JPH07188181A - Inhibitor of glutaminase - Google Patents

Abstract

PURPOSE:To obtain an inhibitor, containing a new imidazole compound capable of suppressing the proliferation of malignant tumor in the intestinum crassum, malignant tumor, originating from the intestinum crassum and remotely metastasized to the liver, etc., and malignant tumor in the stomach, prostate gland, the liver, etc., without manifesting the toxicity to normal tissue. CONSTITUTION:This inhibitor contains 0.1-100wt.% (preferably 5-50wt.%) compound (salt) expressed by the formula (A is a lower alkyl which may have a substituent group or amino which may be protected; dotted line underlined is single or double bond; B is H or a chain hydrocarbon which may have a substituent group) (preferred example: 2-amino-1-methylimidazole, 2- benzyloxycarbonylamino-1-methylimidazole or a salt thereof).

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a glutaminase inhibitor containing an imidazole compound as an active ingredient. More specifically, malignant tumors mainly occurring in the large intestine by inhibiting the enzyme glutaminase (EC3.5.1.2) present in the mitochondrial membrane of tumor cells and malignant tumors derived from large intestine distantly metastasized to organs such as liver, Furthermore, it relates to a pharmaceutical composition for suppressing the growth of malignant tumors occurring in the stomach, prostate, liver and the like. Furthermore, the present invention relates to novel imidazole compounds that can be used in these compositions.

[0002]

2. Description of the Related Art It is an organ responsible for cell energy production as one of enzymes that are characteristically required for the growth of certain tumor cells, but are utilized only in trace amounts by other normal cells. Glutaminase, an enzyme present in the membrane of mitochondria, is known, but inhibition of this enzyme selectively suppresses the growth of tumor cells and does not cause damage to other cells. No successful development of any anti-cancer drug [eg Medina et al., Miguel Angel Medin
a): Molecular and Cellular Biochemistry, 1
13, 1-15 (1992)]. Looking at the types of cancer, colon cancer has the largest number of patients next to lung cancer, and the recurrence rate after surgery is also high. At present, there is no effective single-agent adjuvant chemotherapy after surgery, and treatment with a combination of existing chemotherapeutic agents centered on the antimetabolite 5-fluorouracil (5-FU) has been attempted. , An effective standard therapy has not been established yet (eg, “colorectal cancer”, Medical View, 1989). Although development of new selective therapeutic agents for colorectal cancer has been attempted all over the world, clinically effective ones have not yet been developed (for example, "Colon Cancer Cell
s) "Page 1: Moyer and Post Editing, Academic Press (MP Moyer and GH Poste ed., A.
academic Press, Inc. ) 1990). The following compounds are also known. (1) Cancer Research (Cancer Reserch, 53 , 3)
172-3178, July 1, 1993) (A):

[0003]

[Chemical 5]

(2) Japanese Patent Laid-Open Nos. 62-7732 and 3-
17064, Biochemical Pharmacology (Bioc
hemical Pharmacology, Vol. 43, No. 8, pp1
717-1723, 1992)
(B):

[0005]

[Chemical 6]

(3) Compound (C) described in JP-A-3-5461:

[0007]

[Chemical 7]

(4) Compound (D) described in FR-2681323-A1:

[0009]

[Chemical 8]

(5) Biochemical Society Transactio
ns, 14 , 1053-1054, 1986) (E):

[0011]

[Chemical 9]

The compounds (A), (B), (C) and (D)
Has a different chemical structure and mechanism of action from the present compound.
Further, there is no description that the compound (E) inhibits the enzyme glutaminase and selectively suppresses the growth of tumor cells.

[0013]

Under such circumstances, there is provided a highly selective therapeutic agent having an excellent tumor cell growth inhibitory action and showing no toxicity to normal in vivo tissues. Is required.

[0014]

[Means for Solving the Problems] In order to solve the above problems, the present inventors have conducted a search for a compound that selectively inhibits cell growth of a human colon cancer cell line, and as a result of diligent studies, It was found that the imidazole derivative of (1) specifically inhibits the activity of the glutaminase, a mitochondrial enzyme of human colon cancer cells, and exhibits a colon cancer cell growth inhibitory action with excellent selectivity. The present invention has been completed based on these.

That is, the present invention provides (1) the general formula (I):

[Chemical 10] B represents a hydrogen atom or a chain hydrocarbon group which may have a substituent], or a pharmaceutical composition for inhibiting glutaminase, comprising a compound represented by the formula or a salt thereof.

(2) A is an optionally protected amino group. The pharmaceutical composition according to (1) above, (3) A is an amino optionally protected by a group capable of being eliminated by in vivo metabolism. The pharmaceutical composition according to (1) above, which is a group, (4) A is an amino group which may be protected by an acyl group, and (5) A is an amino group. The pharmaceutical composition according to (1) above, (6) B, which may have a substituent, may be a linear or branched C _1-6 alkyl group, linear or branched. The pharmaceutical composition according to (1) above, which is a C _2-6 alkenyl group or a linear or branched C _2-6 alkynyl group, (7) B is a linear or branched C _1-6 The pharmaceutical composition according to the above (1), which is an alkyl group,

(8)

[Chemical 11] The pharmaceutical composition according to (1) above, wherein

(9) The pharmaceutical composition according to (1) above, which is a mitochondrial glutaminase inhibitor, (10) The pharmaceutical composition according to (1) above, which is a therapeutic agent for glutamine-requiring malignant tumors, (11) colorectal cancer. Alternatively, the pharmaceutical composition according to (1) above, which is a therapeutic agent for colorectal cancer-derived malignant tumor, (12) the pharmaceutical composition according to (1) above, which is a therapeutic agent for colon cancer.
3) The pharmaceutical composition according to the above (1), which is a therapeutic agent for rectal cancer,
(14) The compound represented by the general formula (I) is 2-amino-
The pharmaceutical composition according to (1) above, which is 1-methylimidazole, (15) The pharmaceutical according to (1) above, wherein the compound represented by the general formula (I) is 2-benzyloxycarbonylamino-1-methylimidazole. Composition,

(16) General formula (I):

[Chemical 12] B represents a hydrogen atom or a chain hydrocarbon group which may have a substituent], or a glutaminase inhibitor comprising a salt thereof, (17) the general formula (II):

[0020]

[Chemical 13]

[In the formula, A ′ represents an amino group which may be protected by an acyl group, and B ′ represents a chain hydrocarbon group which may have a substituent. However, when B'is methyl or ethyl, A'represents an amino group protected by an acyl group. ] The compound or its salt represented by these, (18) A'is the compound of the said (17) whose amino group protected by the acyl group, (19) A'is -NHCOOR (In formula, R is a lower alkyl. A group or a benzyl group), and B ′ is a lower alkyl group, (20)
B ′ is a linear or branched C _1-6 alkyl group, a linear or branched C _2-6 alkenyl group, a linear or branched group, each of which may have a substituent. The compound according to the above (17), which is a branched C _2-6 alkynyl group, (21)
The compound according to (17) above, wherein B ′ is a linear or branched C _1-6 alkyl group, (22) the compound according to (17) above, which is 2-amino-1-methylimidazole, and ( 23) 2-Benzyloxycarbonylamino-1-
The present invention relates to the compound described in (17) above, which is methylimidazole.

