Title of invention: Prodrug-type anticancer drug utilizing cancer-specific enzyme activity
Technical field
[0001]
　The present invention relates to a novel compound promising as a prodrug-type anticancer agent, a prodrug-type anticancer agent using the compound, and a pharmaceutical composition.
Background technology
[0002]
　In cancer chemotherapy, which still plays an important role in cancer treatment, the tissue, cells, and target selection system of the anticancer drug itself are not satisfactory, so serious side effects occur at the same time as the onset of drug efficacy. There is always the risk of doing it.
[0003]
　Prodrugization is known as one of the methods for specifically releasing these anticancer agents in cancer cells. Prodrugization is an approach in which a drug is structurally modified so that it changes into an active substance (anticancer drug) for the first time by an enzymatic reaction or a chemical reaction in cancer cells. On the other hand, it is difficult to identify an enzyme that is specifically promoted in cancer cells, and it is not easy to modify the structure of an existing anticancer drug having a complicated structure.
Outline of the invention
Problems to be solved by the invention
[0004]
　An object of the present invention is to provide a promising novel compound as a prodrug-type anticancer agent.
Means to solve problems
[0005]
　In our laboratory, we are conducting research aiming to establish a search platform technology that can comprehensively and non-invasively evaluate the metabolic reaction characteristics characteristic of disease sites on fresh clinical specimens. The present inventors efficiently develop highly specific therapeutic agents by a prodrug-like approach by utilizing useful information on the metabolic reactivity of diseased sites directly obtained from human clinical specimens. I thought it might be possible to do it.
[0006]
　In addition, in the research group of the present inventors, based on an idea based on quinone methide chemistry, it leaks from cells by emitting fluorescence by an enzymatic reaction and being tagged into intracellular thiols. We have developed a SPiDER probe that has the property of disappearing (Fig. 1).
　Here, when the present inventors are conducting live cell application experiments of the present probe, it has been clarified that remarkable cytotoxicity is observed when the probe is used at a high concentration. The mechanism of toxicity has not been clarified, but it is believed that thiols such as intracellular glutathione were consumed by the quinone methide intermediate produced by the enzymatic reaction, and oxidative stress was applied to the cells.
[0007]
　Therefore, the present inventors have conceived that it may be possible to selectively and strongly injure cells by means of cancer cell-specific enzyme activity by utilizing the above-mentioned phenomenon, and a novel prodrug-type anticancer drug. As a result of developing the agent, the present invention has been completed.
[0008]
　That is, the present invention is
a compound represented by the following general formula (I) or a salt thereof.

(In the formula,
X is a fluorine atom, an ester group (-OC (= O) -R'), a carbonate group (-OCO 2- R'), a carbamate group (-OCONH-R'), a phosphoric acid and an ester thereof. Selected from the group consisting of groups (-OP (= O) ( -OR') ( -OR ") , and sulfuric acid and its ester groups (-OSO 2- OR'),
　where R', R'". Are independently selected from substituted or unsubstituted alkyl groups or substituted or unsubstituted aryl groups;
Y is -NH-COL, -NH-L'or -OL',
　where In, L is a partial structure of an amino acid, and
　L'is a saccharide or a partial structure of a saccharide, a saccharide having a self-cleaving linker, an amino acid or peptide having a self-cleaving linker;
R 1 and R. 2 are each independently selected from a hydrogen atom or a monovalent substituent;
R 3 represents a hydrogen atom or 1 to 4 identical or different monovalent substituents present on the benzene ring).
[2] The partial structure of an amino acid of L, together with C = O to which it is bound, constitutes an amino acid, an amino acid residue, a peptide, or a part of an amino acid, according to [1]. Compound or salt thereof.
[3] The compound according to [1] or a salt thereof, wherein the partial structure of the saccharide of L'combines with O to which it is bound to form a saccharide, a part of the saccharide.
[4] -Y in the general formula (I) is bonded to -C (R 1 ) (R 2 ) X on the ortho-position or para-position of the benzene ring, [1] to [3]. The compound according to any one of the above items or a salt thereof.
[5] The compound according to any one of [1] to [4] or a salt thereof, wherein Y has a structure selected from the following.

[6] The compound according to any one of [1] to [5] or a salt thereof, wherein X is a fluorine atom or an ester group (-OCO-R').
[7] The compound according to any one of [1] to [6] or a salt thereof, wherein R 1 and R 2 are independently selected from a hydrogen atom or a fluorine atom.
[8] R 3The monovalent substituent is an alkyl group, an alkoxycarbonyl group, a nitro group, an amino group, a hydroxyl group, an alkylamino group (-NHR', -NHCOR'), an alkoxy group (-OR', -OCOR'), a halogen atom. , A boryl group, a cyano group (R'is a substituted or unsubstituted alkyl group, or a substituted or unsubstituted aryl group), any one of [1] to [7]. The compound according to the section or a salt thereof.
[9] R 3 monovalent substituent of an alkyl group (e.g., methyl group) or alkoxycarbonyl group (e.g., methoxycarbonyl group), a compound or a salt thereof according to [8].
[10] A prodrug-type anticancer agent containing the compound according to any one of [1] to [9] or a pharmaceutically acceptable salt thereof.
[11] A prodrug-type anti-prodrug that acts cell-selectively by cancer cell-specific enzymatic activity, including the compound according to any one of [1] to [9] or a pharmaceutically acceptable salt thereof. Cancer drug.
[12] The prodrug-type anticancer agent according to [11], wherein the enzyme is peptidase or glycosidase.
Is to provide.
Effect of the invention
[0009]
　INDUSTRIAL APPLICABILITY According to the present invention, a novel compound promising as a prodrug-type anticancer agent can be provided.
A brief description of the drawing
[0010]
FIG. 1 is a schematic diagram of fluorescence emission of a SPiDERβ-Gal probe.
FIG. 2 is a schematic diagram of the expression mechanism of the novel prodrug-type anticancer agent of the present invention.
[Fig. 3] Structure of a quinone methide-releasing prodrug compound.
FIG. 4 shows LC-MS analysis results of products obtained by in vitro reaction of compounds 1, 2 and 3 with β-Gal, DPP-IV and GGT, respectively.
FIG. 5: Test Results of CCK-8 Assay for Compound 1.
FIG. 6 is a test result of the CCK-8 assay of Compound 2.
FIG. 7: Test results of the CCK-8 assay for Compound 3.
FIG. 8 shows the results of a study on cell death observation in SHIN3 cells using Compound 3.
[Fig. 9] Results of studies on observation of cell death under co-culture conditions using Compound 3.
FIG. 10 shows the results of flow cytometric analysis of cell proliferation in SHIN3 cells and H226 cells using Compound 3.
FIG. 11 is a structural formula of a derivative having an acyl leaving group synthesized in the examples.
[Fig. 12] Results of confirmation of enzyme recognition ability using a benzyl-leaving group-converting derivative.
FIG. 13: Results of CCK-8 assay for benzyl leaving group conversion derivative.
FIG. 14 shows the results of the CCK-8 assay of the 4-position substituted derivative synthesized in the example.
FIG. 15: CCK-8 assay results for the 5-position substituted derivative synthesized in the Examples.
[Fig. 16] Outline of gGlu-FMA administration test for peritoneal dissemination model mice.
FIG. 17: Tumor imaging results on the mesentery (upper row: PBS-administered mouse, lower row: gGlu-FMA-administered mouse).
Mode for carrying out the invention
[0011]
　As used herein, the term "halogen atom" means a fluorine atom, a chlorine atom, a bromine atom, or an iodine atom.
[0012]
　In the present specification, "alkyl" may be any of linear, branched, cyclic, or a combination thereof, an aliphatic hydrocarbon group. The number of carbon atoms of the alkyl group is not particularly limited, but for example, the number of carbon atoms is 1 to 6 (C 1 to 6 ), the number of carbon atoms is 1 to 10 (C 1 to 10 ), and the number of carbon atoms is 1 to 15 (C 1 to 15). ), The number of carbon atoms is 1 to 20 (C 1 to 20 ). When the number of carbon atoms is specified, it means "alkyl" having the number of carbon atoms in the range of the number of carbon atoms. For example, C 1 ~ 8 Alkyl, methyl, ethyl, n- propyl, isopropyl, n- butyl, isobutyl, sec- butyl, tert- butyl, n- pentyl, isopentyl, neo-pentyl, n- hexyl, isohexyl, Includes n-heptyl, n-octyl and the like. As used herein, the alkyl group may have one or more arbitrary substituents. Examples of such a substituent include, but are not limited to, an alkoxy group, a halogen atom, an amino group, a mono or di-substituted amino group, a substituted silyl group, or an acyl. If the alkyl group has two or more substituents, they may be the same or different. The same applies to the alkyl moiety of other substituents containing an alkyl moiety (eg, an alkyl group, an arylalkyl group, etc.).
[0013]
　In the present specification, when a functional group is defined as "may be substituted", the type of substituent, the position of substitution, and the number of substituents are not particularly limited, and two or more substitutions are made. If they have groups, they may be the same or different. Examples of the substituent include, but are not limited to, an alkyl group, an alkoxy group, a hydroxyl group, a carboxyl group, a halogen atom, a sulfo group, an amino group, an alkoxycarbonyl group and an oxo group. Substituents may be further present in these substituents. Examples of such examples include, but are not limited to, alkyl halide groups, dialkylamino groups, and the like.
[0014]
　In the present specification, "aryl" may be either a monocyclic or condensed polycyclic aromatic hydrocarbon group, and as a ring-constituting atom, a hetero atom (for example, an oxygen atom, a nitrogen atom, or a sulfur atom) may be used. Etc.) may be an aromatic heterocycle containing one or more. In this case, it may be referred to as "heteroaryl" or "heteroaromatic". Whether the aryl is a monocyclic ring or a condensed ring, it can be bonded at all possible positions. Non-limiting examples of monocyclic aryls include phenyl group (Ph), thienyl group (2- or 3-thienyl group), pyridyl group, furyl group, thiazolyl group, oxazolyl group, pyrazolyl group, 2-pyrazinyl. Group, pyrimidinyl group, pyrrolyl group, imidazolyl group, pyridazinyl group, 3-isothiazolyl group, 3-isooxazolyl group, 1,2,4-oxadiazol-5-yl group or 1,2,4-oxadiazol-3 -Il groups and the like can be mentioned. Non-limiting examples of condensed polycyclic aryls include 1-naphthyl group, 2-naphthyl group, 1-indenyl group, 2-indenyl group, 2,3-dihydroindene-1-yl group, 2,3. -Dihydroindene-2-yl group, 2-anthryl group, indazolyl group, quinolyl group, isoquinolyl group, 1,2-dihydroisoquinolyl group, 1,2,3,4-tetrahydroisoquinolyl group, indolyl group, Isoindrill group, phthalazinyl group, quinoxalinyl group, benzofuranyl group, 2,3-dihydrobenzofuran-1-yl group, 2,3-dihydrobenzofuran-2-yl group, 2,3-dihydrobenzothiophen-1-yl group, 2 , 3-Dihydrobenzothiophen-2-yl group, benzothiazolyl group, benzimidazolyl group, fluorenyl group, thioxanthenyl group and the like. In the present specification, the aryl group may have one or more arbitrary substituents on its ring. Examples of the substituent include, but are not limited to, an alkoxy group, a halogen atom, an amino group, a mono or di-substituted amino group, a substituted silyl group, and an acyl. Substituents with two or more aryl groups If they have, they may be the same or different. The same applies to the aryl moiety of other substituents containing an aryl moiety (such as an aryloxy group and an arylalkyl group).
[0015]
　In the present specification, "arylalkyl" represents an alkyl substituted with the above aryl. The arylalkyl may have one or more arbitrary substituents. Examples of the substituent include, but are not limited to, an alkoxy group, a halogen atom, an amino group, a mono or di-substituted amino group, a substituted silyl group, or an acyl group. If the acyl group has two or more substituents, they may be the same or different. Non-limiting examples of arylalkyl include benzyl group, 2-thienylmethyl group, 3-thienylmethyl group, 2-pyridylmethyl group, 3-pyridylmethyl group, 4-pyridylmethyl group, 2-furylmethyl group, 3-furylmethyl group, 2-thiazolylmethyl group, 4-thiazolylmethyl group, 5-thiazolylmethyl group, 2-oxazolylmethyl group, 4-oxazolylmethyl group, 5-oxazolylmethyl group, 1-pyrazolylmethyl group , 3-Pyrazolyl Methyl Group, 4-Pyrazolyl Methyl Group, 2-Pyrazinyl Methyl Group, 2-Pyrimidinyl Methyl Group, 4-Pyrimidinyl Methyl Group, 5-Pyrimidinyl Methyl Group, 1-Pyrrolyl Methyl Group, 2-Pyrrolyl Methyl Group, 3-Pyrrolyl Methyl Group , 1-imidazolylmethyl group, 2-imidazolylmethyl group, 4-imidazolylmethyl group, 3-pyridazinylmethyl group, 4-pyridazinylmethyl group, 3-isothiazolylmethyl group, 3-isooxazoli Examples thereof include a rumethyl group, a 1,2,4-oxadiazol-5-ylmethyl group or a 1,2,4-oxadiazol-3-ylmethyl group.
[0016]
　In the present specification, the "alkoxy group" has a structure in which the alkyl group is bonded to an oxygen atom, and examples thereof include a saturated alkoxy group which is a linear group, a branched group, a cyclic group, or a combination thereof. For example, methoxy group, ethoxy group, n-propoxy group, isopropoxy group, cyclopropoxy group, n-butoxy group, isobutoxy group, s-butoxy group, t-butoxy group, cyclobutoxy group, cyclopropylmethoxy group, n- Pentyloxy group, cyclopentyloxy group, cyclopropylethyloxy group, cyclobutylmethyloxy group, n-hexyloxy group, cyclohexyloxy group, cyclopropylpropyloxy group, cyclobutylethyloxy group, cyclopentylmethyloxy group and the like are preferable. Take as an example.
[0017]
　As used herein, "alkylene" is a divalent group consisting of linear or branched saturated hydrocarbons, such as methylene, 1-methylmethylene, 1,1-dimethylmethylene, ethylene, and the like. 1-Methylethylene, 1-ethylethylene, 1,1-dimethylethylene, 1,2-dimethylethylene, 1,1-diethylethylene, 1,2-diethylethylene, 1-ethyl-2-methylethylene, trimethylene, 1 -Methyltrimethylene, 2-methyltrimethylene, 1,1-dimethyltrimethylene, 1,2-dimethyltrimethylene, 2,2-dimethyltrimethylene, 1-ethyltrimethylene, 2-ethyltrimethylene, 1,1 -Diethyltrimethylene, 1,2-diethyltrimethylene, 2,2-diethyltrimethylene, 2-ethyl-2-methyltrimethylene, tetramethylene, 1-methyltetramethylene, 2-methyltetramethylene, 1,1- Examples thereof include dimethyltetramethylene, 1,2-dimethyltetramethylene, 2,2-dimethyltetramethylene, and 2,2-di-n-propyltrimethylene.
[0018]
1. 1. A compound represented by the general formula (I) or a salt thereof
　One embodiment of the present invention is a compound represented by the following general formula (I) or a salt thereof.

