WO2010090234A1 - 12-deoxyfusicoccin sugar chain-modified derivatives and uses thereof - Google Patents

Abstract

Disclosed are novel water-soluble compounds that have differentiation-inducing effects on mammalian cells, cell differentiation-inducing agents that have said compounds as active ingredients and anti-tumor agents. Disclosed are compounds and salts thereof that are represented by general formula (1) (1) (wherein n represents an integer 0‑5, Me a methyl group, and R¹ a 1- to 6-carbon alkyl group or amino group represented by formula (2) (2) (wherein R² represents a hydrogen atom or 1- to 6-carbon alkyl group, R³ a hydrogen atom, 1- to 6-carbon alkyl group, a 1- to 7-carbon acyl group or amidyl group (–CNHNH₂)])

Description

12-Deoxyfusicoccin sugar chain modified derivatives and uses thereof

The present invention relates to a novel water-soluble compound having a cell differentiation-inducing action. More specifically, the present invention relates to a 12-deoxy derivative of fusicoccin having the above properties and having a modified sugar chain (hereinafter, also simply referred to as “12-deoxyfusucocin sugar chain-modified derivative”). Furthermore, the present invention relates to a cell differentiation inducer and an antitumor agent comprising such a 12-deoxyfusicoccin sugar chain-modified derivative as an active ingredient.

∙ Many chemotherapeutic agents in tumor treatment are cytotoxic substances and may have serious side effects depending on the dose and method of administration. Tumor cells repeat proliferation while remaining undifferentiated during differentiation. Therefore, focusing on this property of tumor cells, a method of suppressing the proliferation of tumor cells by differentiating undifferentiated tumor cells using a cell differentiation inducer is a trial for treating tumors, particularly malignant tumors (cancers). (See Patent Document 1). Among such cell differentiation inducers, cotylenin C, a diterpene glycoside having a plant hormone-like activity (plant growth promoting activity), induces differentiation of mouse myeloid leukemia cells into mature cells, Although it was thought that it had a potential as an anticancer agent for a long time (see Patent Document 2), it was later found that it alone has an effect of inducing differentiation of tumor cells of patients with acute myeloid leukemia into normal mature cells. It is expected to be a therapeutic drug for leukemia (see Non-Patent Document 1).

Also, it is known that cell differentiation inducers are effective in tumors such as cancer by inducing apoptosis when used in combination with interferon α. For example, when cotylenin is used in combination with interferon α as a cell differentiation inducer, it exerts a strong apoptosis-inducing activity against various tumor cells and a remarkable tumor growth-inhibiting effect resulting from it. A remarkable effect is also shown against cellular lung cancer (see Non-Patent Document 2) and ovarian cancer (see Non-Patent Document 3). Furthermore, cotylenin is also effective against cell lines that are resistant to cisplatin and taxol. On the other hand, it has been confirmed that the above effect is highly specific to tumor cells with little effect on normal cells. Cochilenin is effective not only for cultured cells but also for cells collected from actual ovarian cancer patients.

Furthermore, cotylenin also shows a remarkable synergistic effect when used in combination with the cell cycle inhibitor rapamycin, and it has been demonstrated using breast cancer cells that the tumor growth inhibitory effect of cell cycle G1 phase arrest is shown (non-patented). Reference 4).

For this reason, cotylenin is highly expected as a new active ingredient of an antitumor agent that is effective for intractable tumors resistant to various antitumor agents and has low toxicity. However, clinical application of cotylenin has not yet been made.

As described above, a method for industrially synthesizing and stably supplying the necessary amount of cotyrenin has been established as a reason why it has not yet been applied clinically despite its excellent cell differentiation-inducing activity. Not to mention.

On the other hand, Fusicoccin, which is chemically similar to cotyrenin, is stably produced by Pomoopsis amygdali. Although the fusicoccin has a plant hormone-like activity equivalent to that of cotyrenin, it has almost no cell differentiation-inducing activity.

However, subsequent experiments by the present inventors have revealed that the 12-deoxy derivative of the fusicoccin (12-deoxyfusicoccin derivative) exhibits cell differentiation-inducing activity similar to cotyrenin (Patent Document 3). A representative compound is 4 ', 6'-isopropylidene-12-deoxyfusicoccin J (ISIR-005). Such a 12-deoxyfusicoccin derivative has a good differentiation-inducing activity against leukemia tumor cells in addition to low toxicity, and it can induce apoptosis in various solid tumors when used in combination with interferon α. From the viewpoint of induction, it is expected as an active ingredient of antitumor agents. However, there is a problem that it is poor in water solubility and it is difficult to adjust to various dosage forms including liquids such as injections.

Japanese Patent Laid-Open No. 05-178821 JP 04-149133 A International publication WO 2008/010324 A1

An object of the present invention is to provide a water-soluble 12-deoxyfusicoccin sugar chain-modified conductor having a differentiation-inducing action particularly on leukemic tumor cells. Another object of the present invention is to provide a cell differentiation inducer and an antitumor agent comprising such a 12-deoxyfusicoccin sugar chain-modified derivative as an active ingredient.

As a result of intensive research aimed at solving the above problems, the present inventors have found that the above 4 ′, 6′-isopropylidene-12-deoxyfusicoccin J (hereinafter also simply referred to as “ISIR-005”) A modified sugar chain of the following formula (1):

(In the formula, n is an integer of 0 to 5, Me is a methyl group, R ¹ is an alkyl group having 1 to 6 carbon atoms, or an amino group represented by the following formula:

[Wherein R ² is a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; R ³ is a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, or an amidyl group (—CNHNH ₂ ). ]. )
It is found that the compound represented by the formula has cell differentiation-inducing activity superior to ISIR-005, and good apoptosis-inducing activity in combination with interferon α, and the compound (1) has the desired water solubility. It was confirmed.