In the present specification, A represents a lower alkyl group which may have a substituent or an amino group which may be protected. As the lower alkyl group represented by A,
For example, carbon number 1 such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, hexyl.
A straight-chain or branched alkyl group having 1 to 6 is used, and a straight-chain or branched alkyl group having 1 to 3 carbon atoms such as methyl and ethyl is generally used.
The lower alkyl group is hydroxy, amino, carboxyl, nitro, mono- or di-C _1-6 alkylamino (eg, methylamino, ethylamino, propylamino,
Dimethylamino, diethylamino etc.), C _1-6 alkoxy (eg methoxy, ethoxy, propoxy, hexyloxy etc.), C _1-6 alkyl-carbonyloxy (eg acetoxy, ethylcarbonyloxy etc.) and halogen (eg fluorine , Chlorine, bromine, iodine, etc.)
It may have 1 to 3 substituents selected from

The optionally protected amino group represented by A includes a protected amino group in addition to the amino group. The protected group of the protected amino group is a group which can be eliminated by metabolism in the living body, and specific examples thereof include an acyl group and an alkylidene group which may have a substituent. The case where the protecting group is an alkylidene group which may have a substituent means a case where the amino group in the amino group which may be protected forms a Schiff base. As the acyl group as the protecting group, for example, a linear or branched C _1-6 alkanoyl group which may contain an unsaturated bond (eg formyl, acetyl,
Propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl, hexanoyl, acryloyl, methacryloyl, crotonoyl, isocrotonoyl, propioroyl, etc.), C _6-10 aroyl group (for example,
Benzoyl, 1-naphthoyl, 2-naphthoyl, etc.),
C _1-3 alkylsulfonyl group (eg, methanesulfonyl, ethanesulfonyl, etc.), C _6-10 arylsulfonyl group (eg, benzenesulfonyl, p-toluenesulfonyl, 1-naphthalenesulfonyl, 2-naphthalenesulfonyl, etc.), —COOR (In the formula, R has the same meaning as described above), -CONR ¹ R ² (in the formula, R ¹ and R ² are the same or different and each represents a hydrogen atom, a lower alkyl group or an aryl group), and a protective group. (For example, an amino group may be protected with a lower alkoxycarbonyl group such as t-butoxycarbonyl, or a carboxyl group that does not form an acyl group may be protected with a lower alkyl group such as methyl). Examples of alanine, phenylalanine, aspartic acid, glutamic acid, glutamine, cysteine Lysine, such as valine, and the like) and the like. Examples of the lower alkyl group represented by R, R ¹ or R ² include the same as the lower alkyl group represented by A. Examples of the aryl group represented by R ¹ and R ² include a C _6-10 aryl group (eg, phenyl, 1-naphthyl, 2-naphthyl, etc.).

The alkylidene group which may have a substituent as the protecting group includes, for example, ethylidene, isopropylidene, benzylidene, p-nitrobenzylidene,
Examples thereof include salicylidene and dimethylaminomethylene. When the alkylidene group has a substituent, the substituent is, for example, a C _1-6 alkylthio group (eg,
Methylthio, ethylthio, propylthio, etc.), mono-
Or a di-C _1-6 alkylamino group (eg, methylamino, ethylamino, dimethylamino, diethylamino, etc.), a C _1-6 alkoxy group (eg, methoxy, ethoxy,
Propoxy, etc.), halogen (eg fluoro, chloro,
Bromo etc.), hydroxyl group, nitro group and the like. The protecting group is preferably an acyl group, more preferably -COOR (the symbols in the formula have the same meanings as defined above, and a benzyl group is commonly used). In addition, these protecting groups can be prepared by a method known per se [T W Green and PG Wuts, “Protective Groups in -Organic Synthesis (ProtectiveGroups in Organic Synthesis) "
Second Edition, John Wiley & Sons Incorporated (John Wiley & Sons, In
c. ), New York (1991)].

[0025]

[Chemical 14]

B represents a hydrogen atom or a chain hydrocarbon group which may have a substituent. Examples of the chain hydrocarbon group represented by B include a lower alkyl group, a lower alkenyl group and a lower alkynyl group, and a lower alkyl group is commonly used. As the lower alkyl group, the same groups as the lower alkyl groups described above as A are used, and a methyl group is generally used. Examples of the lower alkenyl group include:
Vinyl, allyl, isopropenyl, 2-methallyl, 3-
A straight-chain or branched alkenyl group having 2 to 6 carbon atoms such as methallyl and 3-butenyl is used, and an allyl group is particularly widely used. As the lower alkynyl group, a linear or branched alkynyl group having 2 to 6 carbon atoms such as ethynyl, 2-propynyl and 3-hexynyl is used, and the 2-propynyl group is particularly widely used. Each of the chain hydrocarbon groups represented by B may have 1 to 3 substituents similar to the substituents of the lower alkyl group represented by A, for example. Preferable examples of A include an amino group or an optionally protected amino group such as a benzyloxycarbonylamino group, and the amino group is particularly widely used. Preferable examples of B include C _1-3 alkyl groups such as methyl group. Further, as the compound represented by the general formula (I), a compound represented by the general formula (II)

[0027]

[Chemical 15]

[In the formula, A ′ represents an amino group which may be protected by an acyl group, and B ′ represents a chain hydrocarbon group which may have a substituent. However, when B'is methyl or ethyl, A'represents an amino group protected by an acyl group. ] The compound represented by or a salt thereof is also widely used. Examples of the acyl group of the protective group for A ′ include the same as the acyl group of the protective group for A. Examples of the chain hydrocarbon group which may have a substituent represented by B ′ include the same ones as the chain hydrocarbon group which may have a substituent represented by B. A ′ is preferably an amino group protected by an acyl group, particularly —NHCOOR (in the formula, R represents a lower alkyl group or a benzyl group similar to those described in A above), and a benzyloxycarbonylamino group is particularly preferable. B'is preferably a lower alkyl group similar to those exemplified for A, and particularly preferably a methyl group. General formula (I)
Alternatively, the compound represented by (II) is preferably, for example, 2
-Amino-1-methylimidazole or 2-benzyloxycarbonylamino-1-methylimidazole, or a salt thereof.

The compound represented by the general formula (I) or (II) can be reacted with an acid to form a salt regardless of the kind of the substituent represented by A, B, A'or B'in the general formula. May be formed. As the acid, an organic acid (for example, acetic acid,
Ethyl succinic acid, ethyl carbonic acid, glycopeptonic acid, stearic acid, propionic acid, lactobionic acid, oxalic acid,
Maleic acid, fumaric acid, succinic acid, lactic acid, trifluoroacetic acid, methanesulfonic acid, p-toluenesulfonic acid,
Benzenesulfonic acid, etc.) or mineral acids (for example, hydrochloric acid,
Sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid, etc.) are used,
Preferably, acetic acid, stearic acid, oxalic acid, succinic acid, lactic acid, hydrochloric acid, sulfuric acid, nitric acid and the like, more preferably,
Acetic acid or hydrochloric acid is commonly used.

In the general formula (I),

[Chemical 16] It may be converted into a tautomer, and when a tautomer is produced, it may be present as a mixture thereof.

Compound (I) or a salt thereof can be produced by a method known per se or a method analogous thereto. For example, the Journal of Chemical Society
(J. Chem. Soc.) P. 307 (1956), Kim Geterotsikl. Soedin.
It can be produced by the method described on page 180 (1969) or a method analogous thereto. For example, 1-alkyl-2-
The aminoimidazole derivative (IV) is used in the presence of an acid catalyst,
It can be produced by a cyclization reaction of a guanidine derivative (III). 1-alkyl imidazole derivative (VI)
Can be produced by alkylation of compound (V). When A ′ is a protected amino group, compound (IV) can be produced by deprotecting compound (VI) by a conventional method, and conversely, compound (IV) can be protected by a conventional method. Compound (VI) can also be produced by introducing a group.

[0032]

[Chemical 17]

[0033]

[Chemical 18] The imidazoline derivative represented by the general formula (I) or a salt thereof can be produced by a method known per se or a method analogous thereto. For example, Journal of Organic Chemistry (J. Org. Chem., 24 , 1
157 (1959)) or by subjecting the intermediates or final products in the production of the imidazole derivatives (IV) to (VI) to catalytic reduction reaction at an appropriate stage by a conventional method. Obtainable.

The compound of formula (I) or a salt thereof has an excellent selective growth inhibitory action on tumor cells,
It can be safely used for mammals (eg, humans, mice, etc.), and is particularly useful as a therapeutic agent for malignant tumors that have developed in the large intestine and malignant tumors derived from the large intestine that have metastasized to organs such as the liver.