[0019]
　That is, based on the idea based on quinone methide chemistry, the present inventors have the above-mentioned property of SPIDER having the property of emitting fluorescence by an enzymatic reaction and tagging into intracellular thiols so as not to leak from cells. As a result of molecular design of a compound that is activated by an enzyme activity specific to cancer cells and releases quinonemethide by utilizing the findings of the research of the present inventors obtained from a probe or the like, the above general formula (I) It was found that the compound represented by () can cause strong cell-selective damage due to cancer cell-specific enzyme activity and is useful as a novel prodrug-type anticancer agent (see FIG. 2). ).
[0020]
　Here, in the general formula (I), Y is an enzyme recognition site, and a part thereof is cleaved by a cancer cell-specific enzyme activity to induce the formation of quinone methide.
　Y can be selected according to the type of enzyme. When the target enzyme of the prodrug-type anticancer drug is glycosidase, Y is selected from the groups derived from saccharides, and when the target enzyme is peptidase, Y contains groups derived from amino acids and amino acids. Selected from the group.
[0021]
　In general formula (I), Y is preferably -NH-CO-L, -NH-L'or -OL'.
　Here, L is a partial structure of an amino acid. The partial structure of the amino acid of L means that the amino acid, the amino acid residue, the peptide, and a part of the amino acid are formed together with C = O to which L is bound.
[0022]
　As used herein, the term "amino acid" can be any compound as long as it has both an amino group and a carboxyl group, and includes natural and non-natural compounds. It may be a neutral amino acid, a basic amino acid, or an acidic amino acid, and in addition to an amino acid that itself functions as a transmitter such as a neurotransmitter, a physiologically active peptide (dipeptide, tripeptide, tetrapeptide, as well as Amino acids that are constituents of polypeptide compounds such as (including oligopeptides) and proteins can be used, and may be, for example, α-amino acids, β-amino acids, γ-amino acids, and the like. As the amino acid, it is preferable to use an optically active amino acid. For example, as the α-amino acid, either D- or L-amino acid may be used, but it may be preferable to select an optically active amino acid that functions in a living body.
[0023]
　As used herein, the term "amino acid residue" refers to a structure corresponding to the remaining partial structure obtained by removing a hydroxyl group from a carboxyl group of an amino acid.
　Amino acid residues include α-amino acid residues, β-amino acid residues, and γ-amino acid residues. Preferred amino acid residues include a GGT substrate “γ-glutamyl group” and a DPP4 substrate dipeptide “amino acid-proline dipeptide”.
[0024]
　L'is a saccharide or a partial structure of a saccharide, a saccharide having a self-cleaving linker, an amino acid or peptide having a self-cleaving linker.
　The partial structure of the sugar of L'combines with O to which L'is bound to form a part of the sugar and the sugar.
[0025]
　Examples of saccharides include β-D-glucose, β-D-galactose, β-L-galactose, β-D-xylose, α-D-mannose, β-D-fucose, α-L-fucose, and β-L-. Examples thereof include fucose, β-D-arabinose, β-L-arabinose, β-DN-acetylglucosamine, β-DN-acetylgalactosamine and the like, and β-D-galactose is preferable.
[0026]
　The self-cleaving linker means a linker that is spontaneously cleaved and decomposed, and examples thereof include carbamate, urea, and paraaminobenzyloxy group.
[0027]
　In one preferred aspect of the invention, Y has a structure selected from:

[0028]
　In the general formula (I), X acts as a leaving group that desorbs from the benzene ring by cleaving a part of the enzyme recognition site of Y by a cancer cell-specific enzyme activity, and as a result. , Quinone methide is formed.
[0029]
　X is a fluorine atom, an ester group (-OC (= O) -R'), a carbonate group (-OCO 2- R'), a carbamate group (-OCONH-R'), phosphoric acid and an ester group thereof (-OP). It is selected from the group consisting of (= O) ( -OR') ( -OR ") , and sulfuric acid and its ester group (-OSO 2- OR'), where R'and R'are
　, respectively. It is independently selected from a substituted or unsubstituted alkyl group or a substituted or unsubstituted aryl group. As
　X, a fluorine atom or an ester group (-OCO-R') is preferable. It is bound by theory. Although not intended, when X is a fluorine atom or an ester group (-OCO-R'), quinonemethide is formed promptly when Y is cleaved.
[0030]
　R 1 and R 2 are each independently selected from a hydrogen atom or a monovalent substituent. The monovalent substituent is a halogen atom or an alkyl group having 1 or more carbon atoms (for example, an alkyl group having 1 to 6 carbon atoms).
　R 1 and R 2 are preferably independently selected from hydrogen or fluorine atoms, respectively.
[0031]
It is preferable that 　-Y in the general formula (I) is bonded to -C (R 1 ) (R 2 ) X at the ortho-position or para-position of the benzene ring. When -Y and -C (R 1 ) (R 2 ) X have such a positional relationship on the benzene ring, a quinone methide structure can be formed when Y is cleaved.
[0032]
　R 3 represents a hydrogen atom or 1 to 4 identical or different monovalent substituents present on the benzene ring.
　As the monovalent substituent of R 3 , an alkyl group having 1 or more carbon atoms (for example, an alkyl group having 1 to 6 carbon atoms), an alkoxycarbonyl group, a nitro group, an amino group, a hydroxyl group, and an alkylamino group (-NHR). It is selected from the group consisting of', -NHCOR'), an alkoxy group (-OR', -OCOR'), a halogen atom, a boryl group, and a cyano group. Here, R'is a substituted or unsubstituted alkyl group or a substituted or unsubstituted aryl group. The monovalent substituent of
　R 3 is preferably an alkyl group having 1 or more carbon atoms (for example, an alkyl group having about 1 to 6 carbon atoms (for example, a methyl group)). Although intended to be bound by theory, alkyl groups can improve electron donating and cell killing activity.
[0033]
　As the position of R 3, the 5th position corresponding to the para position of −C (R 1 ) (R 2 ) X or the 4th position corresponding to the meta position is preferable.
[0034]
　Further, the compound represented by the general formula (I) also includes three isomers such as tautomers, geometric isomers (for example, E isomers, Z isomers, etc.) and enantiomers, unless otherwise specified. .. That is, when one or two or more asymmetric carbons are contained in the compound represented by the general formula (I), the stereochemistry of the asymmetric carbons is independently the (R) form or (S). ) Can take any of the isomers and may exist as a stereoisomer such as an enantiomer of the derivative or a diastereoisomer. Therefore, as the active ingredient of the microtubule polymerization inhibitor of the present invention, any stereoisomer in pure form, any mixture of stereoisomers, racemate, etc. can be used, all of which are of the present invention. Included in the range.
[0035]
　Non-limiting examples of the compound represented by the general formula (I) or a salt thereof are shown below.

[0036]
2. 2. Prodrug-type anti-cancer agent
　Another embodiment of the present invention is a pro-drug-type anti-cancer agent containing a compound of the general formula (I) or a pharmaceutically acceptable salt thereof (hereinafter, "the present invention"). Also known as "prodrug-type anticancer drug").
　In addition, another embodiment of the present invention is a prodrug-type anti-drug that acts cell-selectively by a cancer cell-specific enzyme activity, which comprises a compound of general formula (I) or a pharmaceutically acceptable salt thereof. It is a cancer drug.
[0037]
　Cancer cell-specific enzymes are peptidases or glycosidases.
　Examples of the peptidase include γ-glutamyl transpeptidase (GGT), dipeptidyl peptidase IV (DPP-IV), and calpain.
　Examples of glycosidases include β-galactosidase, β-glucosidase, α-mannosidase, α-L-fucosidase, β-hexosaminidase, β-N-acetylgalactosaminidase and the like.
[0038]
　In addition, another embodiment of the present invention comprises a compound of general formula (I) or a pharmaceutically acceptable salt thereof, such as breast cancer, esophageal cancer, lung cancer, head and neck cancer, oral cancer, liver cancer and the like. It is a pharmaceutical composition for treating or preventing cancer (hereinafter, also referred to as "pharmaceutical composition of the present invention").
[0039]
　In addition, another embodiment of the present invention is a method for treating cancers such as breast cancer, esophageal cancer, lung cancer, head and neck cancer, oral cancer, and liver cancer in mammals, particularly humans, and such treatment is performed. An effective amount of the compound of the present invention of the general formula (I) or a pharmaceutically acceptable salt thereof, or the compound represented by the general formula (I) or a pharmaceutically acceptable salt thereof is pharmaceutically acceptable to the required mammal. It is a method of administering a pharmaceutical composition containing a salt.
[0040]
　The prodrug-type anticancer agent or pharmaceutical composition of the present invention may contain not only the compound represented by the general formula (I) but also a salt thereof or a solvate or hydrate thereof. .. The salt is not particularly limited as long as it is a pharmaceutically acceptable salt, and examples thereof include a base addition salt, an acid addition salt, and an amino acid salt. Examples of the base addition salt include alkaline earth metal salts such as sodium salt, potassium salt, calcium salt and magnesium salt, ammonium salt, and organic amine salts such as triethylamine salt, piperidine salt and morpholin salt. Examples of acid addition salts include mineral acid salts such as hydrochloride, hydrobromide, sulfate, nitrate and phosphate; methanesulfonic acid, benzenesulfonic acid, paratoluenesulfonic acid, acetic acid, propionate, etc. Organic acid salts such as tartrate acid, fumaric acid, maleic acid, malic acid, oxalic acid, succinic acid, citric acid, benzoic acid, mandelic acid, silicic acid, lactic acid, glycolic acid, glucuronic acid, ascorbic acid, nicotinic acid and salicylic acid. Can be mentioned. Examples of the amino acid salt include glycine salt, aspartate, glutamic acid and the like. Further, it may be a metal salt such as an aluminum salt.
[0041]
　The type of solvent that forms the solvate is not particularly limited, and examples thereof include solvents such as ethanol, acetone, and isopropanol.
[0042]
　As diseases that can be treated or prevented by the prodrug-type anticancer agent of the present invention, great effects can be expected for a wide range of cancer types, particularly breast cancer, esophageal cancer, lung cancer, head and neck cancer, oral cancer, liver cancer and the like. ..
[0043]
　The prodrug-type anticancer agent or pharmaceutical composition of the present invention contains the active ingredient compound represented by the general formula (I) or a pharmaceutically acceptable salt, hydrate, or solvate itself. Although it may be administered, it is generally desirable to administer it in the form of a pharmaceutical composition containing the above-mentioned substance as an active ingredient and one or more pharmaceutical additives. The term "composition", such as in pharmaceutical compositions, refers to any two or more products as well as products containing the active ingredient and the Inactive ingredients (pharmaceutically acceptable excipients) that make up the carrier. It occurs directly or indirectly as a result of component association, compositing or aggregation, or as a result of dissociation of one or more components, or as a result of another type of reaction or interaction of one or more components. Includes any product.
[0044]
　As the active ingredient of the prodrug-type anticancer agent or the pharmaceutical composition of the present invention, two or more of the above compounds can be used in combination.
[0045]
　In addition, the prodrug-type anticancer agent or pharmaceutical composition of the present invention is an active ingredient compound represented by the general formula (I) or a pharmaceutically acceptable salt, hydrate, or solvate thereof. It is also possible to make a combination drug in combination with an existing anticancer drug. As the existing anticancer agent, those known in the art can be used, and examples thereof include methotrexate, doxorubicin, cisplatin and the like.
[0046]
　The type of the prodrug-type anticancer agent or pharmaceutical composition of the present invention is not particularly limited, and the dosage form includes tablets, capsules, granules, powders, syrups, suspensions, suppositories, ointments, and creams. Examples include agents, gels, patches, inhalants, injections and the like. These formulations are prepared according to conventional methods. In the case of a liquid preparation, it may be dissolved or suspended in water or another suitable solvent at the time of use. Further, tablets and granules may be coated by a well-known method. In the case of an injection, the compound of the present invention is prepared by dissolving it in water, but it may be dissolved in physiological saline or a glucose solution as needed, or a buffer or a preservative may be added. Good. It is provided in any form of formulation for oral or parenteral administration. For example, pharmaceutical compositions for oral administration in the form of granules, fine granules, powders, hard capsules, soft capsules, syrups, emulsions, suspensions or liquids, for intravenous administration, for intramuscular administration, Alternatively, it can be prepared as a pharmaceutical composition for parenteral administration in the form of an injection, a drip, a transdermal absorbent, a transmucosal absorbent, a nasal drop, an inhalant, a suppository, etc. for subcutaneous administration. Injections, drip infusions, and the like can be prepared as powdered dosage forms such as freeze-dried forms, and can be used by dissolving them in an appropriate aqueous medium such as physiological saline at the time of use. It is also possible to directly administer a sustained-release preparation coated with a polymer or the like into the brain.
[0047]
　The type of pharmaceutical additive used in the production of the prodrug-type anticancer agent or pharmaceutical composition of the present invention, the ratio of the pharmaceutical additive to the active ingredient, or the method for producing the pharmaceutical composition is determined in the form of the composition. Those skilled in the art can appropriately select it accordingly. Inorganic or organic substances or solid or liquid substances can be used as the additives for preparation, and generally, they can be blended in an amount of 1% by weight to 90% by weight based on the weight of the active ingredient. Specifically, examples of such substances include lactose, glucose, mannit, dextrin, cyclodextrin, starch, crust, magnesium aluminometasilicate, synthetic aluminum silicate, sodium carboxymethyl cellulose, hydroxypropyl starch, calcium carboxymethyl cellulose. , Ion exchange resin, methyl cellulose, gelatin, gum arabic, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, polyvinyl pyrrolidone, polyvinyl alcohol, light anhydrous silicic acid, magnesium stearate, talc, tragant, bentonite, beagum, titanium oxide, sorbitan fatty acid ester Examples thereof include sodium lauryl sulfate, glycerin, fatty acid glycerin ester, purified lanolin, glycero gelatin, polysorbate, macrogol, vegetable oil, wax, liquid paraffin, white vaseline, fluorocarbon, nonionic surfactant, propylene glycol, water and the like.
[0048]
　To produce a solid preparation for oral administration, the active ingredient and excipient ingredients such as lactose, starch, crystalline cellulose, calcium lactate, silicic anhydride, etc. are mixed to form a powder, or sucrose, if necessary. A binder such as hydroxypropyl cellulose or polyvinylpyrrolidone, a disintegrant such as carboxymethyl cellulose or carboxymethyl cellulose calcium is added, and wet or dry granulation is performed to obtain granules. In order to produce tablets, these powders and granules may be tableted as they are or by adding a lubricant such as magnesium stearate or talc. These granules or tablets may be coated with an enteric solvent base such as hydroxypropylmethylcellulose phthalate or methacrylic acid-methylmethacrylic acid polymer to prepare an enteric solvent preparation or a sustained preparation by coating with ethyl cellulose, carnauba wax, curing oil or the like. it can. To manufacture capsules, hard capsules are filled with powders or granules, or the active ingredient is used as it is or dissolved in glycerin, polyethylene glycol, sesame oil, olive oil, etc. and then coated with a gelatin film to form soft capsules. Can be done.
[0049]
　In order to manufacture injectables, the active ingredient may be used as a pH adjuster such as hydrochloric acid, sodium hydroxide, lactose, lactic acid, sodium, sodium monohydrogen phosphate, sodium dihydrogen phosphate, sodium chloride, glucose, etc. It may be dissolved in distilled water for injection together with an isotonic agent, filtered aseptically and filled in an ampol, or it may be vacuum freeze-dried by adding mannitol, dextrin, cyclodextrin, gelatin, etc. to prepare a solution-type injection. .. Further, reticine, polysorbate 80, polyoxyethylene hydrogenated castor oil and the like can be added to the active ingredient to emulsify in water to prepare an emulsion for injection.
[0050]
　The dose and frequency of administration of the prodrug-type anticancer agent or pharmaceutical composition of the present invention are not particularly limited, and the prevention and / or purpose of treatment, type of disease, weight of patient, etc. It is possible to make an appropriate selection at the discretion of the doctor according to the conditions such as age and severity of the disease. Generally, the daily dose for an adult in oral administration is about 0.01 to 1000 mg (weight of active ingredient), and it may be administered once or divided into several times a day, or every few days. it can. When used as an injection, it is desirable to administer a daily dose of 0.001 to 100 mg (weight of active ingredient) continuously or intermittently to an adult.
[0051]
　The method for producing the compound represented by the general formula (I) is not particularly limited, but a method for synthesizing a representative compound among the compounds included in the general formula (I) is specifically shown in Examples of the present specification. It was. Those skilled in the art can obtain the compounds included in the formula (I) by appropriately modifying or modifying the starting materials, reaction reagents, reaction conditions, etc., as necessary, with reference to the examples of the present specification and the scheme below. Can be manufactured.
Example
[0052]
　Hereinafter, the present invention will be described with reference to Examples, but the present invention is not limited thereto.
[0053]
1. 1. Synthesis of quinone methide-releasing prodrug compound
　First, as shown in FIG. 3, a monocyclic compound having fluorine at the enzyme recognition site and leaving group of three types of enzymes (β-Gal, DPP-IV, GGT) was selected. It was synthesized by the following procedure.
[0054]
[Synthesis Example 1]
　Compound 1 (β-Gal-FMP) of the present invention was synthesized by the following scheme 1.
[0055]
Scheme 1

