synthesized.

First 10 μg RNA, 5 μΜ T7-d (T) 24 primer, lx First strand Buffer, 10 mM DTT, 500 M dNTP mix, and 20 units / μ 1 Superscript II Reverse Transcriptase were added and reacted at 42 ° C for 1 hour to synthesize single-stranded DNA. Next, add lx Second strand buffer, 200 d dNTP mix 67 U / ml DNA ligase, 270 U / ml DNA polymerase I, and 13 U / ml RNase H, and react at 16 ° C for 2 hours 2 Single-stranded cDNA was synthesized. Further, 67 U / ml T4 DNA polymerase I was added and reacted at 16 ° C. for 5 minutes, and then the reaction was stopped by adding ΙΟμΙ 0.5 M EDTA.

The obtained cDNA was purified with phenol / cloform form and labeled with piotinylated UTP and CTP using RNA Transcript Labeling Kit (Enzo Diagnostics) according to the attached procedure. The reaction product was purified with an RNeasy column, and then cRNA was fragmented by heating at 94 ° C for 35 minutes in 200 mM trisacetic acid pH8.1, 150 mM magnesium acetate, 50 mM acetic acid lithium.

Fragmented cHNA was hybridized to GeneChip (Affymetrix) Human Focus array in 100 mM MES, 1 M sodium salt, 20 mM EDTA, 0.01% Tween 20 at 45 ° C. for 16 hours. After hybridization, GeneChip was washed and stained according to the protocol Midi-euk2 attached to the Affymetrix fluidics station. For staining, streptavidin-phycoerythrin and biotinylated anti-streptavidin antibody were used. The stained GeneChip was scanned using an HP argon ion laser confocal microscope (Hewlett Packard), and the fluorescence intensity was measured. For measurement, excitation is performed at a wavelength of 488 nm, and emission is performed at a wavelength of 570 nm.

Do all quantitative data analysis use GeneChip software (Affymetrix) and Gene Spring (Silicongenetics)? 7 pieces. When using GeneChip software to evaluate gene expression changes due to compounds, the RNA quantification value is dissociated more than twice between the two conditions of the compound-treated group and the untreated group. It was judged that gene expression was significantly “increased” or “decreased”. When using Gene Spring to evaluate the similarity of gene expression changes induced by each compound, hierarchical clustering analysis is performed based on the expression changes of all genes on the Human Focus Array. W

went.

The results of hierarchical clustering analysis of HCT116 cells are shown in FIG.

As a result of analysis, adriamycin and daunomycin, ICRF154 and ICRF159, KenpauUone and alsterpullone, which have the same mechanism of action, caused similar genetic changes, respectively (Fig. 3). Therefore, it was confirmed that compounds having the same mechanism of action cause similar genetic changes.

E7070, E7820, LY295501 and CQS caused similar genetic changes (Figure 3). Therefore, this analysis strongly suggests that E7070, E7820, LY295501 and CQS have the same or similar mechanism of action and bring about the same or similar genetic alterations and effects.

[Reference Example 2] DNA microarray analysis

HCT116 cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum, 100 units / ml penicillin, 100 μg / ml streptomycin. Hereinafter, the culture opi compound treatment was carried out in an incubator adjusted to 5% CO ₂ and 37 ° C. 10 cm diameter cell culture dishes at a rate of 2.0 x 10 ⁶ cells / dish

HCT116 cells were seeded and treated with the following compounds after culturing for 24 hours.

In this reference example, using HCT116 cells, E7820 (0 · 16 μΜ), Ε7070 (0 · 26 μΜ),

LY186641 (59 μ Μ), LY295501 (24 μ 、), LY573636 (9.6 Μ) CQS (4.0 μ Μ), MST16 (100 μ Μ), etoposide (3.6 Μ), ethoxzolamide (410 μ Μ), capsaicin

We examined changes in gene expression when treated with 12 compounds (280 Μ), trichostatin A (0.16 μΜ), and kenpauUone (7.1 Μ).

Here, MST16 is a catalytic type of DNA topoisomerase II inhibitor, etoposide (3. DNA topoisomerase II inhibition that forms c 丄 eavable complex lj, ethoxzolamide f or carbonic anhyd ase inhibitory homozygous J, capsaicin ¾ tumor-specinc plasma membrane NADH oxidase inhibitor, trichostatin A ¾ istone deacetylase P, and kenpauUone are known compounds as cyclin-dependent kinases (CDKs) inhibitors. The compound treatment concentration was set as twice the concentration based on the 50% growth inhibitory concentration of each compound on HCT116 cells (based on the 3-day cytostatic activity using MT). Cells were collected after 24 hours of treatment at the set concentration indicated in parentheses following the compound name. In addition, cells cultured for 24 hours without compound addition were also collected.