The compound (I) or a salt thereof may be in the form of bulk powder, but is usually a carrier for pharmaceutical preparations such as excipients (eg calcium carbonate, kaolin, sodium hydrogen carbonate, lactose,
Starch, crystalline cellulose, talc, granulated sugar, porous substance, etc.), binder (for example, dextrin, gums, pregelatinized starch, gelatin, hydroxypropylcellulose, hydroxypropylmethylcellulose, pullulan, etc.), disintegrant (for example, carboxymethylcellulose) Calcium, croscarmellose sodium, crospovidone, low-substituted hydroxypropylcellulose, partially pregelatinized starch, etc.), lubricants (e.g. magnesium stearate, calcium stearate, talc, starch, sodium benzoate, etc.), colorants (e.g. Tar dyes, caramel, iron sesquioxide, titanium oxide, riboflavins, etc.), corrigents (eg sweeteners, flavors etc.), stabilizers (eg sodium sulfite etc.) and preservatives (eg parabens, sorbic acid etc.) During ~ Et appropriately using an appropriate amount,
It is administered in a form prepared according to a conventional method.

The therapeutic agent of the present invention containing the above-mentioned preparation contains a compound (I) or a salt thereof in an amount effective for treating. The content of the compound (I) or a salt thereof in the preparation of the present invention is usually 0.1 to 100% by weight, preferably 5 to 50% by weight of the whole preparation. Specific examples of the dosage form include tablets (including sugar-coated tablets and film-coated tablets), pills, capsules, granules, fine granules, powders, syrups, emulsions, suspensions, injections, etc. . These preparations are prepared according to a conventional method (for example, the method described in the Japanese Pharmacopoeia).

Specifically, the method for producing tablets is as follows: a pharmaceutical product as it is, an excipient, a binder, a disintegrating agent or other appropriate additives are uniformly mixed and granulated by a suitable method. After that, add a lubricant etc. and press-mold, or directly press-mold the drug as it is, or mix it evenly with excipients, binders, disintegrants or other appropriate additives. It is also possible to manufacture by granulation, or by premixing the granules as they are, or by adding appropriate additives and mixing them evenly, and then compression molding. In addition, a colorant, a flavoring agent and the like can be added to the present agent, if necessary. Furthermore, this agent can be coated with a suitable coating agent. The method for producing injections is to make a fixed amount of a fixed amount of a drug by dissolving, suspending or emulsifying it in water for injection in the case of an aqueous solvent, physiological saline, Ringer's solution, etc., and usually in a vegetable oil in the case of a non-aqueous solvent. Alternatively, it can be manufactured by taking a certain amount of a drug and sealing it in a container for injection. As the oral pharmaceutical carrier, substances commonly used in the pharmaceutical field such as starch, mannitol, crystalline cellulose, sodium carboxymethyl cellulose and the like can be used. As the carrier for injection, for example, distilled water, physiological saline, glucose solution, infusion solution and the like are used. In addition, additives generally used in preparations can be added as appropriate.

The preparation of the present invention selectively suppresses the growth of tumor cells, and its action is due to inhibition of the mitochondrial membrane enzyme glutaminase of tumor cells. Therefore, the preparation of the present invention is useful as a therapeutic agent for a malignant tumor in which the requirement for glutamine is involved in its growth. That is, treatment of glutamine-requiring malignant tumors in the mammal, such as colon cancer (including colon cancer and rectal cancer), gastric cancer, prostate cancer, liver cancer, and metastatic lesions in which they metastasize to organs such as lung and liver. , Or their prevention of distant metastases.

The dose of the preparation of the present invention varies depending on age, body weight, symptoms, administration route, number of administrations, etc. For example, in the case of colon cancer, the compound (I) is administered to an adult weighing 60 kg.
Or as its salt, usually 0.1 to 100m per day
It is advisable to administer divided doses of g / kg, preferably 1 to 50 mg / kg, more preferably 1 to 10 mg / kg. The route of administration may be oral or parenteral.

[0040]

EXAMPLES The present invention will be described in more detail with reference to test examples, reference examples and examples, but the present invention is not limited thereto. Test Example 1 In Vitro Inhibition of Cell Proliferation 50 μl (2,000 cells) of a cell suspension of human colon cancer cells WiDr or human normal fetal lung fibroblasts HEL was seeded on a 96-well microplate and 5%. The cells were cultured at 37 ° C in a carbon dioxide gas incubator. On the next day, 50 μl of a 2-fold serially diluted compound solution was added, and the cells were cultured for 3 days. After removing the culture medium containing the compound and washing the cells, the number of surviving cells was quantified by adding an MTT dye solution (Tada et al .: Journal of Immunological Methods.
s), 93, 157-165 (1986)). The compound concentration IC required to suppress the viable cell amount to 50% of the control by obtaining the ratio of the cell amount of each treatment group when the cell amount of the control group to which the compound solution was not added was used as a reference (100%). ₅₀ values were calculated. The results are as shown in Table 1. None of the tested compounds suppressed the cell growth of normal cell HEL, while it suppressed the cell growth of colon cancer WiDr at low concentrations.

[0041]

[Table 1]

Test Example 2 In vitro Inhibitory Effect on Selective Growth of Colorectal Cancer 50 μl (2.0) of cell suspension of various human cancer cells shown in Table 2
00 cells) were seeded on a 96-well microplate and cultured at 37 ° C. in a 5% carbon dioxide gas incubator. The next day, 50 μl of a 2-amino-1-methylimidazole hydrochloride compound solution, which had been 2-fold serially diluted, was added, and the cells were cultured for 3 days.
After removing the culture medium containing the compound and washing the cells, MTT
A dye solution was added to quantify the number of viable cells (Tada et al .: Journal of Immunological Methods (J. Immunol. M.
ethods), 93, 157-165 (1986)). The ratio of the cell amount of each treatment group was calculated when the cell amount of the control group to which the compound solution was not added was 100%, and the viable cell amount was 5
The compound concentration IC ₅₀ value required to suppress to 0% was calculated. The results are as shown in Table 2. From these results, it was shown that the compound 2-amino-1-methylimidazole hydrochloride of the present invention selectively suppresses the growth of colon cancer cell lines.

[0043]

[Table 2]

Test Example 3 In Vivo (In Vivo) Suppression of Colorectal Cancer 5 million human colorectal cancer cells WiDr were subcutaneously transplanted into the chest of Balb / C female nude mice (7 weeks old), and 5 days after the transplantation. Five mice per group were used for the experiment, in which the tumor cell size was measured and the tumor size was made uniform. Compounds of the invention 2-amino-
Physiological saline solution of 1-methylimidazole hydrochloride was orally administered at a concentration of 50 mg / kg, 100 mg / kg and 200 mg / kg, respectively, three times a day for 3 days. The tumor cell diameter was measured the day after the administration, and the formula was: tumor volume = major axis x minor axis x
The tumor volume was calculated from the minor axis x (1/2), and the ratio with the control group administered with physiological saline was determined as the growth rate. The results are shown in Table 3. The compound 2-amino-1-methylimidazole hydrochloride of the present invention suppressed the growth of human colon cancer WiDr cell line transplanted in nude mice in a concentration-dependent manner.

[0045]

[Table 3]

Test Example 4 Action of Compound Having 2-amino Protecting Group 5 million human colon cancer cells WiDr were subcutaneously transplanted into the chest of Balb / C female nude mice (7 weeks old), and 5 days after the transplantation. Five mice per group were used for the experiment, in which the tumor cell diameter was measured and the tumor size was made uniform. The compound of the present invention, 2-benzyloxycarbonylamino-1-methylimidazole hydrochloride, was added to a physiological saline solution at a concentration of 80 mg / kg to give 3 times a day.
Oral administration was performed once for 3 days. The tumor cell diameter was measured on the day after the end of administration, and the formula was: tumor volume = major axis × minor axis × minor axis × (1 /
The tumor volume was calculated according to 2), and the ratio with the control group administered with physiological saline was determined as the growth rate. The results are shown in Table 4.
The compound of the present invention 2-benzyloxycarbonylamino-
1-Methylimidazole hydrochloride suppressed the growth of human colon cancer WiDr cell line transplanted in nude mice.