[0056]
(1) Synthesis of compound 1

[0057]
　2-(((tert-Butyldimethylsilyl) oxy) methyl) phenol (301 mg, 1.26 mmol) and cesium carbonate (4.11 g, 12.6 mmol) are dissolved in dehydrated DMF (7 mL) at 0 ° C. under an argon atmosphere. Was stirred for 5 minutes. (2R, 3S, 4S, 5R, 6R) -2- (acetoxymethyl) -6-bromotetrahydro-2H-pyran-3,4,5-trityltriacetate (4.51 g, 11.0 mmol) was added and 0 ° C. Was stirred for 5 hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous sodium hydrogen carbonate solution and brine, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (626.8 mg, 87%).
1 1 H NMR (CD 2 Cl 2 , 400 MHz): δ7.50 (d, 1H, J = 8.0 Hz), 7.20 (dd, 1H, J = 8.0 Hz, J = 7.3 Hz), 7. 08 (dd, 1H, J = 7.8Hz, J = 7.3Hz), 7.00 (d, 1H, J = 7.8Hz), 5.47-5.43 (m, 2H), 5.13 (Dd, 1H, J = 11Hz, J = 3.7Hz), 5.08 (d, 1H, J = 8.2Hz), 4.75 (d, 1H, J gem = 15Hz), 4.62 (d) , 1H, J gem = 15Hz), 4.22-4.09 (m, 3H), 2.16 (s, 3H, OCOCH 3)), 2.04 (s, 3H, OCOCH 3 ), 2.03 (s3H, OCOCH 3 ), 1.98 (s, 3H, OCOCH 3 ), 0.96 (s, 9H, Si (CH 3 ) 3 ) ), 0.12 (s, 6H, Si (CH 3 ) 2 ).
[0058]
(2) Synthesis of compound 2

[0059]
　Compound 1 (76.7 mg, 0.135 mmol) was dissolved in dehydrated dichloromethane (5 mL) and cooled to −78 ° C. TBAF (ca. 1 mol / L in THF, 39 μL, 0.135 mmol) was added, and the mixture was stirred at −78 ° C. for 1 hour. Subsequently, Deoxo-Flour® (132 μL, 0.675 mmol) was added, and the mixture was further stirred for 1 hour. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with dichloromethane. The ethyl acetate layer was washed with water and brine, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a pale yellow liquid (29.7 mg, 48%).
1 H NMR (CDCl 3 , 400 MHz): δ7.38 (d, 1H, J = 7.8 Hz), 7.32 (dd, 1H, J = 7.8 Hz, J = 7.8 Hz), 7.10 ( dd, 1H, J = 8.2Hz, J = 7.8Hz), 7.07 (d, 1H, J = 8.2Hz), 5.54 (dd, 1H, J = 11Hz, J = 7.8Hz) , 5.47 (dd, 1H, J HBn-F = 48Hz, J gem = 11Hz ), 5.46 (d, 1H, J = 2.7Hz), 5.23 (dd, 1H, J HBn-F = 48Hz, J gem= 11Hz), 5.11 (dd, 1H, J = 11Hz, J = 3.7Hz), 5.03 (d, 1H, J = 7.8Hz), 4.25 (dd, 1H, J = 11Hz, J = 6.9Hz), 4.15 (dd, 1H, J = 11Hz, J = 6.0Hz), 4.08 (dd, 1H, J = 6.9Hz, J = 6.0Hz), 2.18 (S, 3H, OCOCH 3 ), 2.06 (s, 3H, OCOCH 3 ), 2.06 (s, 3H, OCOCH 3 ), 2.01 (s, 3H, OCOCH 3 ).
[0060]
(3) Synthesis of compound 3

[0061]
　Compound 2 (29.7 mg, 0.0650 mmol) was dissolved in dehydrated methanol (5 mL) and cooled to 0 ° C. 28% NaOMe / MeOH (50 μL) was added and the mixture was stirred at −78 ° C. for 12 hours. After confirming the completion of the reaction, neutralization was carried out with Amberlite IR120 (registered trademark), and filtration and concentration were carried out. The residue was produced by reverse phase HPLC (0% → 100% acetonitrile / water) to obtain the desired product as a white solid (2.25 mg, 12%).
1 H NMR (CD 3 OD, 400 MHz): δ7.35 (d, 1H, Ha, JHa -Hb = 7.3 Hz), 7.29 (dd, 1H, Hc, J Hc-Hd = 8.2 Hz, J Hc-Hb = 7.3Hz), 7.21 (d, 1H, Hd, J Hd-Hc = 8.2Hz), 7.03 (dd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz), 5.51 (d, 2H, H Bn , J HBn-F = 48 Hz), 4.85 (d, IH, H1, JH1-H2 = 7.8 Hz), 3.88 (d, IH , H4, J H4-H3 = 3.7Hz), 3.79 (dd, 1H, H2, J H2-H3)= 9.6Hz, J H2-H1 = 7.8Hz), 3.78-3.70 (m, 2H, H6, 6'), 3.65 (dd, 1H, H5, J H5-H6 = 6. 9Hz, J H5-H6' = 5.0Hz), 3.55 (dd, 1H, H3, J H3-H2 = 9.6Hz, J H3-H4 = 3.7Hz); HRMS 311.08976 (M + Na + ) ..
[0062]
[Synthesis Example 2]
　Compound 2 (GP-FMA) of the present invention was synthesized by the following scheme 2.
[0063]
Scheme 2

[0064]
(1) Synthesis of compound 4

[0065]
　tert-Butyl (S)-(2-(2-((2-(((tert-butyldimethylsilyl) oxy) methyl) phenyl) carbamoyl 7) pyrrolidine-1-yl) -2-oxoethyl) carbamate (86. 4 mg (0.176 mmol) was dissolved in dehydrated DMF (2 mL) and cooled to 0 ° C. HATU (206 mg, 0.532 mmol) and DIPEA (183 μL, 1.06 mmol) were added and stirred at 0 ° C. for 5 minutes. Subsequently, 2- (tert-butyldimethylsilyl) oxy) methyl) aniline (101 mg, 0.425 mmol) dissolved in dehydrated DMF (1 mL) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saline solution, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 40% → 50% ethyl acetate / hexane) to obtain the desired product as a white solid (88.1 mg, 51%).
1 H NMR (CD 3 OD, 400 MHz): Deruta7.44-7.41 (m, 2H), 7.28-7.18 (m, 2H), 4.75 (d, IH, HBn, J gem = 14Hz), 4.70 (d, 1H, HBn', J gem = 14Hz ), 4.53 (dd, 1H, J = 8.5Hz, J = 2.7Hz), 3.95 (d, 1H, J) gem = 17Hz), 3.88 (d, 1H, J gem)= 17Hz), 3.71-3.56 (m, 2H), 2.17-1.91 (m, 3H), 1.41 (s, 9H, NHCOO (CH 3 ) 3), 0.91 ( s, 9H, Si (CH 3 ) 3 ), 0.09 (s, 6H, Si (CH 3 ) 2 ).
[0066]
(2) Synthesis of compound 5

[0067]
　Compound 4 (96.5 mg, 0.354 mmol) was dissolved in dehydrated THF (5 mL), TBAF (ca. 1 mol / L in THF, 879 μL, 0.879 mmol) was added, and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was produced by silica gel chromatography (14 g silica gel, 0% → 7% methanol / dichloromethane) to obtain the desired product as a colorless liquid (63.9 mg, 96%).
1 H NMR (CD 3 OD, 400 MHz): δ7.70 (d, 1H, J = 7.8 Hz), 7.30 (d, 1H, J = 7.8 Hz), 7.25 (ddd, 1H, J) = 7.8Hz, J = 7.8Hz, J = 1.4Hz), 7.13 (ddd, 1H, J = 7.8Hz, J = 7.8Hz, J = 1.4Hz), 4.61 (dd) , 1H, HBn, J gem = 14Hz, J = 4.1Hz), 4.57 (dd, 1H, HBn, J gem = 14Hz, J = 4.1Hz), 4.55 (dd, 1H, J = 8) .5Hz, J = 3.7Hz), 3.94 (s, 2H), 3.73-3.55 (m, 2H), 2.30-1.12 (m, 2H), 2.10-2 .03 (m, 2H), 1.43 (s, 9H, NHCOO (CH 3 ) 3 ).
[0068]
(3) Synthesis of compound 6

[0069]
　Compound 5 (63.9 mg, 0.169 mmol) was dissolved in dehydrated dichloromethane (2 mL) and cooled to 0 ° C. Deoxo-Flour® (166 μL, 0.847 mmol) was added, and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saline solution, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 50% → 70% ethyl acetate / hexane) to obtain the desired product as a pale yellow liquid (22.9 mg, 36%).
1 H NMR (CDCl 3 , 400 MHz): δ8.93 (brs, 1H, -CONH-), 7.95 (d, 1H, J = 7.8 Hz), 7.36 (dd, 1H, J = 7. 8Hz, J = 7.8Hz), 7.29 (d, 1H, J = 7.3Hz), 7.13 (dd, 1H, J = 7.8Hz, J = 7.3Hz), 5.41 (brs) , 1H, -OCONH-), 5.39 (d, 2H, HBn, J HBn-F = 48Hz), 4.76 (d, 1H, J = 6.9Hz), 4.03 (dd, 1H, J) gem = 17Hz, J = 5.0Hz), 3.93 (dd, 1H, J gem = 17Hz, J = 4.6Hz), 3.60-3.54 (m, 1H), 3.49-3. 40 (m, 1H), 2.22-2.11 (m, 1H), 2.11-2.02 (m, 1H), 2.01-1.90 (m, 1H), 1.44 ( s, 9H, NHCOO (CH 3 ) 3 ).
[0070]
(4) Synthesis of compound 7