Further, the inventors of the present application said that a part of the compounds included in the compound (1) group more significantly suppresses the growth of solid tumor cells in a Hypoxia (hypoxia) state in addition to the above action. It was found to have a very rare and useful action that was not seen with ISIR-005. The inside of the developed tumor tissue is often under hypoxia, which causes treatment resistance in radiation therapy and chemotherapy. Therefore, it is extremely important in cancer treatment strategies to exert an effective growth inhibitory effect even in a hypoxic environment and to increase the sensitivity of tumor cells to anticancer agents in a hypoxic environment. Therefore, the above compounds can be expected to have an effective antitumor effect against malignant tumors that have developed and become difficult to treat with conventional radiation therapy and chemotherapy, and in a hypoxia state unlike normal cells. It is believed that certain tumor cells (especially solid tumor cells) can be selectively targeted.

The present invention has been completed based on these findings and has the following embodiments.
(I) 12-deoxy fusicoccin sugar chain-modified derivative (I-1) compound represented by general formula (1) or salt thereof:

(Wherein n is an integer of 0 to 5, Me is a methyl group, R ¹ is an alkyl group having 1 to 6 carbon atoms, —N ₃ group or an amino group represented by the following formula:

[Wherein R ² is a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; R ³ is a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, or a p-nitrophenylsulfonyl group. Alternatively, it represents an amidyl group (—CNHNH ₂ ). ]
Indicates. ).

(I-2) In general formula (1),
R ¹ is an alkyl group having 1 to 6 carbon atoms, or an amino group represented by formula (2) (wherein R ³ is a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, The compound according to (I-1), which is an acyl group of 7 or an amidyl group (-CNHNH ₂ ).

(I-3) In general formula (1),
n is 0 and R ¹ is a methyl group, or
n is an integer of 2 to 5, and R ¹ is an amino group represented by the following formula (2):

[Wherein R ² represents a hydrogen atom; R ³ represents a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, or an amidyl group. ]
The compound described in (I-1) or (I-2).

(I-4) The compound described in (I-1), wherein the compound represented by the general formula (1) is a compound represented by any of the following formulas (3) to (12):

(II) Cell differentiation inducer (II-1) A cell differentiation inducer comprising as an active ingredient the compound described in any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof.

(III) Antitumor agent against hematological malignancy comprising as an active ingredient the compound according to any one of (III-1) (I-1) to (I-4) or a pharmaceutically acceptable salt thereof Agent.
(III-2) The antitumor agent according to (III-1), wherein the hematological malignancy is any one selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma.
(III-3) An antitumor agent comprising a combination of the compound described in any of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof and interferon α.
(III-4) The composition according to (III-3), which is a composition comprising the compound according to any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof and interferon α. Antitumor agent.
(III-5) a drug containing the compound according to any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof and a drug containing interferon α as an active ingredient, The antitumor agent according to (III-3), which is provided as a separately packaged combination kit.
(III-6) The antitumor agent according to (III-3), wherein the target tumor is a lung tumor or a breast tumor.

(IV) Use of the compound according to any one of (I-1) to (I-4) (IV-1) The compound according to any one of (I-1) to (I-4) or a pharmaceutical product thereof A method for treating a tumor, comprising administering an effective amount of a pharmaceutically acceptable salt to a tumor patient.
(IV-2) An effective amount of the compound according to any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof and a combination of interferon α is administered to a tumor patient. A method for treating a tumor, comprising:
(IV-3) The hematological malignancy selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma, or a solid cancer selected from the group consisting of breast cancer, pancreatic cancer, lung cancer and ovarian cancer The treatment method described in (IV-1) or (IV-2).
(IV-4) The compound according to any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof, which is used for treating a tumor.
(IV-5) The hematological malignancy selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma, or a solid cancer selected from the group consisting of breast cancer, pancreatic cancer, lung cancer and ovarian cancer The compound or a salt thereof described in (IV-4).
(IV-6) A compound according to any one of (I-1) to (I-4) or a pharmaceutically acceptable salt thereof and a combination of interferon α and used for the treatment of tumors.
(IV-7) Any hematological malignancy selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma, or solid cancer selected from the group consisting of breast cancer, pancreatic cancer, lung cancer and ovarian cancer The combination described in (IV-6).

According to the present invention, there is provided a novel compound (1) which has a cell differentiation inducing action and is useful as a cell differentiation inducer and as an active ingredient of an antitumor agent, particularly an anti-hematologic malignant tumor agent or an intermediate thereof. . The compound (1) (excluding the intermediate) has an effect of enhancing the tumor activity of an antitumor agent such as interferon α and is useful as an active ingredient of an antitumor agent itself. It is also useful as an antitumor effect enhancer of antitumor agents such as