Gene expression analysis using a DNA microarray was performed in the same manner as “(2) Gene expression analysis using DNA microarray” in Reference Example 1.

In addition, this reference example was duplicated for each sample (for convenience of experiments, each sample was assigned branch numbers in the manner of control-1, control_2, Ε7070 · 1, Ε7070-2). Then, gene expression changes induced by each compound were analyzed using GeneChip (Affymetrix) system (Human Focus array).

Applying the RMA method (robust multi-array average method (Biostatistics (2003), 4 _; 249-264)) to .celj files of 26 samples obtained in this reference example (13 samples x 2 of control + 12 compounds) After the normal distribution at the probe level, the log value of the signal intensity at the gene level was calculated, and then the compound intensity was calculated from the log value of the signal intensity in the compound treatment group of each gene. The log value of the signal intensity in the treatment group (control-l) was subtracted to obtain the log value of the signal ratio of the compound treatment group relative to control-l, and the cosine correlation coefficient was calculated and the correlation coefficient between experiments was calculated. Based on this correlation coefficient, hierarchical clustering analysis was performed by the UPGMA method (Unweighted Pair Group method with Arithmetic mean method) (Fig. 5) The same calculation was performed for control-2. (Fig. 6 and Fig. 7) Soft used Air [3 R 2.0.1 (http://www.r-project.org/)], affy package 1.5.8 (http 7 / www. Bioconauctor.org).

4-7, “LY1” is LY186641, “LY2” is LY295501, “LY5” is LY573636, “CAI” is ethoxzolamide, “Cap” is cansaicin “MST” indicates MST16, “Etop” indicates etoposide, “TSA” indicates trichostatin A, and “Kenp” indicates cenpaullone. In Fig. 5 and Fig. 7, "de hclust (*," average ")" is a command used for statistical analysis, indicating that clustering analysis by R was performed using the average value of dupulicate experimental data. Show. As a result of the analysis, E7070, E7820, LY186641, LY295501, LY573636 and CQS showed very high similarity in gene changes caused by HCT116 cells, and other compounds (MST16, etoposide, ethoxzolamide, capsaicin, trichostatin A Kenpaullone) profile (Fig. 4 to Fig. 7). Therefore, this analysis strongly suggests that E7070, E7820, LY186641, LY295501, LY573636, and CQS have the same or similar mechanism of action and bring about the same or similar genetic changes and effects.

[Reference Example 3] Cancer cell line panel experiment

Using a panel of 36 human cancer cells, we investigated the correlation between the cytostatic activity of E7820, E7070, CQS, LY186641, and LY295501. The cancer cell lines used were: DLD-1, HCT15, HCT116, HT29, SW480, SW620, WiDr (above human colon cancer cell line), A427, A549, LX 1, NCI-H460, NCI-H522, PC- 9, PC-10 (Human lung cancer cell line), GT3TKB, HGC27, MKN1, MKN7, MKN28, MKN74 (Human gastric cancer cell line), AsPC-l, KP-1, KP-4, MiaPaCall, PANC-1, SUIT-2 (above human knee cancer cell line), BSY-1, HBC5, MCF-7, MDA-MB-231, MDA-MB-435, MDA-MB-468 (above, hi Breast cancer cell lines), CCRF-CEM, HL60, K562, MOLT-4 (and above, human leukemia cell lines), and all cells are 10% fetal bovine serum, 100 units / ml The cells were cultured in RPMI-1640 medium supplemented with nicillin and 100 / g / ml streptomycin at 37 ° C. under 5% CO ₂ conditions (Table 5). Table 5

36 human cancer cell lines tested in 3-day MTT assays

Colon Stomach Breast

DLD-1 (1250 / well, 16.8 h) GT3TKB (2000 / well, 21.1 h) BSY-l (2000 / well, 46.1 h)

HCT15 (1500 / welI, 14.5 h) HGC27 (1500 / wcll, 14.6 h) HBC5 (2000 / well, 31.8 h)

HCT116 (1250 / well, 13.4 h) M N1 (4000 / well, 35.9 h) MCF-7 (3000 / well, 29.5 h)