[0047]

[Table 4]

Test Example 5 Inhibitory Action of Mitochondrial Membrane Enzyme Glutaminase 1) Preparation of Crude Enzyme Preparation Human colon cancer cells WiDr (wet weight 2 g) in 20 ml of buffer A
(250 mM D-mannitol, 70 mM sucrose, 10 m
The supernatant obtained by crushing in M Hepes (pH 7.4) and centrifugation at 600 xg for 10 minutes was further centrifuged at 8,000 xg for 10 minutes. After washing the obtained precipitate with buffer A,
The mitochondrial fraction was suspended in 5,000 μl of buffer A and used as a crude enzyme preparation. 2) Method for measuring enzyme activity Glutaminase is an enzyme that produces glutamic acid using glutamine as a substrate in a phosphate-dependent manner. Therefore, the enzyme activity was measured by producing radiolabeled glutamic acid using radiolabeled glutamine as a substrate, converting both into derivatives, and then separating and quantifying by high performance liquid chromatography. That is, 50 μl of the crude enzyme preparation was added to Buffer B (400
mM Hepes (pH 8.0), 6 μg / ml antimycin A,
Reaction with tritium-labeled glutamine (0.8 μM, Amersham) in the presence of phosphoric acid (200 mM) in 0.4 mg / ml oligomycin) for 15 minutes at 37 ° C.
The reaction was stopped by adding 00 μl of 6% perchloric acid (Wako Pure Chemical Industries, Ltd.). The precipitated protein is precipitated by centrifugation and the supernatant 1
100 μl with 40 μl 1M sodium hydroxide in 00 μl
Derivatization reagent (108 mg of o-phthalaldehyde (Wako Pure Chemical Industries, Ltd.) dissolved in 1 ml of methanol, and 400 μl of 2-mercaptoethanol (Wako Pure Chemical Industries, Ltd.) and 9 ml of sodium borate (pH 10.5) were added. ) Was added to make glutamine and glutamic acid into derivatives. Add them to buffer C (2
0 mM sodium acetate (pH 4.56): acetonitrile = 9
Octadecyl silica (YMC-
(Pack ODS-A, YMC) was used for separation and quantification by reversed-phase high-performance liquid radiochromatography. The enzyme activity was calculated by multiplying the ratio of the amount of glutamic acid to the sum of the amounts of glutamine and glutamic acid by the amount of glutamine added as a substrate. When the enzyme activity of the control group to which no compound was added during the reaction was set to 100%, the enzyme activity in the presence of each compound was determined, and the compound concentration IC ₅₀ value required to suppress the enzyme activity to 50% of the control was calculated. It was calculated. The results were as shown in Tables 5 and 6. All the tested compounds inhibited the mitochondrial membrane glutaminase activity of colon cancer cell WiDr. Also, the order of strength is
This was in agreement with the order of strength of cell growth inhibitory action shown in Table 1.

[0049]

[Table 5]

[Table 6]

Table 1, Table 2, Table 3, Table 4, Table 5 and Table 6
From this, it can be seen that the compound of the present invention exhibits an excellent enzyme glutaminase inhibitory action and selectively suppresses the growth of tumor cells.

Reference Example 1 Preparation of 2- (dimethylaminomethylene) aminoimidazole 2-Aminoimidazole sulfate (5.0 g) was added to water (100 m).
It was dissolved in l), sodium carbonate (6.0 g) was added, and the mixture was stirred for 30 minutes. After the solvent was distilled off under reduced pressure, ethanol was added to the residue and the insoluble matter was filtered off. The filtrate was concentrated under reduced pressure, and the residue was dissolved in dimethylformamide (DMF: 20 ml), and dimethylformamide dimethylacetal (9.0 g) was added.
Was added and stirred overnight. After the solvent was distilled off under reduced pressure, the residue was dissolved in ethyl acetate and washed with saturated saline. After drying over anhydrous magnesium sulfate, the solvent was distilled off under reduced pressure, and the obtained crude crystals were washed with cold ethyl acetate to give 2
-(Dimethylaminomethylene) aminoimidazole (3.3
8 g, yield: 64%) was obtained. Mp: 146-147 ° C. Elemental analysis: C ₆ H ₁₀ N ₄ Calculated: C: 52.16%, H: 7.29%, N: 40.55
% Measured value: C: 51.97%, H: 7.08%, N: 40.41
% IR (KBr) cm ⁻¹ : 1625,1560,1375 ¹ H-NMR (CDCl ₃ ) δ: 3.01 (3H, s), 3.07 (3
H, s), 6.76 (2H, s), 8.48 (1H, s).

Reference Example 2 Preparation of 2- (dimethylaminomethylene) amino-1- (2-propenyl) imidazole 2- (dimethylaminomethylene) aminoimidazole (0.
2 g) in DMF (3 ml) under ice cooling potassium t-butoxide (0.37 g) and 3-bromo-1-propene (0.3 g).
3 ml) was added, the temperature was raised to room temperature (about 15 to 25 ° C.), and the mixture was stirred for 30 minutes. Water was added to the reaction solution, and the product was extracted with chloroform. The extract was dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The obtained residue was subjected to silica gel column chromatography (developing solvent: chloroform-methanol-ammonia water = 40:
2- (dimethylaminomethylene) after purification with 1: 0.1)
Amino-1- (2-propenyl) imidazole (0.31
g, yield: 80%) as a brown oil. ¹ H-NMR (CDCl ₃ ) δ: 3.01 (3H, s), 3.05 (3
H, s), 4.52 (2H, dt, J = 1.5Hz, 5.5Hz), 5.1
1 (1H, dq, J = 1.5Hz, 17Hz), 5.17 (1H, dq,
J = 1.5Hz, 10Hz), 5.93 (1H, ddt, J = 5.5H
z, 10Hz, 17Hz), 6.65 (1H, d, J = 1.5Hz),
6.75 (1H, d, J = 1.5Hz), 8.46 (1H, s).

Reference Example 3 Preparation of 2- (dimethylaminomethylene) amino-1- (2-propynyl) imidazole In the same manner as in Reference Example 2, 2- (dimethylaminomethylene)
Aminoimidazole (0.3g), potassium t-butoxide
2- (Dimethylaminomethylene) amino-1- (2-propynyl) imidazole (0.27 g, yield: 70%) was brown from (0.37 g) and 3-bromo-1-propyne (0.25 ml). Obtained as an oil. ¹ H-NMR (CDCl ₃ ) δ: 2.33 (1H, t, J = 2.5H
z), 3.03 (3H, s), 3.05 (3H, s), 4.71 (2H, d,
J = 2.5Hz), 6.76 (1H, d, J = 2.5Hz), 6.85
(1H, d, J = 2.5Hz), 8.41 (1H, s).