Compound of the present invention represented by general formula (1) (ISIR-040: Compound (3), ISIR-041: Compound (4), ISIR-042: Compound (5), ISIR-043: Compound (6), ISIR -044: This represents a synthesis scheme of the compound (7)). The abbreviations and abbreviations of substituents in the drawings mean the following terms (the same applies to FIGS. 2 and 3). Me: methyl group, Ac: acetyl group, Ms: methanesulfonyl group, Ns: 4-nitrobenzenesulfonyl group, Cbz: benzyloxycarbonyl group, AcCl: acetyl chloride, DMAP: dimethylaminopyridine, PPTS: pyridinium p-toluenesulfonic acid , MsCl: Methanesulfonyl chloride, aq. NaHCO ₃ : Sodium bicarbonate aqueous solution, THF: Tetrahydrofuran, HMPA: Hexamethylphosphoramide, Et ₃ N: Triethylamine, Ac ₂ O: Acetic anhydride, PS-carbodiimide: Supported on polystyrene resin Carbodiimide reagent, MeI: methyl iodide, DMF: dimethylformamide. The compound of the present invention represented by the general formula (1) (ISIR-042: Compound (5), ISIR-062: Compound (8), ISIR-072: Compound (9), ISIR-082: Compound (10)) Represents a synthesis scheme. This represents a synthesis scheme of the compound of the present invention represented by the general formula (1) (ISIR-051: Compound (11), ISIR-052: Compound (12)). It is a graph which shows the cell differentiation-inducing activity of ISIR-040 (compound (3)) and cotylenin A (CN-A) with respect to human acute myeloid leukemia cells (HL-60 cells) (Test Example 1). ISIR-040 (compound (3)), ISIR-042 (compound (5)), ISIR-062 (compound (8)), ISIR-082 (compound (10)) for human monocytic leukemia cells (U937 cells) 1 is a graph showing the cell differentiation-inducing activity of ISIR-005 and Cochirenin A (CN-A) (Test Example 1). ISIR-041 (compound (4)), ISIR-042 (compound (5)), ISIR-043 (compound (6)), ISIR-044 (compound (7)) for human monocytic leukemia cells (U937 cells) 2 is a graph showing the cell differentiation-inducing activity of ISIR-005 (Test Example 1). It is a graph which shows the cell differentiation-inducing activity of ISIR-042 (compound (5)) and ISIR-072 (compound (9)) against human monocytic leukemia cells (U937 cells) (Test Example 1). The test result which evaluated the inhibitory effect with respect to the lung cancer cell (A549) proliferation by combined use of ISIR-042 (compound (5)) and interferon (alpha) is shown (test example 2). The vertical axis represents the number of viable cells (%) relative to the control. The results of measuring the inhibitory effect of ISIR-042 (compound (5)) on breast cancer cells (MCF-7) under normal oxygen concentration (21%) and hypoxia (1%) are shown (Test Example 3). . The vertical axis represents the number of viable cells (%) relative to the control. The growth inhibitory effect with respect to the pancreatic cancer cell (MiaPaca-2) of the known anticancer drug Gemcitabin is shown in a result of measurement under normal oxygen concentration (21%) and hypoxia (1%) (Test Example 3). The vertical axis represents the number of viable cells (%) relative to the control. The results of measuring the inhibitory effect of ISIR-042, ISIR-005, and CN-A on pancreatic cancer cells (MiaPaca-2) under normal oxygen concentration (21%) and hypoxia (1%) are shown ( Test Example 3). The vertical axis represents the number of viable cells (%) relative to the control. The results of measuring the growth inhibitory action of ISIR-042 and ISIR-041 on pancreatic cancer cells (Panc-1) under normal oxygen concentration (21%) and under hypoxia (1%) are shown (Test Example 3). The vertical axis represents the number of viable cells (%) relative to the control.

(I) 12-Deoxyfuscoccine sugar chain-modified derivative The 12-deoxyfusicoccin sugar chain-modified derivative targeted by the present invention can be represented by the following general formula (1):

In the above formula (1), R ¹ means a linear or branched alkyl group having 1 to 6 carbon atoms, —N ₃ group, or an amino group represented by the following general formula (2):

Here, examples of the linear or branched alkyl group having 1 to 6 carbon atoms represented by R ¹ include a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, a pentyl group, an isopentyl group, A hexyl group and an isohexyl group are included. Particularly preferred is a methyl group.

In the above formula (2), R ² is a hydrogen atom or a linear or branched alkyl group having 1 to 6 carbon atoms. Preferably it is a hydrogen atom. Here, examples of the linear or branched alkyl group having 1 to 6 carbon atoms include the above-described alkyl groups.

In the above formula (2), R ³ represents a hydrogen atom, a linear or branched alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, a p-nitrophenylsulfonyl group, or an amidyl group ( -CNHNH ₂ ). Particularly preferred is a hydrogen atom.

Here, examples of the linear or branched alkyl group having 1 to 6 carbon atoms include the above-described alkyl groups.

The acyl group having 1 to 7 carbon atoms represented by R ³ means a group represented by CO—R ⁴ . Here, R ⁴ includes an alkyl group having 1 to 6 carbon atoms, an alkenyl group having 2 to 6 carbon atoms, and an alkynyl group having 2 to 6 carbon atoms. Here, examples of the alkyl group having 1 to 6 carbon atoms include the above-described alkyl groups. A methyl group is preferred. Examples of the alkenyl group having 2 to 6 carbon atoms include a vinyl group, a propenyl group, a butenyl group, a pentenyl group, and a hexenyl group. A vinyl group is preferred. Examples of the alkynyl group having 2 to 6 carbon atoms include ethynyl group, propynyl group, butyryl group, pentynyl group, and hexynyl group. An ethynyl group is preferable.

The preferred acyl group acetyl group R ⁴ is a methyl group having one carbon atom, and R ⁴ is an ethynyl group or an ethynyl group having 2 carbon atoms.

In the above formula (1), n represents an integer of 0 to 5. In the formula (1), when R ¹ is an alkyl group having 1 to 6 carbon atoms, n is preferably 0. When R ¹ is an amino group represented by the formula (2), n is preferably an integer of 2 to 5.

Specific examples of the compound of the present invention represented by the formula (1) include compounds represented by the following formulas (3) to (12) (hereinafter also referred to as compounds (3) to (12)). Can do.

Of the above compounds (3) to (12), preferably compound (4) wherein n is 2 and R ¹ is an amino group in formula (1), compound (5) where n is 3 and R ¹ is an amino group , And n is 3 and R ¹ is the compound (9) in which R ² is a hydrogen atom and R ³ is an ethynylcarbonyl group in the formula (2). More preferred are compound (5) and compound (9).