HT29 (2500 / elI, 19.8 h) MKN7 (3000 / well, 37.4 h) MDA-MB231 (2000 / well, 21.6 h) SW480 (3000 / well, 19.5 h) MKN28 (2000 / well, 22.7 h) MDA-MB -435 (3000 / well, 24.4 h) SW620 (2500 / well, 17.3 h) MKN74 (4000 / well, 24.8 h) MDA-MB-468 (3000 / well, 34.2 h) WiDr (2000 / welI, 18.9 h)

Pancreas leukemia

Lung AsPC-l (2500 /, vell, 28.4 h) CCRF-CEM (1500 / welI, 27.2 h)

A427 (2500 / well, 32.4 h) KP-1 (2000 / well, 24.8 h) HL60 (1500 / well, 29.5 h)

A549 (1250 / well, 18.9 h) KP-4 (2000 / well, 16.7 h) K562 (1500 / well, 20.6 h)

LX-1 (2000 / weIl, 17.2 h) MiaPaCaH (2500 / well, 19.1 h) MOLT-4 (1500 / well, 22.3 h) NCI-H460 (1000 / eIl, 13.6 h) PANC-1 (2500 / well, 27.9 h)

NCI-H522 (4000 / well, 80.4 h) SUIT-2 (2000 / well, 15.6 h)

PC-9 (2000 / well, 23.7 h)

PC-10 (2000 / eU, 24.0 h)

Cell line (initial cell number, doubling time) Table 5 shows the types of human cancer cell lines, the number of cells and the doubling time in the human cancer cell line panel.

The number of cells listed in Table 5 was plated on a 96-well microphone mouthplate (flat bottom) (50 μ 1 / well), and after 3 hours, a 3-fold dilution series of compounds was added (50 ^ 1 / well). 72 hours later, WST-8 (ΙΟμΙ / weU) was added, and the absorbance at 450 nm was measured. The 50% growth inhibitory inhibitory concentration for all 36 cancer cells was determined by the least squares method, and the patterns were compared among the compounds. As a correlation index, we used Pearson's correlation coefficients (Paull, KD et al. Display and analysis of patterns of differential activity of drugs against human tumor cell lines: development of mean graph and COMPARE algorithm.J. Natl. Cancer Inst. 1989, 81, 1088-1092; Monks, A. et al. Feasibility of a high-flux anticancer drug screen using a diverse panel of cultured human tumor cell lines. J. Natl. Cancer Inst. 1991, 83, 757-766 .) As a result, E7070, E7820, LY186641, LY295501 and CQS showed a high correlation coefficient in the growth inhibitory activity against each cancer cell line (Table 6). Therefore, this analysis strongly suggests that E7070, E7820, LY186641, LY295501, and CQS have the same or similar mechanism of action and bring about the same or similar gene change effect.

Table 6

E7070 E7820 CQS LY186641 LY295501

E7070 1.00 0.98 0.97 0.93 0.80

E7820 0.98 1.00 0.96 0.95 0.82

CQS 0.97 0.96 1.00 0.92 0.82

LY186641 0.93 0.95 0.92 1.00 0.81

LY295501 0.80 0.82 0.82 0.81 1.00

Table 6 shows the correlation coefficients between compounds (E7070, E7820, CQS, LY186641 and LY295501) in a human cancer cell line panel.

[Reference Example 4] Cross resistance in E7070 resistant strain

The cell growth inhibitory activity of E7820, LY186641, LY295501, LY-ASAP and CQS was evaluated using E7070 resistant strains. HCT116-C9 is a substrain isolated from human colorectal cancer-derived HCT116 (American Type Culture Collection, Manassas, VA, USA). This HCT116-C9 is cultured in the presence of E7070, and the E7070 concentration is gradually increased. The E7070 resistant substrains obtained by raising are HCT116-C9-C1 and HCT116-C9-C4 (Molecular Cancer Therapeutics, 2002, 1, 275-286).

Three cell lines HCT116-C9, HCT116-C9-C1, and HCT116-C9-C4 were spread on a 96-well microphone plate (flat bottom) at 3000 cells / well (50 μ 1Λνβ! 1), and tripled 24 hours later Dilution series of compounds were added (50 / zl / well). Furthermore, cell proliferation inhibitory activity was evaluated 72 hours later by the MTT method (Mossmann T., J. Immunol. Methods, 1983, 65, 55-63). The 50% growth inhibitory inhibitory concentration for each cancer cell was determined by the least square method.