Reference Example 4 3- (2-Dimethylaminomethyleneaminoimidazol-1-yl) -N- (benzyloxycarbonyl) -L
-Production of alanine methyl ester N- (benzyloxycarbonyl) under argon atmosphere
-L-serine methyl ester (1.44g) was dissolved in dichloromethane (28ml) and cooled to -50 ° C. Diisopropylethylamine (0.812) at the same temperature
g) and trifluoromethanesulfonic anhydride (1.77 g) were added, and the mixture was stirred for 1.5 hours. Hexane (28 ml) was added to the reaction solution, and this was added to silica gel (13
g)) flash column chromatography (developing solvent: hexane-dichloromethane = 1: 1; 500 m
It was purified in l). The eluate was concentrated to (10 ml) and diluted with dichloromethane (25 ml), and then 2- (dimethylaminomethylene) aminoimidazole (718 mg) was dissolved in DMF (13 ml) under an argon atmosphere, and then at -20 ° C.
It was added dropwise to the cooled solution. After stirring for 2 hours at the same temperature,
Dichloromethane (300 ml) was added, the mixture was washed with saturated brine and dried over anhydrous magnesium sulfate. The solvent was evaporated under reduced pressure and the obtained residue was purified by silica gel flash column chromatography (developing solvent: 7% ammonia-containing ethanol-chloroform = 1: 49) to give the title compound (975 mg). IR (Neat) cm ⁻¹ : 3230, 3030, 2950,
1750, 1720, 1630, 1520, 1480,
1430, 1400, 1370, 1335, 1280,
1205, 1170, 1100, 1080, 1050,
1025. ¹ H-NMR (CDCl ₃ ) δ: 2.91 (3H, s),
3.01 (3H, s), 3.71 (3H, s), 4.30
(2H, d, J = 4.6Hz), 4.55 (1H, dt,
J = 4.6 Hz, 7.0 Hz), 5.04, 5.12 (2
H, ABq, J = 12.0 Hz), 6.54 (1H, d,
J = 1.6Hz), 6.68 (1H, d, J = 1.6H
z), 7.33 (5H, s), 7.59 (1H, d, J =
7.0 Hz), 8.37 (1 H, s).

Reference Example 5 Preparation of 3- (2-Dimethylaminomethyleneaminoimidazol-1-yl) -L-alanine methyl ester 3- (2-Dimethylaminomethyleneaminoimidazol-1-yl) -N- (benzyloxy) Carbonyl) -L
-Alanine methyl ester (937 mg) was dissolved in methanol (25 ml), 50% water content-10% Pd / C (4
(68 mg) was added, and the mixture was vigorously stirred under a hydrogen atmosphere for 2 hours. The catalyst was filtered off, the solvent was evaporated under reduced pressure, and the obtained residue was purified by silica gel flash column chromatography (developing solvent: 7% ammonia-containing ethanol-chloroform = 3:97) to give the title compound (48
2 mg) was obtained. IR (Neat) cm ⁻¹ : 3370, 3300, 2950,
1730, 1630, 1520, 1480, 1460,
1430, 1395, 1365, 1280, 1250,
1200, 1170, 1100, 1070, 1025. ¹ H-NMR (CDCl ₃ ) δ: 3.02 (3H, s),
3.05 (3H, s), 3.72 (3H, s), 3.81
(1H, dd, J = 5.0Hz, 7.0Hz), 4.08
(1H, dd, J = 7.0Hz, 13.8Hz), 4.2
7 (1H, dd, J = 7.0Hz, 13.8Hz), 6.
64 (1H, d, J = 1.6Hz), 6.73 (1H,
d, J = 1.6 Hz), 8.42 (1H, s).

Reference Example 6 Preparation of N- (2-imidazolyl) benzylideneamine 2-Aminoimidazole sulfate (10.0 g) was dissolved in water (70 ml) and sodium carbonate (12 g: 1.5).
Equivalent weight) was used for neutralization. The solvent was evaporated, ethanol was added, and the insoluble material was filtered off. The solvent in the filtrate was evaporated to give a brown oily substance (6.2 g). Of this, 1.0 g was dissolved in ethanol (5 ml), benzaldehyde (1.28 g) was added, and the mixture was stirred at room temperature for 2 days. The solvent was distilled off, the residue was purified by silica gel column chromatography (developing solvent: ethyl acetate-hexane = 1: 1), and the solvent of the target fraction was distilled off. The precipitated pale yellow prism was collected by filtration using diethyl ether-hexane to give the title compound (0.8
8g: 43.0%) was obtained. Melting point: 121.5-123.0 ° C IR (KBr) cm ^-1 : 3130-2800, 1605,
1575, 1445. ¹ H-NMR (CDCl ₃ ) δ: 7.06 (2H, s),
7.46-7.50 (3H, m), 7.89-7.94 (2
H, m), 9.29 (1H, s).

Reference Example 7 Preparation of N- (1-methylimidazol-2-yl) benzylideneamine N- (2-imidazolyl) benzylideneamine (0.2
g) was dissolved in DMF (4 ml), potassium t-butoxide (0.15 g) and iodomethane (0.18 g) were added, and the mixture was stirred at room temperature for 3 hours. The solvent was distilled off, water was added,
It was extracted with ethyl acetate. The organic layer was washed with water and saturated brine, dried over magnesium sulfate, and the solvent was evaporated.
The residue was purified by silica gel column chromatography (developing solvent: chloroform), and the obtained crude crystals were recrystallized from diethyl ether-hexane to give the title compound (0.1 g: 49.9%) as pale yellow needles. Obtained as crystals. Melting point: 71.0-72.0 ° C Elemental analysis: Calculated as C ₁₁ H ₁₁ N ₃ : C: 71.33%, H: 5.99%, N: 2
2.69%. Found: C: 71.40%, H: 6.02%, N: 2
2.62%. IR (KBr) cm ^-1 : 3080-2900, 1605,
1570, 1510, 1480, 760, 685. ¹ H-NMR (CDCl ₃ ) δ: 3.79 (3H, s),
6.92 (1H, d, J = 1.2Hz), 7.01 (1
H, d, J = 1.2 Hz), 7.45-7.49 (3H,
m), 7.93-7.98 (2H, m), 9.22 (1
H, s).

Reference Example 8 Production of N- (1-methylimidazol-2-yl) benzylideneamine.hydrochloride N- (1-Methylimidazol-2-yl) benzylideneamine (0.1 g) was dissolved in diethyl ether, Four
N hydrochloric acid-ethyl acetate (0.2 ml) was added, and the precipitated pale yellow powder 70 mg (21.9%) was collected by filtration. Melting point: 95.0-98.0 ° C IR (KBr) cm ^-1 : 3120-3050, 1665,
1620, 1585. ¹ H-NMR (CDCl ₃ ) δ: 3.83 (3H, s),
6.89 (1H, s), 6.95 (1H, s), 7.59
-7.73 (3H, m), 8.03-8.07 (2H,
m), 9.27 (1H, s).

Reference Example 9 Preparation of N- (1-methylimidazol-2-yl) -4-methoxybenzylideneamine.hydrochloride 2-amino-1-methylimidazole.hydrochloride (0.3
g) was dissolved in ethanol (4 ml), sodium methoxide (28% methanol solution: 0.5 ml) and p-anisaldehyde (0.34 g) were added, and the mixture was heated under reflux overnight.
The solvent was distilled off, and the residue was subjected to silica gel column chromatography (developing solvent: methanol-chloroform = 5: 9).
After purification in 5), the solvent of the target fraction was distilled off. The precipitated pale yellow prism was collected by filtration using hexane. The obtained crystals were dissolved in chloroform, 4N hydrochloric acid-ethyl acetate (0.22 ml) was added, the solvent was evaporated, and the precipitated pale yellow powder was collected by filtration with diethyl ether to give the title compound (0.17 g : 26.9%) was obtained. Melting point: 191.0-192.0 ° C. Elemental analysis: As C ₁₂ H ₁₄ N ₃ OCl.0.3H ₂ O Calculated value: C: 56.06%, H: 5.72%, N: 1
6.34%. Measured value: C: 56.07%, H: 5.62%, N: 1
6.46%. IR (KBr) cm ^-1 : 2840-2670, 1585,
1255, 1160. ¹ H-NMR (DMSO-d ₆ ) δ: 3.80 (3H,
s), 3.90 (3H, s), 7.18 (2H, d, J =
8.8Hz), 7.57 (1H, d, J = 2.0Hz),
7.62 (1H, d, J = 2.0Hz), 8.02 (2
H, d, J = 8.8 Hz), 9.12 (1 H, s).