Among the compounds of the present invention represented by the formula (1), a compound in which R ¹ is a —N ₃ group or R ³ is a p-nitrophenylsulfonyl group is a compound of the present invention having a cell differentiation inducing action. It is useful as a synthetic intermediate for compounds, particularly the above compounds (3) to (12). Specifically, examples of the compound (1) in which R ¹ is a —N ₃ group include SI-05 (Production Example 2 (1)), SI-06 (Production Example 3 (1)), SI-07 ( Production Example 4 (1)) and SI-08 (Production Example 5 (1)) can be mentioned, and these are the above-mentioned compound (4) (see ISIR-041: Production Example 2 (2)), compound ( 5) (ISIR-042: See Preparation Example 3 (2)), Compound (6) (ISIR-043: See Preparation Example 4 (2)), and Compound (7) (ISIR-044: Preparation Example 5 (2) (See FIG. 1). Examples of the compound (1) in which R ³ is a p-nitrophenylsulfonyl group include SI-09 (Production Example 9 (1)) and SI-10 (Production Example 9 (2)). These are all synthetic intermediates of the above compound (10) (see ISIR-082: Production Example 9 (3)) (see FIG. 2).

The compound of the present invention represented by the general formula (1) is produced, for example, by changing reaction conditions, adding a reaction, omitting a reaction, etc., if necessary, based on the synthetic scheme shown in FIGS. be able to.

FIG. 1 is a synthesis scheme using 4 ′, 6′-isopropylidene-12-deoxyfusicoccin J (ISIR-005) described in Patent Document 3 (International Publication WO2008 / 010324) as a starting material. Is a synthesis scheme starting from compound (5) (ISIR-042). FIG. 3 is a synthesis scheme using Compound (4) (ISIR-041) and Compound (5) (ISIR-042) as starting materials.

In addition, ISIR-005 used as a raw material is synthesized from natural fusicoccins that are stably produced by Pomoopsis 病 amygdali, as described in Patent Document 3 (International Publication WO2008 / 010324). be able to. Natural fusicocins include, for example, (1) KD Barrow, D. H. R. Barton, Sir E. Chain, C. Conlay, T. C. Smale, R. Thomas, and E. S. Waight, J. Chem. Soc. (C), 1259 (1971); or (2) N. Tajima, M. Nukina, N.Kato, ukand T. Sassa, Biosci. Biotechnol. Biochem., 68, 1125 (2004) (Japan) It can be obtained by the method described in the literature on cultivation of peach branch rot fungus Phomopsis amygdali Niigata 2-A).

In FIGS. 1, 2 and 3, the compound ISIR-040 is the compound (3) described above, ISIR-041 to 044 is the compound (4) to (7) described above, and ISIR-062 is the compound (8) described above. Compound ISIR-072 corresponds to compound (9) described above, ISIR-082 corresponds to compound (10) described above, and ISIR-051 and ISIR-052 correspond to compounds (11) and (12) described above, respectively.

As shown in FIG. 1, for example, the compound of the general formula (1) in which R ¹ represents an alkyl group having 1 to 6 carbon atoms (compound ISIR-040: compound (3)) is, for example, the above known compound ISIR-005. Is then acetylated to form compound SI-01, followed by ring-opening reaction (generation of SI-02), sulfonylation (generation of SI-03), deacetylation (generation of SI-04), and ring-closing reaction Manufactured. As reaction conditions for acetylation, ring-opening reaction, sulfonylation, deacetylation, and ring-closing reaction, known conditions can be widely applied. Specifically, it can be produced according to the reaction conditions described in Production Example 1 described later. Further, as shown in FIG. 1, the compounds of the general formula (1) in which R ¹ represents an N ₃ — group (compounds SI-05 to 08) are compound (3) (compound ISIR-040) produced by the above method. ) Can be produced by azidation using an azidoalkanol. As the reaction conditions for the azidation reaction, known conditions can be widely applied. Specifically, it can be produced according to the reaction conditions described in Production Examples 2 to 5 described later. The compounds (4) to (7) (compounds ISIR-041 to 044) obtained here can also be prepared in the form of salts by a conventional method shown in Production Example 6.

As shown in FIG. 2, for example, a compound in which R ¹ is an amino group represented by the general formula (2) (wherein R ² is a hydrogen atom and R ³ is an acyl group having 1 to 7 carbon atoms) ( 1) (Compound ISIR-062: Compound (8)) is produced, for example, by acetylating compound ISIR-042 (compound (5)) produced by the above method, and R ¹ is generally the same as above. A compound (compound ISIR-072: compound (9)) which is an amino group represented by the formula (2) can be produced, for example, by ethynylcarbonylating the above compound ISIR-042 (compound (5)).

Compound (1) wherein R ¹ is an amino group represented by the general formula (2) (wherein either R ² or R ³ is a hydrogen atom and the other is an alkyl group having 1 to 6 carbon atoms) (Compound ISIR-082: Compound (10)) sulfonylates the compound ISIR-042 (compound (5)) produced by the above method to produce compound SI-09, which is then methylated (compound SI -10) and desulfonylation.

Further, as shown in FIG. 3, for example, a compound (1) in which R ¹ is an amino group represented by the general formula (2) (where R ² is a hydrogen atom, R ³ is an amidyl group (—CNHNH ₂ )) Compound ISIR-051: Compound (11) and Compound ISIR-052: Compound (12)) are, for example, compound ISIR-041 (compound (4)) and compound ISIR-042 (compound (5)) produced by the above-described method. Can be produced by amidylation.

These compounds have cell differentiation-inducing activity and are useful as active ingredients of cell differentiation-inducing agents or antitumor agents, as shown in the test examples described later. Among them, ISIR-041 (compound (4)) and ISIR-042 (compound (5)) are effective against various malignant tumor cells (breast cancer cells, pancreatic cancer cells, lung cancer cells, ovarian cancer cells) even in a hypoxic environment. Demonstrate cell growth inhibitory effect (Test Example 3). Furthermore, ISIR-042 (compound (5)) exhibits an effect of significantly suppressing the growth of malignant tumor cells when used in combination with interferon α (Test Example 2).