[0071]
　Compound 6 (22.9 mg, 0.0604 mmol) was dissolved in ethyl acetate (1 mL) and 4M. Hydrochloric acid / ethyl acetate (2 mL) was added, and the mixture was stirred at room temperature for 12 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was produced by reverse phase HPLC (0.1% acetic acid, 0% → 100% acetonitrile / water) to obtain the desired product as a white solid (5.7 mg, 30%).
1 1 H NMR (CD 3 OD, 400 MHz): δ7.46 (d, 1H, Ha, JHa -Hb = 7.3 Hz), 7.41-7.27 (m, 3H, Hb, Hc, Hd), 5.38 (ddd, 2H, HBn, J HBn-F = 48Hz, J HBn'-F = 34Hz, J gem = 11Hz ), 4.61 (dd, 1H, Hα 2 , J α2-β2 = 8.2Hz , J α2-β2' = 3.7Hz), 3.88 (d, 2H, Hα1, J gem = 4.1Hz), 3.68-3.52 (m, 2H, Hδ), 2.39-2 .28 (m, 1H, Hβ2), 2.17-1.97 (m, 3H, Hβ2, Hγ2); 13 C NMR (CD 3)OD, 100MHz): δ172.1, 164.8, 128.9, 128.1, 126.6, 125.9, 60.6, 46.4, 40.2, 29.5, 24.5; HRMS 280.14657 (M + H + ).
[0072]
[Synthesis Example 3]
　Compound 3 (gGlu-FMA) of the present invention was synthesized by the following scheme 3.
[0073]
Scheme 3

[0074]
(1) Synthesis of compound 8

[0075]
　(S) -5- (tert-butoxy) -4-((tert-butoxycarbonyl) amino) -5-oxopentanoic acid (1.26 g, 4.17 mmol) was dissolved in dehydrated DMF (15 mL) at 0 ° C. Cooled to. HATU (2.42 g, 6.25 mmol) and DIPEA (2.16 mL, 12.5 mmol) were added and stirred at 0 ° C. for 5 minutes. Subsequently, 2- (tert-butyldimethylsilyl) oxy) methyl) aniline (1.19 g, 5.00 mmol) dissolved in dehydrated DMF (5 mL) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saline solution, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 10% → 30% ethyl acetate / hexane) to obtain the desired product as a yellow liquid (2.18 g, quantitative).
1 1 H NMR (CDCl 3 , 400 MHz): δ8.88 (brs, 1H, -CONH-), 8.15 (d, 1H, J = 8.2 Hz), 7.29 (dd, 1H, J = 8. 2Hz, J = 7.3Hz), 7.10 (d, 1H, J = 6.9Hz), 7.13 (dd, 1H, J = 7.3Hz, J = 6.9Hz), 5.20 (brd) , 1H, -OCONH-, J = 7.8Hz), 4.75 (d, 1H, HBn, J gem = 13Hz ), 4.71 (d, 1H, HBn', J gem = 13Hz ), 4.27 -4.15 (m, 1H), 2.52-2.34 (m, 2H), 2.32-2.20 (m, 1H), 2.07-1.95 (m, 1H), 1 .46 (s, 9H, COO (CH) 3 ) 3 ), 1.42 (s, 9H, NHCOO (CH 3 ) 3 ), 0.90 (s, 9H, Si (CH 3 ) 3 ), 0.07 (s, 6H, Si (CH 3 )) 2 ).
[0076]
(2) Synthesis of compound 9

[0077]
　Compound 8 (2.18 g, 4.17 mmol) was dissolved in dehydrated THF (10 mL), TBAF (ca. 1 mol / L in THF, 10 mL, 10 mmol) was added, and the mixture was stirred at room temperature for 2 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 20% → 80% ethyl acetate / hexane) to obtain the desired product as a white solid (943 mg, 54%).
1 1 H NMR (CD 2 Cl 2 , 400 MHz): δ 8.69 (brs, 1H, -CONH-), 7.91 (d, 1H, J = 7.3 Hz), 7.29 (dd, 1H, J) = 7.8Hz, J = 7.3Hz), 7.23 (d, 1H, J = 7.3Hz), 7.09 (dd, 1H, J = 7.8Hz, J = 6.3Hz), 4. 66 (m, 2H, HBn), 4.22-4.11 (m, 1H), 2.80 (brs, 1H, -CH 2 OH), 2.51-2.35 (m, 2H), 2 .29-2.16 (m, 1H), 1.97-1.85 (m, 1H), 1.44 (s, 9H, COO (CH 3 ) 3 ), 1.40 (s, 9H, NHCOO) (CH 3 ) 3 ).
[0078]
(3) Synthesis of compound 10

[0079]
　Compound 9 (215 mg, 0.527 mmol) was dissolved in dehydrated dichloromethane (10 mL) and cooled to 0 ° C. Deoxo-Flour® (514 μL, 2.64 mmol) was added and stirred at room temperature for 12 hours. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous sodium hydrogen carbonate solution and brine, dried over sodium sulfate, and then concentrated. The residue was produced by silica gel chromatography (34 g silica gel, 20% → 30% ethyl acetate / hexane) to obtain the desired product as a pale yellow liquid (151.8 mg, 70%).
1 1 H NMR (CD 2 Cl 2 , 40 MHz): δ8.04 (brs, 1H, -CONH-), 7.84 (d, 1H, J = 8.2 Hz), 7.37 (dd, 1H, J = 8.2Hz, J = 7.3Hz), 7.33 (d, 1H, J = 6.9Hz), 7.17 (dd, 1H, J = 7.3Hz, J = 6.9Hz), 5.43 (D, 2H, HBn, J HBn-F = 48Hz, J gem = 11Hz), 4.22-4.11 (m, 1H), 2.80 (brs, 1H, -CH 2 OH), 2.51 -2.35 (m, 2H), 2.29-2.16 (m, 1H), 1.97-1.85 (m, 1H), 1.44 (s, 9H, COO (CH 3 ) 3 ), 1.40 (s, 9H, NHCOO (CH) 3 ) 3 ).
[0080]
(4) Synthesis of compound 11

[0081]
　Compound 10 (70.3 mg, 0.171 mmol) was dissolved in ethyl acetate (2 mL) and 4M. Hydrochloric acid / ethyl acetate (2 mL) was added, and the mixture was stirred at room temperature for 12 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was produced by reverse phase HPLC (0.1% acetic acid, 0% → 100% acetonitrile / water) to obtain the desired product as a white solid (28.9 mg, 58%).
1 1 H NMR (CD 2 Cl 2 , 400 MHz): δ7.49 (d, 1H, Ha, JHa -Hb = 7.8 Hz), 7.43 (dd, 1H, Hc, J Hc-Hb = J Hc- Hd = 7.8Hz), 7.36 (dd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.8Hz), 7.29 (d, 1H, Hd, J Hd-Hc = 7.8Hz) ), 5.35 (d, 2H, HBn, J HBn-F = 48Hz), 3.80-3.77 (m, 1H, Hα), 2.67-2.56 (m, 2H, Hγ), 2.20-2.15 (m, 2H, Hβ); 13 C NMR (D 2)O, 100MHz): δ174.7, 173.8, 134.3, 132.0, 130.4, 130.0, 128.1, 127.5, 82.9, 81.3, 54.1, 31 .6,26.3; HRMS 255.11370 (M + H + ).
[0082]
[Example 1]
　Next, an enzymatic reaction was carried out under the following conditions using the three synthesized compounds and the purified enzyme.
Compound final concentration: 100 μM
Enzyme final concentration: 4 nM (β-Gal), 8.5 μg / mL (DPP-IV), 10 U / mL (GGT)
Reaction temperature: 37 ° C.
[0083]
(1) Enzyme reaction
　model using β-galactosidase Model: ACQUITY UPLC (manufactured by Waters)
　Column: Poroshell 120, 4.6 × 100 mm (manufactured by Agient)
　Mobile phase A: Water (0.01 M ammonium formate)
　Mobile phase B : 80% acetonitrile / water (0.01M ammonium formate)
　gradient: A / B: 95/5 → 5/95, 5 minutes
[0084]
(2) Enzyme reaction
　model using DPP-IV and GGT : 1260 Infinity (manufactured by
　Agent ) Column: Poroshell 120, 4.6 × 100 mm (manufactured by Agent)
　Mobile phase A: Water (0.01 M ammonium formate)
　transfer Phase B: 80% acetonitrile / water (0.01M ammonium formate)
　Gradient: A / B: 95/5 → 50/50, 20 minutes
[0085]
　The results are shown in FIG.
　Compound 1-3, β-Gal, DPP-IV , is cleaved respectively by GGT, quinone methide is H 2 was reacted with O 2- hydroxybenzyl alcohol (β-Gal) or 2-aminobenzyl alcohol (DPP-IV, GGT ) Was confirmed as a reaction product.
　For GGT, an experiment in which an inhibitor was added was also conducted, and it was confirmed that the enzymatic reaction of the model compound was inhibited by adding GGsTop® (registered trademark), which is a GGT inhibitor, at a concentration of 100 μM.
[0086]
[Example 2] In
vitro drug efficacy test using high-expression / low-expressing enzyme cells
　Next, when compounds 1 to 3 were administered to high-expressing / low-expressing enzyme cells, they functioned as prodrugs for cell survival. We verified whether it was possible to change the rate. The drug efficacy test uses the CCK-8 assay (a method of quantifying the dehydrogenase activity in living cells in the form of reduction from colorless WST-8 to orange formazan), which is a general colorimetric quantification method. This was done by quantifying the number of living cells. The evaluation method of the CCK-8 assay is described below.
[0087]
(1) Cultured cells used
　β-galactosidase activation prodrugs were evaluated by HEK / lacZ cells (human kidney cell-derived cells, β-Gal high expression) and HEK293 cells (human kidney cell-derived cells, β-Gal low expression). ) Was used. Evaluation of DPP-IV activated prodrug was performed using H226 cells (human squamous cell lung cancer cells, high expression of DPP-IV) and H460 cells (human non-small cell epithelial lung cancer cells, low expression of DPP-IV). I went. Evaluation of the GGT-activated prodrug was performed using SHIN3 cells (human ovarian cancer cells, high GGT expression) and H226 cells (human lung squamous cell carcinoma cells, low GGT expression).
[0088]
(2) Evaluation method
　Various cells were seeded on a 96-well plate (cell density: 1.0 × 10 4 / well) and incubated overnight. The medium was replaced with fresh medium and the synthesized derivative was added (final concentration 1-50 μM, 0.5% DMSO, n = 3). After further culturing the cells for 24 hours, Cell counting Kit-8 (10 μL / well, manufactured by Promega) was added, and after 2.5 hours, the absorbance at 450 nm was measured with a plate reader to quantify the number of surviving cells. It was.
[0089]
　The results of the CCK-8 assay are shown in Figures 5-7.
　First, compound 1 (β-Gal-FMP) for β-galactosidase was administered to high-expressing cells (HEK / lacZ) and low-expressing cells (HEK293), and the survival rate was calculated after 24 hours. As a result, the survival rate of both cells did not decrease at all even when administered at a high concentration of 50 μM, and no significant difference was observed (Fig. 5). In contrast to the results with β-galactosidase, when the aminopeptidases DPP-IV and GGT compounds 2 and 3 (GP-FMA, gGlu-FMA) were used, the enzyme was highly expressed / low expressed between cells. Changes in cell viability were observed in (Figs. 6 and 7). From this result, (i) the difference in localization between DPP-IV and GGT (cell membrane) and β-galactosidase (cytoplasm), and (ii) the difference in quinone methide species (DPP-IV and GGT release azaquinone methide) are important. It was suggested, but it is unknown at this stage. In particular, GGT prodrugs have high cell type selectivity, and the survival rate was completely restored by the GGT inhibitor GGsTop (registered trademark). Therefore, it was expected that most of the drug efficacy was dependent on GGT activity. ..
[0090]
[Example 3]
Examination of observation of cell death under co-culture conditions Since
　co-culture became possible by changing the cell type and unifying the medium to RPMI-1640, cells under co-culture conditions were subsequently subjected to. We aimed to establish a death observation and quantitative method. Based on the results so far, we decided to conduct these studies using GGT compound 3 (gGlu-FMA). First, as an initial study, is it possible to visually observe cell death by fluorescence observation using one type of cell? It was examined (Fig. 8). The experiment was performed using the following protocol 1.
[0091]
(Protocol 1)

[0092]
　FIG. 8 shows fluorescence imaging over time of SHIN3 cells (top) and H226 cells (bottom) using Compound 3 and EthD-1 (dead cell staining, Ex / Em = 525 nm / 511-564 nm).
Lens 63x / 1.4 Oil.
Scan modes: xyzt, x = 512, y = 512, z = 4, t = 24. Scale bar, 50 μm.
[0093]
　From the results shown in FIG. 8, it was confirmed that SHIN3 cells, which are GGT high-expressing cell lines, caused more cell death, and therefore, evaluation was carried out in a co-culture system. The experiment was performed using the following protocol 2.
[0094]
(Protocol 2)

[0095]
　The results are shown in FIG.
　FIG. 9 shows time-lapse fluorescence imaging of co-cultured cells using Compound 3 and EthD-1 (dead cell staining, Ex / Em = 525 nm / 511-564 nm) (top).
Confocal imaging of cells in the other three fields of view after 24-hour imaging (bottom).
Lens 63x / 1.4 Oil.
Scan modes: xyzt, x = 512, y = 512, z = 4, t = 24. Scale bar, 50 μm.
[0096]
　As can be seen from the results of the 24-hour time-lapse imaging shown in the upper part of FIG. 9, it was clarified that the compound 3 for GGT (gGlu-FMA) selectively injured highly expressed cells even in the co-culture system. In addition, since phototoxicity was observed when 10 sections x 24 imagings were performed in the previous studies (many cells in the field of view die), this time the imaging was 4 sections x 24 times, but phototoxicity was observed. There was concern about the impact of. Therefore, when three different fields of view were imaged after the completion of 24-hour imaging (lower part of FIG. 9), a similar tendency was observed in which dead cells were present in the gaps between low-expressing cells (green), so that compound 3 worked well. It was judged that the killing was achieved by working.
[0097]
[Example 4]
Evaluation of drug efficacy by Flow Cytometry
　It was shown that cell death under co-culture conditions can be observed in real time by imaging using a fluorescent dye. On the other hand, since it is not possible to quantitatively discuss how much the survival rate of high-expression / low-expression cells has changed, it was examined whether these can be evaluated using flow cytometry (Fig. 10). The experiment was carried out according to Protocol 3 below.
[0098]
(Protocol 3)