Example 1 Preparation of 2-amino-1- (2-propenyl) imidazole hydrochloride 2- (dimethylaminomethylene) amino-1- (2-propenyl) imidazole (0.2 g) was added to 6N hydrochloric acid ( It was dissolved in 5 ml) and heated under reflux for 1 hour. The solvent was evaporated under reduced pressure, and the obtained residue was dissolved in saturated aqueous sodium carbonate solution (1 ml). After concentrating under reduced pressure, the organic matter was extracted with chloroform, and the solvent was distilled off under reduced pressure. The obtained residue was purified by silica gel column chromatography (developing solvent: chloroform-methanol-ammonia water = 20: 1: 0.1) to give 2-amino-1- (2-propenyl) imidazole. This was dissolved in dichloromethane, 4N hydrochloric acid-ethyl acetate (2 ml) was added to give a hydrochloride, and recrystallized from acetonitrile to give 2-amino-1- (2-propenyl) imidazole hydrochloride (0.19 g, Yield: 71%) was obtained as pale yellow needle crystals. Melting point: 98-99 ° C ¹ H-NMR (DMSO-d ₆ ) δ: 4.54 (2H, dt, J =
1.5Hz, 5.5Hz), 5.10 (1H, dq, J = 1.5Hz, 1
7Hz), 5.26 (1H, dq, J = 1.5Hz, 10Hz), 5.9
1 (1H, ddt, J = 5.5Hz, 10Hz, 17Hz), 6.94
(2H, s), 7.76 (2H, brs), 12.20 (1H, brs).

Example 2 Preparation of 2-amino-1- (2-propynyl) imidazole-hydrochloride 2- (dimethylaminomethylene) amino-1- (2-propynyl) imidazole (0.2 g) Was treated with 6N hydrochloric acid to give 2-amino-1- (2-propynyl) imidazole hydrochloride (0.12 g, yield: 54%). Melting point: 119-120 ° C ¹ H-NMR (CDCl ₃ ) δ: 3.06 (1H, t, J = 2.5H
z), 4.84 (2H, d, J = 2.5Hz), 6.95 (1H, d, J
= 2.5 Hz), 7.03 (1 H, d, J = 2.5 Hz), 7.89
(2H, brs), 12.21 (1H, brs).

Example 3 Preparation of 2-Ethoxycarbonylamino-1-methylimidazole Hydrochloride 2-Amino-1-methylimidazole Hydrochloride (0.3 g)
Ethyl chlorocarbonate (0.6 g) was added to the pyridine (5 ml) solution of and the mixture was heated with stirring at 100 ° C for 2 hours. The solvent was distilled off under reduced pressure, the residue was dissolved in water, and sodium carbonate was added until the solution became alkaline. After distilling off the solvent under reduced pressure, the residue was subjected to silica gel column chromatography.
Purification with (developing solvent: chloroform) gave 2-ethoxycarbonylamino-1-methylimidazole. This was dissolved in dichloromethane, 4N hydrochloric acid-ethyl acetate (2 ml) was added to give the hydrochloride, and the obtained crude crystals were washed with ether to give 2-ethoxycarbonylamino-1-methylimidazole hydrochloride (0 .29 g, yield: 56%) was obtained as a colorless prism. Melting point: 129-131 ° C. Elemental analysis: Calculated as C ₇ H ₁₂ ClN ₃ O ₂ .0.2H ₂ O: C: 40.18%, H: 5.97%, N: 20.08
% Measured value: C: 40.00%, H: 6.40%, N: 20.05
% IR (KBr) cm ^-1 : 3170-2990,1745,164
0.1250 ¹ H-NMR (DMSO-d ₆ ) δ: 1.30 (3 H, t, J = 7.
1Hz), 3.66 (3H, s), 4.28 (2H, q, J = 7.1H
z), 7.32 (1H, d, J = 1.4Hz), 7.43 (1H, d, J
= 1.4 Hz).

Example 4 Preparation of 2-benzyloxycarbonylamino-1-methylimidazole-hydrochloride 2-amino-1-methylimidazole-hydrochloride (0.3 g)
Was dissolved in 1N aqueous sodium hydroxide solution (5 ml), benzyloxycarbonyl chloride (0.43 g) and ether
(2 ml) was added and stirred for 1 hour. The product was extracted with ethyl acetate, the extract was washed with water and saturated aqueous sodium chloride solution, dried over magnesium sulfate, and the solvent was evaporated under reduced pressure. Diethyl ether-hexane was added to the obtained residue to crystallize and collect. 2 thus obtained
The crystals of -benzyloxycarbonylamino-1-methylimidazole were dissolved in chloroform, 4N hydrochloric acid-ethyl acetate (0.5 ml) was added to give a hydrochloride, and the obtained crude crystals were washed with ether to give 2-benzyl. Oxycarbonylamino-1-methylimidazole hydrochloride (0.2 g, yield: 30%) was obtained as a white powder. Melting point: 141-142 ° C. Elemental analysis: Calculated as C ₁₂ H ₁₄ ClN ₃ O ₂ .0.2H ₂ O: C: 53.12%, H: 5.35%, N: 15.94
% Measured value: C: 53.17%, H: 5.24%, N: 15.45
% IR (KBr) cm ^-1 : 3070,1740,1625,125
0,1215 ¹ H-NMR (DMSO-d ₆ ) δ: 3.64 (3H, s), 5.2
9 (2H, s), 7.30 (1H, d, J = 2.2Hz), 7.40-
7.47 (6H, m).

Example 5 Preparation of 3- (2-aminoimidazol-1-yl) -L-alanine methyl ester dihydrochloride 3- (2-Dimethylaminomethyleneaminoimidazol-1-yl) -L-alanine methyl Ester (470
mg) was suspended in 20% hydrogen chloride-methanol (40 ml), and the mixture was stirred at 55 ° C for 6 days. The solvent was distilled off under reduced pressure, methanol was added to the obtained residue, and the solvent was distilled off again.
Methanol-ether was added to this substance and the mixture was allowed to stand, whereby colorless needle crystals were precipitated. The crystals were collected by filtration, washed with methanol and then with ether, and dried to obtain the title compound (292 mg). IR (KBr) cm ⁻¹ : 3380, 3200, 3150,
3060, 2950, 2900, 2830, 2780,
2630, 1750, 1660, 1520, 1435,
1360, 1290, 1260, 1150, 1125,
1075. ¹ H-NMR (DMSO-d ₆ ) δ: 3.78 (3H,
s), 4.48 (3H, s), 6.94 (1H, d, J =
2.4Hz), 6.98 (1H, d, J = 2.4Hz),
8.01 (2H, s), 9.02 (2H, brs).

Example 6 Preparation of 3- (2-Aminoimidazol-1-yl) -L-alanine dihydrochloride 3- (2-Aminoimidazol-1-yl) -L-alanine methyl ester dihydrochloride (103 mg) was dissolved in 6N hydrochloric acid (4 ml), and the mixture was stirred at 55 ° C for 5 days. Methanol was added to the residue obtained by evaporating the solvent under reduced pressure, the insoluble matter was filtered off, and the filtrate was evaporated under reduced pressure to give the title compound (57 mg). IR (KBr) cm ⁻¹ : 3360, 3150, 2970,
1740, 1660, 1535, 1500, 1450,
1260, 1120, 1090, 1040. ¹ H-NM
R (CD ₃ OD) δ: 4.42-4.60 (3H, m),
6.93 (2H, s).