Of the compounds shown in FIGS. 1, 2 and 3, the compound represented by SI is useful as an intermediate of the above compounds (3) to (12) which can be an active ingredient of an antitumor agent.

Among these compounds, (4) to (7) and (10) having an amino group, and (11) and (12) having a guanidyl group can have a free or salt form. Examples of the salt include pharmaceutically acceptable salts such as acid addition salts such as inorganic acids, organic acids, or acidic amino acids.

Examples of inorganic acids that form acid addition salts include hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, and the like. Examples of organic acids include formic acid, acetic acid, lactic acid, trifluoroacetic acid, fumaric acid, oxalic acid, tartaric acid, maleic acid, benzoic acid, citric acid, succinic acid, malic acid, ascorbic acid, methanesulfonic acid, ethanesulfonic acid Benzenesulfonic acid, p-toluenesulfonic acid and the like. Examples of acidic amino acids include aspartic acid and glutamic acid.

(II) Drug, cell differentiation inducer and antitumor agent The compound of the present invention represented by the general formula (1) has cell differentiation induction activity. In particular, the compounds (3) to (12) are used in the cell differentiation-inducing activity test (testability 1) using human leukemia cells (human myeloid leukemia HL-60 cells or human monocytic leukemia U937 cells) described later. It exhibits an activity similar to that of cotylenin A and has an effect of inducing cell differentiation from abnormal cells such as tumor cells to mature cells. Since the cell differentiation inducing activity induces cell differentiation, the compound (1) of the present invention can suppress the proliferation of cells exhibiting abnormal differentiation such as tumor cells. Therefore, the compound of the present invention represented by the general formula (1) is useful as a cell differentiation inducer. Moreover, since proliferation of tumor cells can be suppressed based on cell differentiation-inducing activity, it is also useful as a drug, specifically as an antitumor agent. The target tumors include benign tumors and malignant tumors. A malignant tumor (cancer) is preferred.

Examples of the cells in which the cell growth inhibitory action by the cell differentiation-inducing activity of the compound (1) of the present invention is effective include tumor cells such as leukemia cells, malignant lymphomas, and hematological malignancies including malignant myeloma. . Leukemia cells, malignant lymphoma, and malignant myeloma are preferable as growth inhibition targets, and leukemia cells (acute myeloid leukemia cells, monocytic leukemia cells) are more preferable. In addition, solid cancers such as breast cancer, pancreatic cancer, lung cancer, and ovarian cancer can also be targeted.

Furthermore, the compound (1) of the present invention can also be used in combination with interferon α. Among them, as shown in Test Example 2, compound (5) can synergistically enhance the antitumor activity of interferon α when used in combination with interferon α. Therefore, the compound of the present invention can be used as an antitumor activity enhancer of interferon α, or can be used as an antitumor agent in combination with interferon α. The embodiment to be combined here is not particularly limited, and is an embodiment in which the compound of the present invention and interferon α are mixed in the form of a composition (mixture) from the beginning, and the compound of the present invention and interferon α are separately packaged in packaging form. And any of the aspects (combination kit) used combining both at the time of use may be sufficient.

As shown in Test Example 3, among the compounds (1) of the present invention, particularly the compound (5) remarkably suppresses the growth of cancer cells in the Hypoxia state. In general, it is known that cancer tissue is hypoxic and resistant to chemotherapeutic agents. Therefore, the compound of the present invention having the above characteristics is extremely effective as an antitumor agent (anticancer agent) that exhibits an antitumor action more effectively.

Of the compounds (1) of the present invention, compounds (4) to (7) and (10) having an amino group, those having a guanidyl group (11) and (12), and pharmaceutically acceptable compounds thereof The salt has sufficient water solubility and can be prepared as a liquid such as an injection.

The compound of the present invention can be orally or non-humanly administered to humans or mammals other than humans in a known manner, in known unit dosages, and as a pharmaceutical composition together with known carriers or excipients or pharmaceutically acceptable additives. Administer orally (eg, intramuscular, subcutaneous, intravenous injection or infusion, transpulmonary administration), or topical (mucosal administration) or external preparation (suppository, ointment, cream, patch, etc.) Can do.

In the case of oral administration, the compound of the present invention is converted into a conventional pharmaceutical preparation, for example, a solid preparation such as tablet, capsule, granule, fine granule, powder, lozenge, troche, jelly; or solution, emulsion (water-in-oil type) Emulsions, etc.), suspensions, syrups, etc. To produce tablets or other solid preparations, one or more of the compounds of the present invention can be combined with conventional excipients (sodium citrate, lactose, microcrystalline cellulose, starch, etc.), lubricants (anhydrous silicic acid, Hydrogenated castor oil, magnesium stearate, sodium lauryl sulfate, talc, etc.), binders (starch paste, glucose, lactose, gum arabic, gelatin, mannitol, etc.) and other normal additives (flavors, colorants, antioxidants) Preservatives including surfactants, surfactants, suspending agents, emulsifiers and the like), and the mixture is formulated by a known method. For the production of the liquid preparation, a known liquid carrier such as water, physiological saline or oil is used.

For parenteral administration, the compound of the present invention is used as a sterile oily or aqueous preparation. Injections are usually prepared by dissolving the compound of the present invention in distilled water for injection and then buffering or isotonizing with glucose, saline, etc. if necessary. The external preparation is preferably an ointment, but includes liniments, lotions, oil-in-water or water-in-oil emulsions (for example, creams), solutions, suspensions, and the like. Ointments are prepared in a known manner by conventional ointment bases such as fats, fatty acids (olive oil, sesame oil, triglycerides of medium chain fatty acids, etc.), lanolin, wax, paraffin, glycols, higher alcohols, surfactants and the like. Can be manufactured together.