[0099]
　FIG. 10 shows the results of flow cytometric analysis of cell proliferation in SHIN3 cells (green) and H226 cells (unstained) using Compound 3. (Top) 25 μM model compound, (Center) 25 μM model compound + 100 μM GGsTop, (Bottom) 0.25% DMSO control. The analysis was performed after 24 hours of incubation.
[0100]
　From the results shown in FIG. 10, it is flow cytometry that cell death is induced only in the GGT prodrug-administered group (death staining with EthD-1 is observed) and causes SHIN3 cell-selective cell death, which is a GGT highly expressing cell. Also became clear.
[0101]
Synthesis of benzyl-leaving group-converted derivative
　Next, the synthesis of a benzyl-converted derivative targeting GGT was examined. Evans of pK a table; a (DH Ripin DA Evans, see Http://Evans.Rc.Fas.Harvard.Edu/pdf/evans_pKa_table.Pdf) with reference acyl-based leaving group such as shown in FIG. 11 It was verified whether the derivative having the above can be synthesized.
[0102]
[Synthesis Example 4]
　The compound of the present invention was synthesized by the following scheme 4.
[0103]
Scheme 4

[0104]
(1) Synthesis of compound 11

[0105]
　(S) -5- (allyloxy) -4-(((allyloxy) carbonyl) amino) -5-oxopentanoic acid (930 mg, 3.45 mmol) was dissolved in dehydrated DMF (17 mL) and cooled to 0 ° C. HATU (1.96 g, 5.14 mmol) and DIPEA (1.75 mL, 10.3 mmol) were added and stirred at 0 ° C. for 5 minutes. Subsequently, 2- (tert-butyldimethylsilyl) oxy) methyl) aniline (1.22 g, 5.14 mmol) dissolved in dehydrated DMF (5 mL) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 50% → 60% ethyl acetate / hexane) to obtain the desired yellow liquid (848 mg, 86%).
1 1 H NMR (CDCl 3), 400MHz): δ8.91 (brs, 1H, -CONH-), 8.16 (d, 1H, J = 7.8Hz), 7.29 (dd, 1H, J = 7.8Hz, J = 7. 3Hz), 7.08 (d, 1H, J = 7.3Hz), 7.02 (dd, 1H, J = 7.3Hz, J = 7.3Hz, 5.95-5.82 (m, 2H) , 5.61 (brd, 1H, -OCONH-, J = 7.8Hz), 5.32 (d, 1H, J = 17Hz), 5.28 (d, 1H, J = 17Hz), 5.24 ( d, 1H, J = 11Hz), 5.18 (d, 1H, J = 11Hz), 4.73 (s, 2H, HBn), 4.64 (d, 1H, J = 5.5Hz), 4. 56-4.52 (m, 2H), 4.47-4.37 (m, 1H), 2.56-2.39 (m, 2H), 2.38-2.27 (m, 1H), 2.18-2.05 (m, 1H), 0.90 (s, 9H, Si (CH 3 ) 3 ), 0.08 (s, 6H, Si (CH 3 ) 2 ).
[0106]
(2) Synthesis of compound 12

[0107]
　Compound 11 (1.29 g, 2.63 mmol) is dissolved in dehydrated THF (20 mL) and TBAF (ca. 1 mol / L in THF, 7.89 mL, 7.89 mmol) and acetic acid (304 μL, 5.26 mmol) are added. , Stirred at room temperature for 3 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 50% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (848 mg, 86%).
1 H NMR (CDCl 3 , 400 MHz): δ8.73 (brs, 1H, -CONH-), 7.97 (d, 1H, J = 8.2 Hz), 7.30 (dd, 1H, J = 8. 2Hz, J = 7.8Hz), 7.18 (d, 1H, J = 7.3Hz), 7.07 (dd, 1H, J = 7.8Hz, J = 7.3Hz), 5.95-5 .79 (m, 2H, 5.67 (brd, 1H, -OCONH-, J = 7.8Hz), 5.32 (d, 1H, J = 17Hz), 5.27 (d, 1H, J = 17Hz) ), 5.24 (d, 1H, J = 10Hz), 5.19 (d, 1H, J = 10Hz), 4.74-4.61 (m, 2H, HBn), 4.63 (d, 1H) , J = 6.0Hz), 4.57-4.45 (m, 2H), 4.45-4.35 (m, 1H), 2.97 (brs, 1H, CH 2 OH), 2.55 -2.40 (m, 2H), 2.39-2.26 (m, 1H), 2.12-1.96 (m, 1H).
[0108]
(3) Synthesis of compound 13

[0109]
　Compound 12 (29.6 mg, 0.0786 mmol) and triethylamine (110 μL, 0.786 mmol) were dissolved in dehydrated dichloromethane (1 mL), acetic anhydride (15 μL, 0.157 mmol) was added, and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 30% → 40% ethyl acetate / hexane) to obtain the desired product as a white solid (31.8 mg, 97%).
1 1 H NMR (CDCl 3 , 400 MHz): δ8.86 (brs, 1H, -CONH-), 7.96 (d, 1H, J = 7.8 Hz), 7.35 (dd, 1H, J = 7. 8Hz, J = 7.3Hz), 7.34 (d, 1H, J = 7.3Hz), 7.13 (dd, 1H, J = 7.8Hz, J = 7.3Hz), 5.95-5 .81 (m, 2H), 5.68 (brd, 1H, -OCONH-, J = 7.3Hz), 5.32 (d, 1H, J = 17Hz), 5.27 (d, 1H, J = 17Hz), 5.24 (d, 1H, J = 11Hz), 5.18 (d, 1H, J = 11Hz), 5.13 (d, 1H, HBn, J gem = 12Hz ), 5.08 (d) , 1H, HBn', J gem = 12Hz ), 4.63 (d, 1H, J = 5.5Hz), 4.58-4.48 (m, 2H), 4.48-4.39 (m, 1H), 2.62-2.46 (m, 2H), 2.41-2.27 (m, 1H), 2.19-2.05 (m, 1H), 2.08 (s, 3H, OCOCH 3 ).
[0110]
(4) Synthesis of compound 14

[0111]
　Compound 13 (31.7 mg, 0.0758 mmol) and phenylsilane (234 μL, 1.89 mmol) are dissolved in dehydrated dichloromethane (20 mL), tetrakis (triphenylphosphine) palladium (21.8 mg, 0.0189 mmol) is added. The mixture was stirred at room temperature for 2 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (0% → 100% acetonitrile / water) to obtain the desired product as a white solid (8.0 mg, 36%).
1 H NMR (CD 3 OD, 400 MHz): δ7.40 (d, 1H, Ha, JHa -Hb = 7.3 Hz), 7.38 (d, 1H, Hd, JHd -Hc = 7.3 Hz) , 7.32 (dd, 1H, Hc, J Hc-Hb = J Hc-Hd = 7.3 Hz), 7.24 (dd, 1 H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz) , 5.09 (s, 2H, HBn), 3.63 (t, 1H, Hα, J Hα-Hβ = 6.2Hz), 2.65 (t, 2H, Hγ, J Hγ-Hα = J Hγ- Hβ = 7.3Hz), 2.18 (td, 2H, Hβ, J Hβ-Hγ= 7.3 Hz, J Hβ-Hα = 6.2 Hz), 2.05 (s, 3H, CH 3 COO-); 13 C NMR (CD 3 OD, 100 MHz): δ 172.3, 172.4, 171. 4,135.3,131.1,129.4,128.6,126.3,126,1,62.6,54.3,32.0,26.5,19.5; HRMS 317.1124 (M + Na + ).
[0112]
(5) Synthesis of compound 15

[0113]
　Compound 12 (35.1 mg, 0.0933 mmol), DIPEA (96 μL, 0.560 mmol) and DMAP (3.5 mg) were dissolved in dehydrated dichloromethane (1 mL) to add 4-methoxybenzoyl chloride (19 mg, 0.112 mmol). In addition, it was stirred at room temperature for 4 hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a white solid (43.2 mg, 91%).
1 H NMR (CDCl 3 , 400 MHz): δ9.20 (brs, 1H, -CONH-), 8.00 (d, 2H, J = 9.2 Hz), 8.00 (d, 1H, J = 7. 3Hz), 7.43 (d, 1H, J = 7.8Hz), 7.37 (dd, 1H, J = 7.8Hz, J = 7.3Hz), 7.14 (dd, 1H, J = 7) .3Hz, J = 7.3Hz), 6.90 (d, 2H, J = 9.2Hz), 5.95-5.82 (m, 2H), 5.71 (brd, 1H, -OCONH-, J = 6.9Hz), 5.35 (d, 1H, HBn, J gem = 12Hz ), 5.34 (d, 1H, J = 19Hz), 5.31 (d, 1H, HBn', J gem = 12Hz), 5.27 (d, 1H, J = 19Hz), 5.23 (d, 1H, J = 11Hz), 5.17 (d, 1H, J = 11Hz), 4.63 (d, 1H, J = 6.0Hz), 4.58-4.50 (m, 2H), 4.50-4.41 (m, 1H), 3.85 (s, 3H, ArOCH) 3 ), 2.67-2.52 (m, 2H), 2.42-2.30 (m, 1H), 2.24-2.07 (m, 1H).
[0114]
(6) Synthesis of compound 16

[0115]
　Compound 15 (26.4 mg, 0.0517 mmol) and phenylsilane (160 μL, 1.29 mmol) are dissolved in dehydrated dichloromethane (2 mL), tetrakis (triphenylphosphine) palladium (14.9 mg, 0.0129 mmol) is added. The mixture was stirred at room temperature for 3 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (7.8 mg, 39%).
1 1 H NMR (CD 3 OD, 400 MHz): δ7.96 (d, 2H, He, Hf, J He-Hh = J Hf-Hg = 9.2 Hz), 7.50 (d, 1H, Ha, J Ha) -Hb = 7.3Hz), 7.41 (d, 1H, Hd, J Hd-Hc = 6.9Hz), 7.34 (dd, 1H, Hc, J Hc-Hb = 7.8Hz, J Hc- Hd = 6.9Hz), 7.26 (dd, 1H, Hb, J Hb-Hc = 7.8Hz, J Hb-Ha = 7.3Hz), 6.97 (d, 2H, Hg, Hh, J Hg) -Hf = J Hh-He= 9.2Hz), 5.32 (s, 2H, HBn), 3.83 (s, 3H, -OCH 3 ), 3.62 (t, 1H, Hα, J Hα-Hβ = 6.0Hz), 2.66 (t, 2H, Hγ, J Hγ-Hα = J Hγ-Hβ = 7.3Hz), 2.17 (td, 2H, Hβ, J Hβ-Hγ = 7.3Hz, J Hβ-Hα = 6 .0Hz); 13 C NMR (CD 3 OD, 100MHz): δ172.9, 166.5, 164.0, 135.3, 131.4, 131.3, 129.4, 128.6, 126.4 , 126.0, 122.0, 113.6, 62.8, 54.7, 54.4, 32.0, 26.5; HRMS 409.13271 (M + Na + ).
[0116]
(7) Synthesis of compound 17

[0117]
　Compound 12 (29.7 mg, 0.0789 mmol), DIPEA (82 μL, 0.473 mmol) and DMAP (3.0 mg) were dissolved in dehydrated dichloromethane (1 mL) to add 4-chlorobenzoyl chloride (13 μL, 0.0947 mmol). In addition, it was stirred at room temperature for 4 hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a white solid (33.8 mg, 83%).
1 H NMR (CDCl 3 , 400 MHz): δ9.02 (brs, 1H, -CONH-), 7.97 (d, 2H, J = 8.7 Hz), 7.95 (d, 1H, J = 7. 3Hz), 7.43 (d, 1H, J = 7.8Hz), 7.43 (d, 2H, J = 8.7Hz), 7.37 (dd, 1H, J = 7.8Hz, J = 7) .3Hz), 7.16 (dd, 1H, J = 7.3Hz, J = 7.3Hz), 5.95-5.79 (m, 2H), 5.69 (brd, 1H, -OCONH-, J = 7.8Hz), 5.38 (d, 1H, HBn, J gem = 12Hz ), 5.35 (d, 1H, J = 17Hz), 5.33 (d, 1H, HBn', J gem)= 12Hz), 5.26 (d, 1H, J = 17Hz), 5.23 (d, 1H, J = 9.6Hz), 5.16 (d, 1H, J = 9.6Hz), 4.63 (D, 1H, J = 6.0Hz), 4.56-4.48 (m, 2H), 4.49-4.40 (m, 1H), 2.66-2.51 (m, 2H) , 2.43-2.30 (m, 1H), 2.20-2.05 (m, 1H).
[0118]
(8) Synthesis of compound 18

[0119]
　Compound 17 (33.7 mg, 0.0654 mmol) and phenylsilane (203 μL, 1.63 mmol) are dissolved in dehydrated dichloromethane (2 mL), tetrakis (triphenylphosphine) palladium (18.9 mg, 0.0163 mmol) is added. The mixture was stirred at room temperature for 3 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (3.8 mg, 15%).
1 1 H NMR (CD 3 OD, 400 MHz): δ7.99 (d, 2H, He, Hf, J He-Hh = J Hf-Hg = 7.8 Hz), 7.51 (d, 1H, Ha, J Ha) -Hb = 7.8Hz), 7.48 (d, 2H, Hg, Hh, J Hg-Hf = J Hh-He = 7.8Hz), 7.40 (d, 1H, Hd, J Hd-Hc = 7.8Hz), 7.34 (dd, 1H, Hc, J Hc-Hd = 7.8Hz, J Hc-Hb = 7.3Hz), 7.27 (dd, 1H, Hb, J Hb-Ha = 7) .8Hz, J Hb-Hc= 7.3 Hz), 5.36 (s, 2H, HBn), 3.62 (t, 1H, Hα, J Hα-Hβ = 6.0 Hz), 2.66 (t, 2H, Hγ, J Hγ) -Hα = J Hγ-Hβ = 7.3 Hz), 2.16 (td, 2H, Hβ, J Hβ-Hγ = 7.3 Hz , J Hβ-Hα = 6.0 Hz); 13 C NMR (CD 3 OD) , 100 MHz): δ172.9, 172.5, 165.6, 139.4, 135.4, 131.1, 130.9, 129.5, 128.8, 128.6, 126.5, 126. 2,63,54.3,32.0,26.5; HRMS 413.80817 (M + Na + ).
[0120]
(9) Synthesis of compound 19