As a result, the cytostatic activity of E7070 is IC50 against HCT116-C9 (C9). W 200

Was 0 · 127μΜ. In contrast, the activity against HCT116-C9-C1 (C9C1) and HCT116-C9-C4 (C9C4) is IC50 = 31.9 μΜ and 26.9 それぞれ, respectively, and the cell growth inhibitory activity of 97070 against C9C1 and C9C4 is remarkable. It was confirmed that it decreased (Fig. 8). In addition, the cell growth inhibitory activities of E7820, CQS, LY186641, LY295501, and LY-ASAP were HCT116-C9 activity at IC50 = 0,080 / M, 1.73 ^ Μ, 33.6μΜ, 10.9 Μ 1.63Μ, respectively. In contrast, the activity against HCT116-C9-C1 and HCT116-C9-C4 was IC50 = 51.2 / ζΜ, 634Μ, 134 μΜ, 111 // Μ, 113 / Μ for HCT116-C9-C1, respectively. IC50 = 52.8 μΜ, 517 μΜ, 138 μΜ, 110 μΜ, and 90.3 μΜ for HCT116-C9-C4, respectively. Therefore, the cell growth inhibitory activity of Ε7820, CQS, LY186641, LY295501, and LY-ASA was remarkably reduced compared to the activity against C9. The activity against C9C1 and C9C4 was significantly reduced (FIG. 8). Therefore, it was strongly suggested that E7070, E7820, LY186641, LY295501, LY-ASAP and CQS have the same or similar mechanism of action, and bring about the same or similar genetic changes and effects.

[Reference Example 5] Cross resistance in E7070 resistant strain

In the same manner as in Reference Example 4, the cell growth inhibitory activity of LY573636 was evaluated at the same time as E7070 using an E7070 resistant strain.

As a result, the cytostatic activity of E7070 was higher than that of HCT116-C9 (IC50 = 0.127 t M), but that of HCT116-C9-C1 and HCT116-C9-C4 (IC50 = 32.7 M and 28.0 それぞれ, respectively). It was confirmed again that it was significantly reduced (Figure 9). In addition, LY573636 has a cytostatic activity compared to that of HCT116-C9 (IC50 = 5.11 zM), activity against HCT116-C9-C1 and HCT116-C9-C4 (IC50 = 264 μΜ and 240 μΜ, respectively). ) Was significantly reduced (Figure 9). Therefore, LY573636 was considered to have the same or similar mechanism of action as Ε7070, and strongly suggested that LY573636 produces the same or similar genetic changes and effects.

These results (Reference Example 1-5) Force, et al., Ε7070, Ε7820, LY186641, W

LY295501, LY-ASAP, LY573636 or CQS or their combinational powers were found to produce the same or similar genetic changes and the same or similar actions and effects.

Therefore, as with E7070 (Examples 1 and 2), the sulfoneamide compound, preferably E7820, LY186641, LY295501, LY-ASAP, LY573636, or CQS or a combination of these, has an NSE content. It was clarified that it can be predicted by measuring and using the NSE amount as an indicator. Industrial applicability

The present invention provides a method for predicting the antitumor effect of a sulfonamide compound.

More specifically, the antitumor effect of a sulfonamide compound can be predicted by measuring the amount of NSE and using the amount of NSE as an index. In other words, in cancer patients, when the amount of NSE was high, it was possible to judge that cancer patients were highly sensitive to sulfonamide compounds.

The method according to the present invention makes it possible to predict an antitumor effect without administering a sulfonamide compound to a patient, so that a patient who can expect a more antitumor effect can be selected and treated. It has become possible to contribute to the patient's QOL.

Claims

A method for predicting the antitumor effect of a sulfonamide compound comprising:

Measuring NSE amount;

Including the method,

Sulfonamide compounds are

Formula (I)

[Where

A ring is an optionally substituted monocyclic or bicyclic aromatic ring,

Ring B is an optionally substituted 6-membered cyclic unsaturated hydrocarbon or unsaturated 6-membered heterocycle containing one nitrogen atom as a hetero atom,

W is a single bond or one C H = C H—

X is one N (R i) — or oxygen atom,

Y is

— C (R ³ ) — or one N— and Z means one N (R2) one,

Where R i, R 2 and R ³ are independently the same or different and hydrogen It means an atom or a lower alkyl group.