The dose of the compound or pharmaceutical composition of the present invention varies depending on the administration method, the sex and age of the patient, the degree of symptoms, etc., but when administered parenterally, such as subcutaneous injection, About 1 to 100 mg / kg, preferably about 3 to 40 mg / kg is used. When administered orally, although not limited, 3 to 300 mg / kg, preferably about 9 to 120 mg / kg, of the compound can be used per day for an adult.

The compound of the present invention may be administered alone or in combination with another antitumor agent (for example, interferon α). When the compound of the present invention is used in combination with another antitumor agent, the amount used is not particularly limited as long as the antitumor action of the other antitumor agent is enhanced, but preferably 100 parts by weight of the other antitumor agent 0.0001 to 500 parts by weight, more preferably 0.001 to 300 parts by weight.

Hereinafter, the present invention will be specifically described with reference to production examples and test examples, but the present invention is not limited thereto.

The compound of the present invention was produced based on the synthetic schemes shown in FIG. 1, FIG. 2 and FIG. The starting material ISIR-005 was synthesized according to the method described in Patent Document 3 (International Publication WO2008 / 010324).

Production Example 1 Production of Compound (3) (FIG. 1, ISIR-040)
(1) Synthesis of SI-01 198 mg (0.37 mmol) of ISIR-005 was dissolved in 4 mL of dichloromethane, and 181 mg (1.48 mmol) of dimethylaminopyridine (DMAP) and 60 μL (0.85 mmol) of ice were cooled. Acetyl chloride (AcCl) was added and stirred for 45 minutes. The reaction was terminated with saturated aqueous NH ₄ Cl, extracted with dichloromethane, and the organic layer was dried over MgSO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 1: 2) to quantitatively obtain 229 mg of SI-01. The ¹³ C-NMR data and ESI-MS data of SI-01 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.4, 18.9, 20.6, 20.8, 20.9, 21.0, 26.7, 26.8, 27.7, 27.9, 28.9, 35.8, 40.5, 40.8, 42.5, 48.4, 52.5, 58.6, 62.3, 63.9, 69.7, 71.7, 72.0, 76.2, 77.6, 78.4, 99.7, 100.0, 132.9, 135.0, 138.7, 149.8, 169.7, and 170.1.
ESI-MS: m / z 643.3434 (M + Na ⁺ ).

(2) Synthesis of SI-02 To SI-01 (215 mg (0.35 mmol)) synthesized in methanol (10 mL), 53 mg (0.21 mmol) of pyridinium p-toluenesulfonic acid (PPTS) was cooled with ice. ) Was added. After stirring at 50 ° C. for 2 hours, the mixture was diluted with saturated aqueous KHSO ₄ solution, extracted with dichloromethane, and the organic layer was washed with saturated NaHCO ₃ . The extract was dried over MgSO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 1: 1) to obtain 191 mg of SI-02.
Yield 94%.
The ¹³ C-NMR data and ESI-MS data of SI-02 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.4, 20.7, 20.8, 20.9, 21.3, 26.6, 26.9, 27.7, 28.0, 35.8, 40.7, 40.9, 42.4, 48.5, 52.4, 58.7, 61.6, 69.5, 70.6, 71.8, 73.6, 76.2, 77.9, 78.3, 99.5, 132.9, 135.2, 138.7, 149.7, 169.6, and 172.1.
ESI-MS: m / z 603.3165 (M + Na ⁺ ).

(3) Synthesis of SI-03 SI-02 (236 mg (0.41 mmol)) synthesized above was dissolved in 5 mL of dichloromethane, and 400 μL (2.44 mmol) triethylamine and 94 μL (1.22 mmol) were added under ice cooling. Methanesulfonyl chloride (MsCl) was added and stirred at room temperature for 16 hours. Saturated KHSO ₄ was added to the reaction solution, followed by extraction with dichloromethane. The organic layer was washed with saturated NaHCO ₃ and NaCl aqueous solution in this order, dried over MgSO ₄ , and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 2: 3) to obtain 255 mg of SI-03. Yield 85%.
The ¹³ C-NMR data and ESI-MS data of SI-03 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.4, 20.6, 20.8, 20.9, 21.2, 26.8, 26.9, 27.8, 28.0, 35.9, 37.6, 38.6, 40.6, 41.0, 42.3, 48.4, 52.4, 58.7, 65.8, 67.6, 69.2, 70.2, 73.3, 76.2, 77.8, 79.0, 99.0, 132.6, 135.3, 138.9, 150.0, 169.2, and 170.2.
ESI-MS: m / z 759.2691 (M + Na ⁺ ).

(4) Synthesis of SI-04 SI-03 (126 mg (0.17 mmol)) synthesized above was dissolved in 5 mL of methanol, a saturated aqueous NaHCO ₃ solution (0.5 mL) was added, and the mixture was stirred at room temperature for 4 hours. The mixture was extracted with dichloromethane, and the organic layer was washed with a saturated aqueous NaCl solution, dried over MgSO ₄ , and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 3: 1) to obtain 109 mg of SI-04. Yield 98%.
The ¹³ C-NMR data and ESI-MS data of SI-04 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.5, 20.5, 21.5, 26.8, 26.9, 27.9, 28.0, 29.7, 36.0, 37.5, 38.7, 40.6, 41.9, 42.1, 48.2, 52.4, 58.7, 66.7, 67.5, 71.9, 72.3, 76.2, 78.1, 78.2, 100.6, 132.8, 135.7, 138.8, and 149.4.
ESI-MS: m / z 675.2464 (M + Na ⁺ ).