[0121]
　Compound 12 (31.5 mg, 0.0837 mmol), DIPEA (87 μL, 0.502 mmol) and DMAP (3.2 mg) were dissolved in dehydrated dichloromethane (1 mL) and 4-nitrobenzoyl chloride (18.6 mg, 0.100 mmol). ) Was added, and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 30% → 40% ethyl acetate / hexane) to obtain the desired product as a white solid (40.5 mg, 92%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.86 (brs, 1H, -CONH-), 8.27 (d, 2H, J = 8.7 Hz), 8.21 (d, 2H, J = 8.7 Hz), 7.91 (d , 1H, J = 7.8 Hz), 7.45 (d, 1H, J = 7.3 Hz), 7.38 (dd, 1H, J = 7.8 Hz, J = 7.3 Hz), 7.18 (dd, 1H, J = 7.3 Hz, J = 7.3 Hz), 5.95-5.78 (m, 2H), 5.67 (brd, 1H, -OCONH-, J = 7.8 Hz), 5.44 (d, 1H, HBn, J gem = 12 Hz), 5.39 (d, 1H) , HBn', J gem = 12 Hz), 5.31 (d, 1H, J = 17 Hz), 5.25 (d, 1H, J = 17 Hz), 5.23 (d, 1H, J = 11 Hz), 5.16 (d, 1H, J = 11 Hz), 4.63 (d, 1H, J = 6.0 Hz), 4.56-4.39 (m, 3H), 2.65-2.51 (m, 2H), 2.43-2.29 (m, 1H), 2.20-2.03 (m, 1H) ..
[0122]
(10) Synthesis of compound 20 (N 5- (2-(((4-nitrobenzoyl) oxy) methyl) phenyl) -L-glutamine)

[0123]
　Compound 19 (40.5 mg, 0.0771 mmol) and phenylsilane (239 μL, 1.92 mmol) are dissolved in dehydrated dichloromethane (2 mL), tetrakis (triphenylphosphine) palladium (22.3 mg, 0.0192 mmol) is added. The mixture was stirred at room temperature for 2 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (6.3 mg, 20%).
1 H NMR (CD 3 OD, 400 MHz): δ 8.31 (d, 2H, He, Hf, J He-Hh = J Hf-Hg = 9.2 Hz), 8.22 (d, 2H, Hg, Hh, J Hg- Hf = J Hh-He = 9.2 Hz), 7.53 (d, 1H, Ha, J Ha-Hb = 7.3 Hz), 7.40-7.34 (m, 2H, Hc, Hd), 7.29 (ddd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz, J Hb-Hd = 1.8 Hz), 5.41 (s, 2H, HBn), 3.62 (t, 1H, Hα, J Hα-Hβ= 6.0 Hz), 2.66 (t, 2H, Hγ, J Hγ-Hα = J Hγ-Hβ = 7.3 Hz), 2.16 (td, 2H, Hβ, J Hβ-Hγ = 7.3 Hz, J Hβ-Hα = 6.0 Hz ); 13 C NMR (CD 3 OD, 100 MHz): δ 173.0, 172.5, 164.7, 150.8, 135.5, 130.9, 130.5, 129.7, 129.0, 128.0, 127.6, 126.5, 125.6, 124.6, 123.3, 63.9, 54.3, 32.0 , 26.6; HRMS 413.11126 (M + Na + ).
[0124]
(11) Synthesis of compound 21

[0125]
　Compound 12 (30.8 mg, 0.0818 mmol), DIPEA (85 μL, 0.491 mmol) and DMAP (3.1 mg) were dissolved in dehydrated dichloromethane (1 mL) and 2-chlorobenzoyl chloride (12.4 μL, 0.0978 mmol). ) Was added, and the mixture was stirred at room temperature for one and a half hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 20% → 30% ethyl acetate / hexane) to obtain the desired product as a white solid (27.3 mg, 63%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.74 (brs, 1H, -CONH-), 7.94 (d, 1H, J = 8.2 Hz), 7.83 (d, 1H, J = 7.6 Hz), 7.47-7.41 (m, 3H), 7.37 (ddd, 1H, J = 7.8 Hz, J = 7.8 Hz, J = 1.4 Hz), 7.30 (ddd, 1H, J = 7.3 Hz, J = 6.9 Hz, J = 1.8 Hz), 7.16 (dd, 1H, J = 7.3 Hz, J = 7.3 Hz), 5.95-5.80 (m, 2H), 5.66 (brd, 1H, -OCONH-, J = 7.8 Hz), 5.40 (d, 1H, HBn, J gem = 12 Hz), 5.36 (d, 1H, HBn', J gem = 12 Hz), 5.31 (d, 1H, J = 17 Hz), 5.26 (d, 1H, J = 17 Hz), 5.22 (d, 1H, J = 11 Hz), 5.16 (d, 1H, J = 11 Hz), 4.63 (d, 1H, J = 6.0 Hz), 4.58-4.49 (m, 2H), 4.49-4.39 (m, 1H), 2.67-2.50 (m, 2H), 2.44-2.30 (m, 1H) , 2.18-2.04 (m, 1H).
[0126]
(12) Synthesis of compound 22

[0127]
　Compound 21 (27.3 mg, 0.0530 mmol) and phenylsilane (164 μL, 1.32 mmol) are dissolved in dehydrated dichloromethane (2 mL), tetrakis (triphenylphosphine) palladium (15.3 mg, 0.0132 mmol) is added. The mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (11.1 mg, 54%).
1 H NMR (CD 3 OD, 400 MHz): δ 7.81 (d, 1H, He, J He-Hf = 7.3 Hz), 7.53 (d, 1H, Ha, J Ha-Hb = 7.3 Hz), 7.49-7.48 (m, 2H, Hc, Hd), 7.41-7.34 (m, 3H, Hb, Hg, Hh), 7.29 (dd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz), 5.37 (s) , 2H, HBn), 3.63 (t, 1H, Hα, J Hα-Hβ = 6.0 Hz), 2.67 (t, 2H, Hγ, J Hγ-Hα = J Hγ-Hβ = 7.3 Hz), 2.18 (td, 2H) , Hβ, J Hβ-Hγ = 7.3 Hz, J Hβ-Hα = 6.0 Hz); 13 C NMR (CD 3 OD, 100 MHz): δ 172.9, 172.7, 135.4, 133.1, 132.7, 131.1, 130.8, 130.7, 130.1, 129.6, 128.8 , 126.7, 126.4, 126.1, 63.6, 54.4, 32.0, 26.6; HRMS 413.08882 (M + Na + ).
[0128]
(13) Synthesis of compound 23

[0129]
　Compound 12 (32.2 mg, 0.0855 mmol), DIPEA (89 μL, 0.513 mmol) and DMAP (3.2 mg) were dissolved in dehydrated dichloromethane (1 mL) to give 3-nitrobenzoyl chloride (14 μL, 0.103 mmol). In addition, the mixture was stirred at room temperature for 3 hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 30% → 50% ethyl acetate / hexane) to obtain the desired product as a white solid (37.8 mg, 84%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.90 (brs, 1H, -CONH-), 8.85 (s, 1H), 8.41 (dd, 1H, J = 8.0 Hz, J = 2.3 Hz), 8.36 (d , 1H, J = 7.8 Hz), 7.91 (d, 1H, J = 7.8 Hz), 7.65 (dd, 1H, J = 8.2 Hz, J = 7.8 Hz), 7.47 (d, 1H, J = 7.8 Hz), 7.38 (dd, 1H, J = 7.8 Hz, J = 7.3 Hz), 7.18 (dd, 1H, J = 7.3 Hz, J = 7.3 Hz), 5.95-5.78 (m, 2H), 5.68 (brd, 1H,- OCONH-, J = 7.8 Hz), 5.45 (d, 1H, HBn, J gem = 12 Hz), 5.40 (d, 1H, HBn', J gem = 12 Hz), 5.31 (d, 1H, J = 17 Hz), 5.25 (d, 1H, J = 17 Hz), 5.23 (d, 1H, J = 11 Hz), 5.15 (d, 1H, J = 11 Hz), 4.63 (d, 1H, J = 6.0 Hz), 4.56-4.40 (m, 2H), 2.67-2.51 (m, 2H), 2.44-2.30 (m, 1H), 2.22-2.05 (m, 1H) ..
[0130]
(14) Synthesis of compound 24

[0131]
　Compound 23 (36.9 mg, 0.0702 mmol) and phenylsilane (217 μL, 1.76 mmol) were dissolved in dehydrated DMF (2 mL), tetrakis (triphenylphosphine) palladium (20.3 mg, 0.0176 mmol) was added. The mixture was stirred at room temperature for 2 hours. After confirming the completion of the reaction, the reaction solution was directly subjected to purification by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (10.0 mg, 35%).
1 H NMR (CD 3 OD, 400 MHz): δ 8.78 (s, 1H, He), 8.45 (d, 1H, Hh, J Hh-Hg = 8.2 Hz), 8.39 (d, 1H, Hf, J Hf- Hg = 7.6 Hz), 7.75 (dd, 1H, Hg, J Hg-Hh = 8.2 Hz, J Hg-Hf = 7.6 Hz), 7.54 (d, 1H, Ha, J Ha-Hb = 7.3 Hz), 7.43- 7.35 (m, 2H, Hc, Hd), 7.41-7.34 (m, 3H, Hb, Hg, Hh), 7.30 (dd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz), 5.43 s, 2H, HBn), 3.60 (t, 1H, Hα, J Hα-Hβ = 6.0 Hz), 2.66 (t, 2H, Hγ, J Hγ-Hα = J Hγ-Hβ = 7.3 Hz), 2.16 (td, 2H, Hβ, J Hβ-Hγ = 7.3 Hz, J Hβ-Hα = 6.0 Hz); 13 C NMR (CD 3 OD, 100 MHz): no data; HRMS 424.11063 (M + Na + ).
[0132]
(15) Synthesis of compound 25

[0133]
　Compound 12 (37.3 mg, 0.0991 mmol), DIPEA (103 μL, 0.595 mmol) and DMAP (3.7 mg) were dissolved in dehydrated dichloromethane (1 mL) to give 2-nitrobenzoyl chloride (16 μL, 0.119 mmol). In addition, the mixture was stirred at room temperature for one and a half hours. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous ammonium chloride solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 30% → 50% ethyl acetate / hexane) to obtain the desired product as a white solid (40.5 mg, 78%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.23 ​​(brs, 1H, -CONH-), 7.91 (d, 1H, J = 7.8 Hz), 7.74 (d, 1H, J = 8.0 Hz), 7.71-7.60 (m, 2H), 7.38 (dd, 1H, J = 7.8 Hz, J = 7.3 Hz), 7.37 (d, 1H, J = 7.3 Hz), 7.16 (dd, 1H, J = 7.3 Hz, J = 7.3 Hz ), 5.95-5.80 (m, 2H), 5.69 (brd, 1H, -OCONH-, J = 7.8 Hz), 5.38 (d, 1H, HBn, J gem = 12 Hz), 5.31 (d, 1H, HBn' , J gem = 12 Hz), 5.31 (d, 1H, J = 17 Hz), 5.26 (d, 1H, J = 17 Hz), 5.22 (d, 1H, J = 11 Hz), 5.15 (d, 1H, J = 11 Hz), 4.63 (d, 1H, J = 6.0 Hz), 4.58-4.40 (m, 2H), 2.68-2.53 (m, 2H), 2.42-2.30 (m, 1H), 2.18-2.20 (m, 1H) ..
[0134]
(16) Synthesis of compound 26

[0135]
　Compound 25 (40.2 mg, 0.0765 mmol) and phenylsilane (236 μL, 1.91 mmol) are dissolved in dehydrated dichloromethane (2 mL), tetrakis (triphenylphosphine) palladium (22.1 mg, 0.0191 mmol) is added. The mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, the reaction solution was directly subjected to purification by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (7.2 mg, 23%).
1 H NMR (CD 3 OD, 400 MHz): δ 7.94 (d, 1H, He, J He-Hf = 7.8 Hz), 7.82-7.69 (m, 3H, Hf, Hg, Hh), 7.46 (d, 1H) , Ha, J Ha-Hb = 7.3 Hz), 7.42-7.33 (m, 2H, Hc, Hd), 7.27 (dd, 1H, Hb, J Hb-Ha = J Hb-Hc = 7.3 Hz), 5.34 (s) , 2H, HBn), 3.63 (t, 1H, Hα, J Hα-Hβ = 5.3 Hz), 2.69 (t, 2H, Hγ, J Hγ-Hα = J Hγ-Hβ = 6.6 Hz), 2.18 (td, 2H) , Hβ, J Hβ-Hγ = 6.6 Hz, J Hβ-Hα = 5.3 Hz); 13 C NMR (CD 3 OD, 100 MHz): 173.0, 172.6, 165.3, 135.5, 133.0, 132.3, 130.2, 129.8, 129.0, 126.9 , 126.4, 126.1, 123.8, 64.4, 54.4, 32.0, 26.5; HRMS 424.11207 (M + Na + ).
[0136]
[Example 5]
(1) Confirmation of enzyme recognition ability using a benzyl-leaving group-converting derivative
　Next, an enzyme reaction was carried out using the synthesized benzyl-leaving group-converting derivative and a purified enzyme under the following conditions. ..
Compound final concentration: 100 μM
Enzyme final concentration: 10 U / mL (GGT)
Reaction temperature: 37 ° C
[0137]
Enzyme reaction
　model using GGT : 1260 Infinity (manufactured by
　Agent ) Column: Poroshell 120, 4.6 × 100 mm (manufactured by Agent)
　Mobile phase A: Water (0.01 M ammonium formate)
　Mobile phase B: 80% acetonitrile / Water (0.01M ammonium formate)
　Gradient: A / B: 95/5 → 50/50, 20 minutes
[0138]
　The results are shown in FIG. FIG. 12 shows a chromatogram 0 hour and 1 hour after the mass peak of the raw material and a chromatogram 0 hour and 1 hour after the mass peak of the enzyme reaction product, respectively.
　From the results of FIG. 12, it was confirmed that the derivative group having an acyl leaving group releases azaquinone methide at a different rate depending on the purified GGT.
[0139]
(2) In vitro drug efficacy test using benzyl leaving group-converting derivative (CCK-8 assay)
　Next, the above-mentioned CCK-8 assay was performed on the benzyl leaving leaving group-converting derivative. The results are shown in FIG.
　As shown in FIG. 13, at a concentration of 25 μM in which the model compound having a leaving group of fluorine showed a medicinal effect, none of the derivatives having an acyl leaving group showed antitumor activity.
[0140]
Synthesis of Substituent Converting Derivative on Benzene Ring
　Further, the following four compounds in which a substituent was introduced at the 4th and 5th positions on the benzene ring as a GGT prodrug derivative were synthesized. By introducing a methyl group (electron donating group) and a methyl ester group (electron attracting group), the electron density of the benzene ring changes and the reactivity of the released azaquinone methide changes, which also affects cell death. is expected.
[0141]