A compound represented by

Formula (II)

[Wherein E is 101, 1 N (CH ₃ )-, 1 CH ₂ —, 1 CH ₂ CH ₂ — or — CH ₂ 0 1, D is 1 CH ₂ — or 1 O— Ria represents a hydrogen atom or a halogen atom, and R ^2a represents a halogen atom or a trifluoromethyl group, respectively. ]

A compound represented by

General formula (III)

[Wherein J represents 1O— or 1 NH—, Rib represents a hydrogen atom, a halogen atom, an optionally substituted Ci 1 C ₆ alkyl group, or an optionally substituted group. Ci 1 C4 alkoxy group, optionally substituted Ci 1 C ₄ alkylthio group — CF ₃ , _OCF ₃ , 1 S CF ₃ , optionally substituted d—C ₄ alkoxy carboxyl Group, etro group, azido group, 1 O (S0 ₂ ) CH ₃ , -N (CH ₃ ) 2, hydroxyl group, phenyl group, substituted furan group, pyridinyl group, chenyl group, furyl group, quinolinyl group Or a triazole group, R ² b is a hydrogen atom, a halogen atom, a cyano group, one CF ₃ , an optionally substituted d_Cs alkyl group, or an optionally substituted Ci—C ₄ alkoxycarbonyl two group, which may have a substituent Ci one C ₄ alkoxy group, which may have a substituent There Hue - the group or may have a substituent group quinolinyl group, R3b is a hydrogen atom or an optionally substituted Ci _C ₄ alkoxy radical, R ^4b represents a hydrogen atom or an optionally substituted d—C ₆ alkyl group (provided that at least one of R ^3b and R ^4b is a hydrogen atom), R ⁵ b represents a hydrogen atom, halogen atoms, one may have a substituent group C ₆ alkyl group, an CF ₃ or nitro, R ^6b represents a hydrogen atom, a halogen atom or a substituent chromatic and have good Ci one C ₆ alkyl, A group (provided that when R ^6b is a Ci—C ₆ alkyl group which may have a substituent, R ^5b is a hydrogen atom and R ^7b is a halogen atom), R ^7b is a halogen atom D—C ₆ alkyl group or _CF ₃ (provided that either R ⁵ b or R? B may have a substituent) C 1 C ₆ alkyl group optionally having substituent (s) in a force or if it has a R ^7b force halo gen atom or a substituent is also optionally Ci one C ₆ alkyl group , One of R ^5b and R ^6b are hydrogen atoms) mean respectively. ]

A compound represented by

Formula (IV)

A compound represented by

Formula (V)

The above method, which is at least one compound selected from the group consisting of the compounds represented by: or a pharmacologically acceptable salt thereof, or a solvate thereof.

2. The sulfonamide compound is

N- (3—Chloro 1 H—Indanol 7—Inol) 1 1, 4 1 Benzene Disinole Honamid,

N— (3-cyanone 4-methyl- 1 H-indole one 7 f) 1 3-ciano benzenesulfonamide,

N — [[((4) cyclophenyl) amino] carbonyl] 1,2,3-dihydrone 1 H—indene 1-5-sulfonamide,

N — [[((3,4-dicyclophenylamine) amino) force ruponyl] 1,2,3-dihydrobenzofuran-5-sulfonamide,

N— (2, 4-dichlorobenzoinole), 4-one-mouthed phenylolene amide, N— (2, 4-dichlorobenzoinole), 1-5-bromothiophene, 2-snorethone amide

and

2-Sunorefinolide MIDO—Black mouth quinoxaline

The method according to claim 1, which is at least one compound selected from the group consisting of: or a pharmacologically acceptable salt thereof, or a solvate thereof.

3. The sulfonamide compound is N- (3-chloro-1H-indole-17-yl) 1,1,4-benzenedisulfonamide, or a pharmacologically acceptable salt thereof, or a solvate thereof. The method of claim 1, wherein

4. The sulfonamide compound is N— (3-Cyanol 4-methyl-1 H—Indolulu 7 f) _3-Cyananobenzenesulfonamide, or a pharmacologically acceptable salt thereof, or a solvent thereof. The method according to claim 1, which is a Japanese product.

5. Sulfonamide compounds are N — [[((3,4-Diphenyl) phenyl) amino] carbonyl] 1,2,3-Dihydrobenzofuran 1 5-sulfonamide and N 1 (2,4-Dichlorobenzobenzore) 2. The method according to claim 1, which is at least one compound selected from the group consisting of 1-bromothiophene 2-snorephoneamide, or a pharmaceutically acceptable salt thereof, or a solvate thereof. .

6. The method according to claim 1, wherein the sulfonamide compound is a sodium salt of N- (2,4-diglycol benzoyl) -5-promothiophene-2-sulfonamide.

7. The method according to claim 1, wherein the amount of NSE is the amount of NSE in serum.

8. The method according to claim 1, wherein the amount of NSE is the amount of NSE in the tumor.

9. The method according to claim 1, wherein the NSE amount is measured by an immunochemical method.

10. A kit for use in the method according to any one of claims 1-9, comprising:

The kit comprising an anti-NSE antibody.