(5) Synthesis of ISIR-040 (compound (3)) SI-04 (83 mg (0.13 mmol)) synthesized above was dissolved in tetrahydrofuran (THF: 2.0 mL), and NaOMe (0.5 M methanol was added under ice cooling. (Solution: 381 μL) was added and stirred for 24 hours under ice cooling. Dilute with saturated aqueous NH ₄ Cl and extract with ethyl acetate. The organic layer was dried over Na ₂ SO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 3: 2) to quantitatively obtain 66 mg of ISIR-040.
¹ H-NMR data and ESI-MS data of ISIR-040 are shown below.
¹ H-NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 7.0), 0.96 (3H, d, J = 7.0), 1.05 (3H, d, J = 7.0), 1.20 (3H, s), 1.47-1.60 (3H, m), 1.64-1.72 (2H, m), 1.75 (1H, m), 1.89-1.98 (2H.m), 2.09-2.13 (2H, m), 2.61 (1H, t, J = 7.0), 2.66 (1H, m), 2.79 (1H, m), 2.95 (1H, brs), 3.13 (1H, m, J = 7.0), 3.29 (2H, d, J = 8.1), 3.31 (3H , s), 3.35 (3H, s), 3.80 (1H, dd, J = 4.0, 10.3), 3.84 (1H, d, J = 10.3), 3.93 (1H, d, J = 10.3) 3.97 (1H, dd , J = 4.0, 10.3), 3.97 (1H, t, J = 4.0), 4.75 (1H, dd, J = 4.0, 7.3), 5.01 (1H, d, J = 0), 5.06 (1H, d, J = 4.0), and 5.31 (1H, t, J = 1.7).
ESI-MS: m / z 515 (M + Na ⁺ ).

Production Example 2 Production of Compound (4) (FIG. 1, ISIR-041)
(1) Synthesis of SI-05 Sodium hydride (7.8 mg, 0.20 mmol) was added to a solution of 2-azido-1-ethanol (601 μL, 7.94 mmol) in tetrahydrofuran (THF: 1.0 mL) under ice cooling. Then, a solution prepared by dissolving SI-04 (30 mg (0.046 mmol)) synthesized in Production Example 1 (4) in 2 mL of THF containing 30 μL of hexamethylphosphoric triamide (HMPA) was added. After stirring at room temperature for 24 hours, the mixture was diluted with saturated aqueous NH ₄ Cl and extracted with ethyl acetate. The organic layer was dried over MgSO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (dichloromethane: acetone: methanol = 100: 5: 1) to obtain 19.9 mg of SI-05. Yield 74%.
The ¹ H-NMR data of SI-05 is shown below.
¹ H-NMR (CDCl ₃ ): δ 0.83 (3H, d, J = 7.3 Hz), 0.97 (3H, d, J = 6.8 Hz), 1.06 (3H, d, J = 6.5 Hz), 1.20 (3H, s), 1.53 (1H, s), 1.63-1.81 (2H, m), 1.87-2.01 (2H, m), 2.12 (2H, dd, J = 4.0 Hz, 4.8 Hz), 2.18 (1H, s), 2.65 (1H, d, J = 9.9 Hz), 2.67 (2H, dd, J = 7.7, 7.7 Hz), 2.76-2.90 (2H, m), 3.13 (1H, quint), 3.30 (2H, d, J = 7.9 Hz), 3.35 (3H, s), 3.34-3.40 (3H, m), 3.60 (1H, quin), 3.76-3.89 (2H, m), 3.87 (1H, d, J = 2.2 Hz), 3.95 ( 1H, dd, J = 10.7, 11.0 Hz), 3.97 (1H, d, J = 3.9 Hz), 4.75-4.83 (1H, m), 5.07 (1H, d, J = 4.9 Hz), 5.16 (1H, s ), and 5.32 (1H, s).

(2) Synthesis of ISIR-041 (Compound (4 )) In a THF solution (1.0 mL) of SI-05 (19.9 mg (0.036 mmol)) synthesized in (1), THF of lithium aluminum hydride was added under ice cooling. The solution (180 μL: 0.36 mmol) was added and stirred for 4 hours. The inorganic compound was removed by filtration as it was, and the residue obtained by distilling off the solvent was directly separated and purified by silica gel column chromatography (ethanol: aqueous ammonia = 100: 1) to obtain 12.2 mg of ISIR-041 (compound (4)) was obtained. Yield 65%.
The ¹³ C-NMR data of ISIR-041 is shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.6,20.5, 21.3, 26., 27.7, 29.7, 31.4, 35.9, 36.5, 39.5, 40.4, 42.2, 42.8, 48.4, 52.3, 55.6, 58.7, 63.0, 71.6, 75.8 , 76.2, 77.2, 81.2, 105.6, 107.5, 132.6, 14.8, 138.4, and 150.3

Production Example 3 Production of Compound (5) (FIG. 1, ISIR-042)
(1) Synthesis of SI-06 SI-04 (53 mg (0.081 mmol)) synthesized in Production Example 1 (4) was synthesized in the same manner as the synthesis of SI-05 described in Production Example 2 (1). Reaction with azido-1-propanol gave 37.7 mg of SI-06. Yield 83%.
The ¹³ C-NMR data and ESI-MS data of SI-06 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.5, 20.3, 21.2, 26.5, 26.9, 27.7, 28.3, 28.9, 35.9, 40.6, 41.9, 42.6, 48.4, 48.5, 52.4,56.6, 58.7, 63.9, 71.2, 75.8, 76.3, 76.6, 77.8, 81.0, 104.7, 107.2, 132.8, 135.4, 138.5, and 149.8.
ESI-MS: m / z 584.3317 (M + Na ⁺ ).