[0142]
[Synthesis Example 5] A
　4-position substituted product was synthesized according to the following synthesis scheme 5.
[0143]
(Scheme 5)

[0144]
(1) Synthesis of compound 27

[0145]
　(S) -5- (tert-butoxy) -4-((tert-butoxycarbonyl) amino) -5-oxopentanoic acid (347 mg, 1.14 mmol) was dissolved in dehydrated DMF (10 mL) and cooled to 0 ° C. did. HATU (408 mg, 1.72 mmol) and DIPEA (743 μL, 3.43 mmol) were added, and the mixture was stirred at 0 ° C. for 5 minutes. Subsequently, methyl-4-amino-3-(((tert-butyldimethylsilyl) oxy) methyl) benzoate (405 mg, 1.37 mmol) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 10% → 20% ethyl acetate / hexane) to obtain the desired product as a mixture (197 mg). This mixture was dissolved in dehydrated THF (5 mL), TBAF (ca. 1 mol / L in THF, 1 mL, 1.00 mmol) and acetic acid (46 μL, 0.678 mmol) were added, and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, saturated aqueous ammonium chloride solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 40% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (39.9 mg, 2 steps 6%).
1 1 H NMR (CDCl 3), 400 MHz): δ 9.16 (brs, 1H, -CONH-), 8.24 (d, 1H, J = 8.2 Hz), 7.97 (d, 1H, J = 8.2 Hz), 7.85 (s, 1H), 5.31 ( brd, 1H, -OCONH-, J = 7.3 Hz), 4.84-4.64 (m, 2H, HBn), 4.27-4.15 (m, 1H), 3.88 (s, 3H, ArCOOCH 3 ), 3.26-3.18 (m, 1H, ArCOOCH 3 ) 1H, CH 2 OH), 2.36-2.22 (m, 1H), 2.03-1.88 (m, 1H), 1.45 (s, 9H, COO (CH 3 ) 3 ), 1.41 (s, 9H, NHCOO (CH 3 )) 3 ).
[0146]
(2) Synthesis of compound 28

[0147]
　Compound 27 (38.6 mg, 0.0824 mmol) was dissolved in dehydrated dichloromethane (2 mL) and cooled to 0 ° C. DAST (54 μL, 0.412 mmol) was added and the mixture was stirred at room temperature for 1 hour. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous sodium hydrogen carbonate solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 20% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (25.9 mg, 67%).
1 1 H NMR (CDCl 3 , 400 MHz): δ 8.48 (brs, 1H, -CONH-), 8.23 ​​(d, 1H, J = 8.2 Hz), 8.07 (d, 1H, J = 8.2 Hz), 7.99 (s , 1H), 5.70-5.38 (m, 2H, HBn), 5.30 (brs, 1H, -OCONH-), 4.28-4.18 (m, 1H), 3.91 (s, 3H, ArCOOCH 3 ), 2.62-2.42 (m) , 2H), 2.40-2.26 (m, 1H), 2.20-1.82 (m, 1H), 1.46 (s, 9H, COO (CH 3 ) 3 ), 1.44 (s, 9H, NHCOO (CH 3 ) 3 ).
[0148]
(3) Synthesis of compound 29

[0149]
　Compound 28 (29.5 mg, 0.0553 mmol) was added to 4M. It was dissolved in hydrochloric acid / ethyl acetate (2 mL) and stirred at room temperature for 12 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (0% → 100% acetonitrile / water) to obtain the desired product as a white solid (11 mg, 64%).
1 1 H NMR (CD 3 OD, 400 MHz): δ 8.09 (s, 1H, Ha), 7.99 (d, 1H, Hb, J Hb-Hc = 8.7 Hz), 7.66 (d, 1H, Hb, J Hb- Hc = 8.7 Hz), 5.44 (d, 2H, HBn, J HBn-F = 48 Hz), 4.06 (t, 1H, Hα, J Hα-Hβ = 6.4 Hz), 3.89 (s, 3H, ArCOOCH 3 ), 2.76 (t, 2H, Hγ, J Hγ-Hα = 6.9 Hz), 2.25 (tt, 2H, Hβ, J Hβ-Hγ = 6.9 Hz, J Hβ-Hα = 6.4 Hz); 13 C NMR (CD 3)OD, 100 MHz): δ 173.1, 171.4, 167.7, 140.7, 131.9, 131.4, 131.0, 128.7, 126.0, 82.2 (d, CH 2 F, J CF = 164 Hz), 53.4, 52.7, 32.7, 26.8; HRMS 335.10105 (M + Na + ).
[0150]
[Synthesis Example 6] A
　4-position substituted product was synthesized according to the following synthesis scheme 6.
[0151]
(Scheme 6)

[0152]
(1) Synthesis of compound 30

[0153]
　(S) -5- (tert-butoxy) -4-((tert-butoxycarbonyl) amino) -5-oxopentanoic acid (811 mg, 2.67 mmol) was dissolved in dehydrated DMF (20 mL) and cooled to 0 ° C. did. HATU (952 mg, 4.01 mmol) and DIPEA (174 μL, 8.02 mmol) were added and stirred at 0 ° C. for 5 minutes. Subsequently, 2- (tert-butyldimethylsilyl) oxy) methyl) -4-methylaniline (807 mg, 3.21 mmol) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 10% → 20% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (1.31 g, 91%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.78 (brs, 1H, -CONH-), 8.00 (d, 1H, J = 7.8 Hz), 7.09 (d, 1H, J = 7.8 Hz), 6.91 (s) , 1H), 5.21 (brd, 1H, -OCONH-, J = 7.8 Hz), 4.70 (d, 1H, HBn, J gem = 13 Hz), 4.67 (d, 1H, HBn', J gem = 13 Hz) , 4.28-4.14 (m, 1H), 2.52-2.33 (m, 2H), 2.32-2.18 (m, 1H), 2.29 (s, 3H, ArCH 3)), 2.06-1.92 (m, 1H), 1.46 (s, 9H, COO (CH 3 ) 3 ), 1.42 (s, 9H, NHCOO (CH 3 ) 3 ), 0.90 (s, 9H, Si (CH 3 )) 3 ), 0.08 (s, 6H, Si (CH 3 ) 2 ).
[0154]
(2) Synthesis of compound 31

[0155]
　Compound 30 (1.31 g, 2.43 mmol) is dissolved in dehydrated THF (14 mL), TBAF (ca. 1 mol / L in THF, 7.3 mL, 7.3 mmol) and acetic acid (329 μL, 4.87 mmol) are added. , Stirred for 1 hour at room temperature. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 40% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (1.03 g, quantitative).
1 1 H NMR (CDCl 3 , 400 MHz): δ 8.75 (brs, 1H, -CONH-), 7.68 (d, 1H, J = 7.8 Hz), 7.07 (d, 1H, J = 7.8 Hz), 7.01 (s) , 1H), 5.36 (brd, 1H, -OCONH-, J = 8.2 Hz), 4.60 (d, 1H, HBn, J gem = 13 Hz), 4.54 (d, 1H, HBn', J gem = 13 Hz) , 4.22-4.10 (m, 1H), 2.48-2.32 (m, 2H), 2.32-2.06 (m, 1H), 2.27 (s, 3H, ArCH 3 ), 2.20-1.82 (m, 1H), 1.44 (s) , 9H, COO (CH 3 ) 3), 1.40 (s, 9H, NHCOO (CH 3 ) 3 ).
[0156]
(3) Synthesis of compound 32

[0157]
　Compound 31 (476 mg, 1.13 mmol) was dissolved in dehydrated dichloromethane (10 mL) and cooled to 0 ° C. DAST (738 μL, 5.63 mmol) was added and the mixture was stirred for 1 hour. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous sodium hydrogen carbonate solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 20% → 30% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (129 mg, 27%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.14 (brs, 1H, -CONH-), 7.68 (d, 1H, J = 8.2 Hz), 7.17 (d, 1H, J = 8.2 Hz), 7.12 (s) , 1H), 5.38 (d, 2H, HBn, J HBn-F = 48 Hz, J gem = 11 Hz), 5.30 (brd, 1H, -OCONH-, J = 6.0 Hz), 4.32-4.16 (m, 1H) ), 2.56-2.36 (m, 2H), 2.36-2.20 (m, 1H), 2.31 (s, 3H, ArCH 3 ), 2.20-1.84 (m, 1H), 1.45 (s, 9H, COO (CH 3 )) 3 ), 1.43 (s, 9H, NHCOO (CH) 3 ) 3 ).
[0158]
(4) Synthesis of compound 33

[0159]
　Compound 32 (38.5 mg, 0.0907 mmol) was added to 4M. It was dissolved in hydrochloric acid / ethyl acetate (1 mL) and stirred at room temperature for 12 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (0% → 100% acetonitrile / water) to obtain the desired product as a white solid (7.5 mg, 31%).
1 H NMR (CD 2 Cl 2 , 400 MHz): δ 7.26 (s, 1H), 7.23 (d, 1H, Hc, J Hc-Hb = 7.8 Hz), 7.17 (d, 1H, Hb, J Hb-Hc) = 7.8 Hz), 5.32 (d, 2H, HBn, J HBn-F = 48 Hz), 3.62 (t, 1H, Hα, J Ha-Hβ = 6.0 Hz), 2.63 (t, 2H, Hγ, J Hγ- Hβ = 7.3 Hz), 2.33 (s, 3H, ArCH 3 ), 2.22-2.10 (m, 2H, Hβ); 13 C NMR (CD 3)OD, 100 MHz): δ 174.2, 173.6, 137.7, 133.6, 132.8, 130.8, 130.2, 127.2, 82.5 (d, CH 2 F, J CF = 163 Hz), 55.4, 33.2, 27.8, 21.0; HRMS 269.13277 (M) + H + ).
[0160]
[Synthesis Example 7]
　Subsequently, a 5-position substituent was synthesized according to the following scheme 7.
[0161]
(Scheme 7)

[0162]
(1) Synthesis of compound 34

[0163]
　(S) -5 (tert-butoxy) -4-((tert-butoxycarbonyl) amino) -5-oxopentanoic acid (170 mg, 0.560 mmol) was dissolved in dehydrated DMF (6 mL) and cooled to 0 ° C. did. HATU (200 mg, 0.840 mmol) and DIPEA (364 μL, 1.68 mmol) were added, and the mixture was stirred at 0 ° C. for 5 minutes. Subsequently, methyl-3-amino-4-(((tert-butyldimethylsilyl) oxy) methyl) benzoate (198 mg, 0.672 mmol) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (199 mg, 61%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.93 (brs, 1H, -CONH-), 8.77 (s, 1H), 7.73 (d, 1H, J = 7.8 Hz), 7.20 (d, 1H, J = 7.8 Hz), 5.20 (brd, 1H, -OCONH-, J = 7.8 Hz), 4.78 (d, 1H, HBn, J gem = 13 Hz), 4.74 (d, 1H, HBn', J gem = 13 Hz) , 4.26-4.16 (m, 1H), 3.88 (s, 3H, ArCOOCH 3), 2.53-2.34 (m, 2H), 2.34-2.20 (m, 1H), 2.06-1.90 (m, 1H), 1.45 (s, 9H, COO (CH 3 ) 3 ), 1.41 (s, 9H, NHCOO) (CH 3 ) 3 ), 0.90 (s, 9H, Si (CH 3 ) 3 ), 0.08 (s, 6H, Si (CH 3 ) 2 ).
[0164]
(2) Synthesis of compound 35

[0165]
　Compound 34 (199 mg, 0.343 mmol) is dissolved in dehydrated THF (10 mL), TBAF (ca. 1 mol / L in THF, 1.03 mL, 1.03 mmol) and acetic acid (42 μL, 0.687 mmol) are added and room temperature. Was stirred for 2 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 40% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (116 mg, 73%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.96 (brs, 1H, -CONH-), 8.55 (d, 1H, J = 8.2 Hz), 7.75 (d, 1H, J = 7.8 Hz), 7.29 (d , 1H, J = 7.8 Hz), 5.34 (brd, 1H, -OCONH-, J = 7.8 Hz), 4.74 (dd, 1H, HBn, J gem = 13 Hz, J = 6.0 Hz), 4.67 (dd, 1H) , HBn', J gem = 13 Hz, J = 5.5 Hz), 4.26-4.12 (m, 1H), 3.88 (s, 3H, ArCOOCH 3 ), 3.51 (dd, 1H, CH 2)OH, J = 6.0 Hz, J = 5.5 Hz), 2.54-2.38 (m, 2H), 2.34-2.20 (m, 1H), 2.01-1.84 (m, 1H), 1.45 (s, 9H, COO (CH 3) ) 3 ), 1.41 (s, 9H, NHCOO (CH 3 ) 3 ).
[0166]
(3) Synthesis of compound 36