Example 7 (2S) -2-t-butoxycarbonylamino-4-
[(1-Methylimidazol-2-yl) carbamoyl]
Preparation of methyl butyrate 2-amino-1-methylimidazole hydrochloride (668
mg) and N-t-butoxycarbonyl-L-glutamic acid α-methyl ester (1.31 g) were dissolved in dry pyridine (25 ml), phosphorus trichloride (687 mg) was added under ice cooling, and the mixture was returned to room temperature. Stir for hours. The precipitate was filtered off, the solvent was evaporated under reduced pressure, and the obtained residue was dissolved in chloroform, washed with saturated brine, and dried over anhydrous magnesium sulfate. The residue obtained by distilling off the solvent under reduced pressure was subjected to silica gel flash column chromatography (developing solvent: methanol-chloroform = 3:97 → 1:19).
The title compound (300 mg) was obtained by purification with. IR (KBr) cm ⁻¹ : 3370, 3130, 2970,
2950, 1735, 1680, 1555, 1520,
1480, 1460, 1440, 1360, 1320,
1300, 1260, 1180, 1160, 1080,
1060. ¹ H-NMR (DMSO-d ₆ ) δ: 1.39 (9H,
s), 1.60-2.10 (2H, m), 2.40 (2
H, t, J = 7.0 Hz), 3.39 (3H, s), 3.
63 (3H, s), 3.95-4.08 (1H, m),
6.73 (1H, d, J = 1.2Hz), 7.03 (1
H, d, J = 1.2 Hz), 7.29 (1 H, d, J =
8.0 Hz), 9.99 (1 H, s).

Example 8 Preparation of methyl (2S) -2-amino-4-[(1-methylimidazol-2-yl) carbamoyl] butyrate dihydrochloride (2S) -2-t-butoxycarbonylamino-4 −
[(1-Methylimidazol-2-yl) carbamoyl]
Methyl butyrate (34.1 mg) was dissolved in methanol (1 ml), 10% hydrogen chloride / ether (1 ml) was added, and the mixture was stirred at room temperature for 3 days.
Stir for hours. The solvent was distilled off under reduced pressure, and the residue was triturated in ether. This was collected by filtration and dried to obtain the title compound (27 mg). IR (KBr) cm ⁻¹ : 3420, 3130, 2950,
2920, 1740, 1710, 1615, 1555,
1480, 1440, 1350, 1280, 1230,
1170, 1140, 1070. ¹ H-NMR (CD ₃ OD) δ: 2.22-2.42 (2
H, m), 2.93 (2H, t, J = 7.2 Hz), 3.
80 (3H, s), 3.86 (3H, s), 4.19 (1
H, t, J = 7.2 Hz), 7.22 (1H, d, J =
2.2 Hz), 7.28 (1H, d, J = 2.2 Hz).

Example 9 Preparation of (2S) -2-amino-4-[(1-methylimidazol-2-yl) carbamoyl] butyric acid (2S) -2-t-butoxycarbonylamino-4-
[(1-Methylimidazol-2-yl) carbamoyl]
Methyl butyrate (341 mg) was dissolved in dichloromethane (5 ml), trifluoroacetic acid (1 ml) was added, and the mixture was stirred at room temperature for 2 hr. After adding methanol and toluene, the solvent was distilled off under reduced pressure. The obtained residue was dissolved in water (4 ml), 1N sodium hydroxide (4 ml) was added, and the mixture was stirred at room temperature for 3 hr. After neutralizing by adding 1N hydrochloric acid, the solvent was concentrated under reduced pressure to precipitate colorless needle crystals, which was collected by filtration, water,
The title compound (72 mg) was obtained by washing with methanol and ether and then drying. IR (KBr) cm ⁻¹ : 3440, 3180, 3070,
2980, 1695, 1645, 1620, 1550,
1480, 1440, 1420, 1400, 1355,
1320, 1280, 1230, 1200, 1180,
1155, 1140, 1080, 1045. ¹ H-NMR (D ₂ O) δ: 2.09-2.19 (2H,
m), 2.58 (2H, t, J = 7.4Hz), 3.45
(3H, s), 3.74 (1H, t, J = 6.2Hz),
6.84 (1H, d, J = 1.6Hz), 6.96 (1
H, d, J = 1.6 Hz).

Example 10 (4S) -4-t-butoxycarbonylamino-4-
[(1-Methylimidazol-2-yl) carbamoyl]
Production of benzyl butyrate The same procedure as in Example 7 was carried out to prepare 2-amino-1-methylimidazole hydrochloride (668 mg) and Nt-butoxycarbonyl-L-glutamic acid (γ-benzyl ester) (1.69 g). The title compound (857 mg) was obtained from dry pyridine (25 ml) and phosphorus trichloride (687 mg). IR (neat) cm ⁻¹ : 3400, 3280, 2970, ¹
730, 1700, 1585, 1550, 1480, 1
450, 1380, 1360, 1330, 1280, 1
240, 1160, 1050, 1020. ¹ H-NMR (CDCl ₃ ) δ: 1.45 (9H, s),
2.00-2.60 (4H, m), 3.49 (3H,
s), 4.23-4.46 (1H, m), 5.08 (2
H, s), 5.62 (1H, d, J = 7.0 Hz), 6.
51 (1H, d, J = 2.4Hz), 6.65 (1H,
d, J = 2.4 Hz), 7.33 (5H, s).

Example 11 Preparation of methyl (4S) -4-amino-4-[(1-methylimidazol-2-yl) carbamoyl] butyrate dihydrochloride (4S) -4-t-butoxycarbonylamino-4 −
[(1-Methylimidazol-2-yl) carbamoyl]
Benzyl butyrate (113 mg) was dissolved in methanol (2.7 ml), 20% hydrogen chloride-methanol (1.8 ml) was added, and the mixture was stirred at room temperature for 2 hours. The solvent was evaporated under reduced pressure and the residue was triturated in ether. The title compound (83 mg) was obtained by filtering this, washing with ether, and drying. IR (KBr) cm ⁻¹ : 3420, 3280, 3030,
2950, 2850, 1725, 1690, 1625,
1565, 1540, 1490, 1430, 1380,
1340, 1280, 1200, 1140, 1100,
1070. ¹ H-NMR (CD ₃ OD) δ: 2.29-2.43 (2
H, m), 2.67 (2H, t, J = 7.2 Hz), 3.
68 (3H, s), 3.87 (3H, s), 4.51 (1
H, t, J = 5.9 Hz), 7.31 (1H, d, J =
2.2 Hz), 7.3 (1 H, d, J = 2.2 Hz).

Example 12 (2S) -2-t-butoxycarbonylamino-3-
[(1-Methylimidazol-2-yl) carbamoyl]
Production of Methyl Propionate The same operation as in Example 7 was carried out to prepare 2-amino-1-methylimidazole hydrochloride (1.34 g) and Nt-butoxycarbonyl-L-aspartic acid (α-methyl ester) ( The title compound (300 mg) was obtained from 1.24 g), dry pyridine (35 ml) and phosphorus trichloride (687 mg). IR (KBr) cm ⁻¹ : 3420, 2970, 1740,
1710, 1585, 1550, 1490, 1390,
1360, 1340, 1280, 1240, 1210,
1160, 1050, 1020. ¹ H-NMR (CDCl ₃ ) δ: 1.45 (9H, s),
2.82 (1H, dd, J = 4.4Hz, 16.8H
z), 3.11 (1H, dd, J = 4.4Hz, 16.8)
Hz), 3.50 (3H, s), 3.73 (3H, s),
4.53-4.62 (1H, m), 5.40-5.80
(H, brs), 6.03 (1H, d, J = 8.2H
z), 6.55 (1H, d, J = 2.4 Hz), 6.67
(1H, d, J = 2.4 Hz).

Example 13 Methyl (2S) -2-amino-3-[(1-methylimidazol-2-yl) carbamoyl] propionate.2
Production of Hydrochloride The same procedure as in Example 11 was conducted to prepare methyl (2S) -2-t-butoxycarbonylamino-3-[(1-methylimidazol-2-yl) carbamoyl] propionate (258 mg) and methanol ( 7.9 ml) and 20% hydrogen chloride-methanol (5.0 ml) to give the title compound (220 m).
g) was obtained. IR (KBr) cm ⁻¹ : 3430, 3120, 2920,
1745, 1710, 1620, 1550, 1480,
1440, 1370, 1300, 1250, 1180,
1140, 1090, 1070. ¹ H-NMR (CD ₃ OD) δ: 3.40 (2H, d, J
= 5.6 Hz), 3.80 (3H, s), 3.87 (3
H, s), 4.55 (1H, t, J = 5.6 Hz), 7.
27 (1H, d, J = 2.4Hz), 7.33 (1H,
d, J = 2.4 Hz).