[0167]
　Compound 35 (110 mg, 0.236 mmol) was dissolved in dehydrated dichloromethane (10 mL) and cooled to 0 ° C. Fluoled (registered trademark) (119 mg, 0.472 mmol) was added, and the mixture was stirred for one and a half hours. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with dichloromethane. The dichloromethane layer was washed with saturated aqueous sodium hydrogen carbonate solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (43.6 mg, 39%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.47 (brs, 1H, -CONH-), 8.45 (s, 1H), 7.85 (d, 1H, J = 8.2 Hz), 7.43 (d, 1H, J = 8.2 Hz), 5.48 (d, 2H, HBn, J HBn-F = 48 Hz, J gem = 12 Hz), 5.33 (brd, 1H, -OCONH-, J = 7.8 Hz), 4.28-4.17 (m, 1H) ), 3.90 (s, 3H, ArCOOCH 3 ), 2.57-2.40 (m, 2H), 2.35-2.22 (m, 1H), 1.97 -1.82 (m, 1H), 1.45 (s, 9H, COO (CH 3 )) 3), 1.43 (s, 9H, NHCOO (CH 3 ) 3 ).
[0168]
(4) Synthesis of compound 37

[0169]
　Compound 36 (43.6 mg, 0.0931 mmol) was added to 4M. It was dissolved in hydrochloric acid / ethyl acetate (10 mL) and stirred at room temperature for 12 hours. After confirming the completion of the reaction, the reaction solution was concentrated. The residue was purified by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (19.4 mg, 60%).
1 1 H NMR (CD 3 OD, 400 MHz): δ 8.04 (s, 1H, Hc), 7.91 (d, 1H, Hb, J Hb-Ha = 7.8 Hz), 7.56 (d, 1H, Ha, J Ha- Hb = 7.8 Hz), 5.44 (d, 2H, HBn, J HBn-F = 48 Hz), 4.07 (t, 1H, Hα, J Hα-Hβ = 6.4 Hz), 2.75 (t, 2H, Hγ, J Hγ -Hα = 7.3 Hz), 2.26 (tt, 2H, Hβ, J Hβ-Hγ = 7.3 Hz, J Hβ-Hα = 6.4 Hz); 13 C NMR (CD 3)OD, 100 MHz): δ 172.0, 170.2, 166.3, 136.5, 134.6, 130.7, 127.8, 126.9, 126.6, 80.7 (d, CH 2 F, J CF = 165 Hz), 52.1, 51.5, 31.1, 25.6; HRMS 313.11879 (M + H + ).
[0170]
[Synthesis Example 8]
　Further, a 5-position substituted product was synthesized according to the following scheme 8.
[0171]
(Scheme 8)

[0172]
(1) Synthesis of compound 38

[0173]
　(S) -5- (allyloxy) -4-(((allyloxy) carbonyl) amino) -5-oxopentanoic acid (63.0 mg, 0.232 mmol) was dissolved in dehydrated DMF (5 mL) and cooled to 0 ° C. did. HATU (83.0 mg, 0.348 mmol) and DIPEA (151 μL, 0.697 mmol) were added, and the mixture was stirred at 0 ° C. for 5 minutes. Subsequently, 2- (tert-butyldimethylsilyl) oxy) methyl) -5-methylaniline (70 mg, 0.279 mmol) was added, the temperature was raised to room temperature, and the mixture was further stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water, a saturated aqueous sodium hydrogen carbonate solution and a saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (34 g silica gel, 20% → 30% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (59.0 mg, 50%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.80 (brs, 1H, -CONH-), 7.93 (s, 1H), 6.89 (d, 1H, J = 7.6 Hz), 6.76 (d, 1H, J = 7.6 Hz), 5.90-5.70 (m, 2H), 5.56 (brd, 1H, -OCONH-, J = 8.0 Hz), 5.28-5.10 (m, 4H), 4.62 (s, 2H, HBn), 4.58-4.40 (m, 4H), 4.40-4.30 (m, 1H), 2.51-2.30 (m, 2H), 2.36-2.21 (m, 1H), 2.26 (s, 3H, ArCH 3)), 2.13-1.95 (m, 1H), 0.83 (s, 9H, Si (CH 3 ) 3 ), 0.00 (s, 6H, Si (CH 3 ) 2 ).
[0174]
(2) Synthesis of compound 39

[0175]
　Compound 39 (59.0 mg, 0.117 mmol) is dissolved in dehydrated THF (3 mL), TBAF (ca. 1 mol / L in THF, 351 μL, 0.351 mmol) and acetic acid (16 μL, 0.234 mmol) are added and room temperature. Was stirred for 12 hours. After confirming the completion of the reaction, water was added and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with water and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 40% → 60% ethyl acetate / hexane) to obtain the desired product as a colorless liquid (32.6 mg, 71%).
1 H NMR (CDCl 3 , 400 MHz): δ 8.68 (brs, 1H, -CONH-), 7.80 (s, 1H), 7.06 (d, 1H, J = 7.8 Hz), 6.88 (d, 1H, J = 7.8 Hz), 5.96-5.80 (m, 2H), 5.66 (brd, 1H, -OCONH-, J = 7.8 Hz), 5.36-5.14 (m, 4H), 4.71-4.35 (m, 8H), 2.87 (brs) , 1H, CH 2 OH), 2.55-2.41 (m, 2H), 2.39-2.27 (m, 1H), 2.32 (s, 3H, ArCH 3 ), 2.14-1.97 (m, 1H).
[0176]
(3) Synthesis of compound 40

[0177]
Compound 39 (32.6 mg, 0.0835 mmol) was dissolved in dehydrated dichloromethane (2 mL) and cooled to 0 ° C. DAST (55 μL, 0.417 mmol) was added and the mixture was stirred for 3 hours. After confirming the completion of the reaction, saturated aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The ethyl acetate layer was washed with saturated aqueous sodium hydrogen carbonate solution and saturated brine, dried over sodium sulfate, and then concentrated. The residue was purified by silica gel chromatography (14 g silica gel, 20% → 40% ethyl acetate / hexane) to obtain the desired product as a pale yellow liquid (10.3 mg, 31%).
1 H NMR (CDCl 3 , 400 MHz): δ 7.77 (s, 1H), 7.17 (d, 1H, J = 7.3 Hz), 6.96 (d, 1H, J = 7.3 Hz), 5.97-5.82 (m, 2H) ), 5.57 (brd, 1H, -OCONH-, J = 6.9 Hz), 5.40 (d, 2H, HBn, J HBn-F = 48 Hz, J gem = 11 Hz), 5.36-5.17 (m, 4H), 4.65 (d, 2H, J = 5.5 Hz), 4.59-4.42 (m, 3H), 2.59-2.42 (m, 2H), 2.45-2.27 (m, 1H), 2.36 (s, 3H, ArCH 3 ), 2.15 -2.00 (m, 1H).
[0178]
(4) Synthesis of compound 41

[0179]
　Compound 40 (10.3 mg, 0.0262 mmol) and phenylsilane (33 μL, 0.262 mmol) are dissolved in dehydrated dichloromethane (1 mL), tetrakis (triphenylphosphine) palladium (7.6 mg, 25 mol%) is added, and room temperature is reached. Was stirred for 1 hour. After confirming the completion of the reaction, the reaction solution was directly subjected to purification by reverse phase HPLC (20% → 100% acetonitrile / water) to obtain the desired product as a white solid (3.1 mg, 44%).
1 H NMR (CD 3 OD, 400 MHz): δ 7.31 (d, 1H, Hb, J Hb-Ha = 7.8 Hz), 7.22 (s, 1H, Hc), 7.09 (d, 1H, Ha, J Ha- Hb = 7.8 Hz), 5.32 (d, 2H, HBn, J HBn-F = 48 Hz), 3.65 (t, 1H, Hα, J Hα-Hβ = 6.0 Hz), 2.71-2.59 (m, 2H, Hγ) , 2.32 (s, 3H, ArCH 3 ), 2.23-2.10 (m, 2H, Hβ); 13 C NMR (CD 3 OD, 100 MHz): δ 172.0, 170.1, 139.4, 134.8, 130.0, 128.7, 127.0, 126.4 , 81.1 (d, CH 2 F, J CF = 163 Hz), 52.3, 31.2, 25.7, 19.8; HRMS 291.08980 (M + Na + ).
[0180]
[Example 6]
Evaluation of in vitro drug efficacy of benzene ring substituent conversion derivative
　The result of CCK-8 assay of the 4-position substituted derivative is shown in FIG. Derivatives substituted with electron donating groups tended to exhibit relatively strong antitumor activity.
[0181]
　The results of the CCK-8 assay for the 5-position substituted derivative are shown in FIG. The effect of the substituent at the 5-position, which corresponds to the para-position of the leaving group, is stronger than that at the 4-position, and the difference in drug efficacy between 5-COOME, 5-H, and 5-Me is more remarkable. In addition, the drug efficacy evaluation of 5-COOME derivatives by GGT low-expressing cells is shown in the lower part of FIG.
[0182]
　As shown above, the prodrug-type anticancer agent of the present invention has a high reaction having an azaquinone methide structure at the same time that the L-glutamate moiety is cleaved by GGT on the cancer cell membrane that highly expresses GGT. It shows an antitumor effect by the mechanism that a sex substance (toxicity inducer) is released. As a result of confirming the efficacy by co-culture imaging using a GGT high expression / low expression cell line, the prodrug type anticancer agent of the present invention is a GGT high expression cell line without killing adjacent GGT low expression cells. It was possible to selectively kill only. This result suggests that the prodrug-type anticancer agent of the present invention may show a wide safety margin even in vivo.
[0183]
[Example 7]
Administration test of gGlu-FMA (Compound 3) to peritoneal dissemination model mice
(1) Peritoneal dissemination
　Peritoneal dissemination is a state in which tumor cells are scattered on the surface of the peritoneum covering the peritoneum and engrafted. As expected. In clinical practice, metastasis often occurs from gastric cancer, colorectal cancer, ovarian cancer, etc., and no epoch-making treatment method including chemotherapy has been established.
[0184]
(2) Preparation of model mouse and outline of test
　A model mouse for peritoneal dissemination was prepared by intraperitoneally administering SHIN3 cells (ovarian cancer-derived cancer cell line, high GGT activity) suspended in PBS. From 7 days after cell seeding, 5 mg / kg of gGlu-FMA or PBS (control) was intraperitoneally administered daily, and 21 days after cell seeding, gGlu-HMRG was intraperitoneally administered, and 10 minutes later, the mice were sacrificed. After the abdomen was opened, a fluorescent image was acquired with an in vivo imaging device maestro (FIG. 16).
[0185]
(3) Results
　FIG. 17 shows a macro image of the mesentery of mice administered with PBS and gGlu-FMA and two fluorescent images thereof. Both macro and fluorescent images appear to have an overall reduction in tumors in gGlu-FMA-treated mice.
[0186]
[Industrial Availability]
　The prodrug-type anticancer agent of the present invention recognizes the difference in metabolic enzyme activity between two adjacent cells and kills only cancer cells in which the enzyme activity is enhanced. It is expected to be an innovative clinical drug that can improve the major problem in cancer chemotherapy of lowering the QOL of patients due to serious side effects. Furthermore, since the azaquinone-methide-releasing prodrug of the present invention that does not use an existing anticancer agent can be easily synthesized, it is expected to contribute significantly to the development cost. In this way, the medical and industrial utility value and economic value of the developed prodrug are considered to be extremely large.
The scope of the claims
[Claim 1]
　A compound represented by the following general formula (I) or a salt thereof.

(In the formula,
X is a fluorine atom, an ester group (-OC (= O) -R'), a carbonate group (-OCO 2- R'), a carbamate group (-OCONH-R'), a phosphoric acid and an ester thereof. Selected from the group consisting of groups (-OP (= O) ( -OR') ( -OR ") , and sulfuric acid and its ester groups (-OSO 2- OR'),
　where R', R'". Are independently selected from substituted or unsubstituted alkyl groups or substituted or unsubstituted aryl groups;
Y is -NH-COL, -NH-L'or -OL',
　where In, L is a partial structure of an amino acid, and
　L'is a saccharide or a partial structure of a saccharide, a saccharide having a self-cleaving linker, an amino acid or peptide having a self-cleaving linker;
R 1 and R. 2 are each independently selected from a hydrogen atom or a monovalent substituent;
R 3 represents a hydrogen atom or 1 to 4 identical or different monovalent substituents present on the benzene ring).
[Claim 2]
　The compound according to claim 1, wherein the partial structure of the amino acid of L, together with C = O to which it is bound, constitutes an amino acid, an amino acid residue, a peptide, a part of an amino acid. salt.
[Claim 3]
　The compound according to claim 1, or a salt thereof, wherein the partial structure of the saccharide of L'combines with O to which it is bound to form a saccharide, a part of the saccharide.
[Claim 4]
Any one of claims 1 to 3, wherein 　-Y in the general formula (I) is bonded to -C (R 1 ) (R 2 ) X on the ortho-position or para-position of the benzene ring. The compound or salt thereof.
[Claim 5]
　The compound according to any one of claims 1 to 4, or a salt thereof, wherein Y has a structure selected from the following.

[Claim 6]
　The compound according to any one of claims 1 to 5, or a salt thereof, wherein X is a fluorine atom or an ester group (-OCO-R').
[Claim 7]
The compound according to any one of claims 1 to 6, or a salt thereof, wherein 　R 1 and R 2 are independently selected from a hydrogen atom or a fluorine atom.
[Claim 8]
The monovalent substituent of 　R 3 is an alkyl group, an alkoxycarbonyl group, a nitro group, an amino group, a hydroxyl group, an alkylamino group (-NHR', -NHCOR'), an alkoxy group (-OR', -OCOR'), Any one of claims 1 to 7 selected from the group consisting of a halogen atom, a boryl group and a cyano group (R'is a substituted or unsubstituted alkyl group or a substituted or unsubstituted aryl group). The compound according to the section or a salt thereof.
[Claim 9]
　R 3 monovalent substituent of an alkyl group (e.g., methyl group) or alkoxycarbonyl group (e.g., methoxycarbonyl group), a compound or a salt thereof according to claim 8.
[Claim 10]
　A prodrug-type anticancer agent comprising the compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof.
[Claim 11]
　A prodrug-type anticancer agent containing the compound according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof, which acts cell-selectively by a cancer cell-specific enzyme activity.
[Claim 12]
　The prodrug-type anticancer agent according to claim 11, wherein the enzyme is peptidase or glycosidase.