Example 14 Preparation of 2- (N-Boc Phenylalanyl) amino-1-methylimidazole 2-Amino-1-methylimidazole hydrochloride (0.3
g) was dissolved in water, neutralized with sodium carbonate, and the solvent was evaporated to dryness. Dissolve in DMF (5 ml), N-Boc phenylalanine (1.0 g), triethylamine (0.7 m).
l) was added, and under cooling with ice, diethyl phosphate cyanide (D
0.86 ml of EPC) was added dropwise, and the mixture was stirred at room temperature for 2 hours. 1 ml of methanol and water were added, and the mixture was extracted with ethyl acetate. The organic layer was washed with water and saturated brine, dried over magnesium sulfate, and the solvent was evaporated. The residue was purified by silica gel column chromatography (developing solvent: methanol-chloroform = 5:95), and the solvent of the target fraction was distilled off. A pale brown oily substance (0.39 g: 45.1%) was obtained. IR (KBr) cm ^-1 : 3280, 3030-2940,
1700, 1600-1550, 1490, 1390-
1365. ¹ H-NMR (CDCl ₃ ) δ: 1.42 (9H, s),
3.16-3.23 (2H, m), 3.46 (3H,
s), 4.52-4.57 (1H, m), 5.44 (1
H, d, J = 8.6 Hz), 6.54 (1H, d, J =
2.4Hz), 6.67 (1H, d, J = 2.4Hz),
7.21 (5H, s).

Example 15 Preparation of 1-methyl-2- (phenylalanyl) aminoimidazole dihydrochloride 2- (N-Boc phenylalanyl) amino-1-methylimidazole (0.3 g) was added to concentrated hydrochloric acid ( 10 ml) and stirred at room temperature for 30 minutes. The solvent was distilled off, and the precipitated colorless powder was collected by filtration using diethyl ether to obtain the title compound (0.23 g: 66.3%). Melting point: 231.0-232.0 ° C (decomposition). Elemental analysis: Calculated as C ₁₃ H ₁₈ Cl ₂ N ₄ O.0.5H ₂ O: C: 47.86%, H: 5.87%, N: 1
7.17%. Found: C: 47.48%, H: 5.79%, N: 1.
7.1%. IR (KBr) cm ⁻¹ : 3150-2850, 1705
1625, 1500. ¹ H-NMR (D ₂ O) δ: 3.17-3.40 (2H,
m), 3.43 (3H, s), 4.20 (1H, t, J =
6.6 Hz), 6.91 (1H, d, J = 2.4 Hz),
6.95 (1H, d, J = 2.4Hz), 7.29-7.4
0 (5H, m).

Example 16 (1) 2-Amino-1-methylimidazole 10.0 mg (2) Lactose 60.0 mg (3) Corn starch 35.0 mg (4) Gelatin 3.0 mg (5) Magnesium stearate 2.0 mg A mixture of 10.0 mg of 2-amino-1-methylimidazole, 60.0 mg of lactose and 35.0 mg of corn starch was added to 1 part.
0.03 ml of 0% gelatin aqueous solution (3.0 mg as gelatin)
, And after granulating through a 1 mm mesh sieve,
It was dried at ℃ and sieved again. The granules thus obtained were mixed with 2.0 mg of magnesium stearate and compressed. The obtained central tablets were sugar-coated with an aqueous solution of sucrose, titanium dioxide, talc and gum arabic. The coated tablets were polished with beeswax to obtain coated tablets.

Example 17 (1) 2-Amino-1-methylimidazole 10.0 mg (2) Lactose 70.0 mg (3) Corn starch 50.0 mg (4) Soluble starch 7.0 mg (5) Magnesium stearate 3. 0 mg 2-amino-1-methylimidazole 10.0 mg and magnesium stearate 3.0 mg were granulated with 0.07 ml of an aqueous solution of soluble starch (7.0 mg as soluble starch), dried and then lactose 70.0 mg and corn starch. 5
Mixed with 0.0 mg. The mixture was compressed to give tablets.

Example 18 (1) 2-Amino-1-methylimidazole 5.0 mg (2) Table salt 20.0 mg (3) Distilled water Total amount 2 ml 5.0-amino-1-methylimidazole 5.0 mg and table salt 20 Distilled water was added to make the total amount 2.0 mg. The solution was filtered and filled under aseptic conditions into 2 ml ampoules. The ampoule was sterilized and then sealed to obtain an injection solution.

[0078]

INDUSTRIAL APPLICABILITY According to the present invention, it has an excellent inhibitory effect on malignant tumors, and shows an excellent therapeutic effect particularly on colon cancer,
Provided is a glutaminase inhibitor useful as an anticancer agent, particularly a pharmaceutical composition for inhibiting glutaminase. Furthermore, novel imidazole derivatives having these malignant tumor suppressive effects are provided.

Claims (23)

[Claims] 1. A compound represented by the general formula (I): B represents a hydrogen atom or a chain hydrocarbon group which may have a substituent], or a salt thereof, or a pharmaceutical composition for inhibiting glutaminase. 2. The pharmaceutical composition according to claim 1, wherein A is an optionally protected amino group. 3. The pharmaceutical composition according to claim 1, wherein A is an amino group which may be protected by a group capable of leaving by in vivo metabolism. 4. The pharmaceutical composition according to claim 1, wherein A is an amino group which may be protected by an acyl group. 5. The pharmaceutical composition according to claim 1, wherein A is an amino group. 6. B is a linear or branched C _1-6 alkyl group, a linear or branched C _2-6 alkenyl group, or a straight chain which may each have a substituent. The pharmaceutical composition according to claim 1, which is a chain or branched C _2-6 alkynyl group. 7. The pharmaceutical composition according to claim 1, wherein B is a linear or branched C _1-6 alkyl group. 8. A chemical formula: The pharmaceutical composition according to claim 1, which is 9. The pharmaceutical composition according to claim 1, which is a mitochondrial glutaminase inhibitor. 10. The pharmaceutical composition according to claim 1, which is a therapeutic agent for a glutamine-requiring malignant tumor. 11. The pharmaceutical composition according to claim 1, which is a therapeutic agent for colorectal cancer or a malignant tumor derived from colorectal cancer. 12. The pharmaceutical composition according to claim 1, which is a therapeutic agent for colon cancer. 13. The pharmaceutical composition according to claim 1, which is a therapeutic agent for rectal cancer. 14. The compound represented by the general formula (I) is 2-
The pharmaceutical composition according to claim 1, which is amino-1-methylimidazole. 15. The compound represented by the general formula (I) is 2-
The pharmaceutical composition according to claim 1, which is benzyloxycarbonylamino-1-methylimidazole. 16. A compound represented by the general formula (I): B represents a hydrogen atom or a chain hydrocarbon group which may have a substituent]] or a salt thereof, which is a glutaminase inhibitor. 17. A compound represented by the general formula (II): [In the formula, A ′ represents an amino group which may be protected by an acyl group, and B ′ represents a chain hydrocarbon group which may have a substituent. However, when B'is methyl or ethyl, A'represents an amino group protected by an acyl group. ] The compound or its salt represented by these. 18. The compound according to claim 17, wherein A ′ is an amino group protected by an acyl group. 19. The compound according to claim 17, wherein A ′ is —NHCOOR (wherein R represents a lower alkyl group or a benzyl group) and B ′ is a lower alkyl group. 20. B ′ is a linear or branched C _1-6 alkyl group, a linear or branched C _2-6 alkenyl group each optionally having a substituent, or The compound according to claim 17, which is a linear or branched C _2-6 alkynyl group. 21. The compound according to claim 17, wherein B ′ is a linear or branched C _1-6 alkyl group. 22. The compound according to claim 17, which is 2-amino-1-methylimidazole. 23. The compound according to claim 17, which is 2-benzyloxycarbonylamino-1-methylimidazole.