(2) Synthesis of ISIR-042 (compound of formula (5)) In the same manner as ISIR-041 described in Production Example 2 (2), 48 mg (0.086 mmol) of SI-06 was replaced with lithium aluminum hydride. Reduction to obtain 31.9 mg of ISIR-042 (compound (5)). Yield 70%.
The ¹³ C-NMR data and ESI-MS data of ISIR-042 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.5, 20.1, 20.6, 26.1, 26.7, 26.8, 27.5, 27.8, 35.8, 35.8, 40.3, 42.6, 48.2, 52.0, 55.9, 58.5, 65.3, 71.4, 75.0, 76.2, 76.3, 77.4, 77.6, 81.2, 104.9, 107.0, 132.7, 134.6, 138.2, and 150.7.
ESI-MS: m / z 536.3571 (M + Na ⁺ ).

Production Example 4 Production of Compound (6) (FIG. 1, ISIR-043)
(1) Synthesis of SI-07 SI-04 (17 mg (0.026 mmol)) synthesized in Production Example 1 (4) was converted to 4-azido-, similarly to the synthesis of SI-05 in Production Example 2 (1). Reaction with 1-butanol gave 9.8 mg of SI-07. Yield 66%.
The ¹ H-NMR data of SI-07 is shown below.
¹ H-NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 7.3 Hz), 0.97 (3H, d, J = 6.9 Hz), 1.07 (3H, d, J = 7.0 Hz), 1.20 (3H, s), 1.45-1.60 (3H, m), 1.60-1.70 (6H, m), 1.70-1.79 (1H, m), 1.87-2.00 (2H, m), 2.12 (2H, dd, J = 3.4, 5.3 Hz), 2.61 (1H, t, J = 6.7 Hz), 2.66 (1H, quin), 2.79 (1H, dd, J = 6.2, 6.8 Hz), 3.13 (1H, quin), 3.29 (2H, d, J = 7.7 Hz), 3.29 (2H, t, J = 5.6 Hz), 3.35 (3H, s), 3.38-3.50 (1H, m), 3.65 (1H, dd, J = 4.9,6.5 Hz), 3.69 (1H , t, J = 6.2, 6.2 Hz), 3.76-3.87 (2H, m), 3.92-2-4.01 (3H, m), 4.76 (1H, dd, J = 3.9, 7.8 Hz), 5.07 (1H, d , J = 4.5 Hz), 5.10 (1H, s), and 5.32 (1H, s).

(2) Synthesis of ISIR-043 (Compound (6)) In the same manner as ISIR-041 described in Production Example 2 (2), 9.8 mg (0.017 mmol) of SI-07 was replaced with lithium aluminum hydride. Reduction gave 6.1 mg of ISIR-043 (compound (6)). Yield 65%.

Production Example 5 Production of Compound (7) (FIG. 1, ISIR-044)
(1) Synthesis of SI-08 SI-04 (19.5 mg (0.030 mmol)) synthesized in Production Example 1 (4) was converted to 4-azido-, similarly to the synthesis of SI-05 in Production Example 2 (1). Reaction with 1-pentanol gave 11.5 mg of SI-08. Yield 65%.
The ¹³ C-NMR data of SI-08 is shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.6, 20.3, 21.3, 23.5, 26.6, 26.9, 27.7, 28.2, 28.6, 29.0, 35.9, 40.6, 42.0, 42.6, 48.4, 51.3, 52.4, 56.4, 58.7, 67.0, 71.1, 75.9, 76.3, 76.6, 77.8, 81.1, 104.8, 107.1, 132.8, 135.4, 138.5, and 149.7.

(2) Synthesis of ISIR-044 (compound (7)) 11.5 mg (0.020 mmol) of SI-08 was reduced with lithium aluminum hydride in the same manner as ISIR-041 described in Production Example 2 (2). 8.5 mg of ISIR-044 (compound (7)) was obtained. Yield 75%.
The ¹ H-NMR data of ISIR-044 is shown below.
¹ H NMR (CDCl ₃ ): δ 0.80 (3H, d, J = 7.0 Hz), 0.97 (3H, d, J = 6.9 Hz), 1.07 (3H, d, J = 6.6 Hz), 1.20 (3H, s ), 1.38-1.92 (7H, m), 2.08-2.13 (2H, m), 2.60-2.72 (2H, m), 2.76-2.84 (3H, m), 3.03 (4H, t, J = 7.1 Hz), 3.24 (1H, t, J = 6.0 Hz), 3.28 (2H, d, J = 6.9 Hz), 3.34 (3H, s), 3.44 (3H, dd, J = 5.6, 6.0 Hz), 3.62-3.72 (4H , m), 3.82 (2H, s), 3.86 (1H, d, J = 10.6 Hz), 3.97-4.06 (3H, m), 4.80-4.84 (1H, m), 5.01 (1H, d, J = 4.4 Hz), 5.21 (1H, s), and 5.30 (1H, s).

Production Example 6 Production of Compound (5) Salt (Hydrochloride, Ascorbic Acid, Citrate, Benzoate) Compound (5) (ISIR-042) synthesized in Production Example 3 (2) was added to an anhydrous diethyl ether solution. By adding the equivalent amount of hydrogen chloride ether solution, ascorbic acid, citric acid or benzoic acid and diluting with petroleum ether, the resulting powder is filtered to obtain the corresponding hydrochloride salt and ascorbate salt of compound (5). Citrate or benzoate was obtained.

Production Example 7 Production of Compound (8) (Figure 2, ISIR-062)
Compound (5) (ISIR-042) (23.8 mg, 44 μmol) synthesized in Production Example 3 (2) is dissolved in 1.0 mL of dichloromethane, and 0.1 mL of pyridine and 8.3 μL (88 μmol) of acetic anhydride are dissolved under ice cooling. And then stirred at room temperature for 1.5 hours. Dilute with saturated aqueous NH ₄ Cl and extract with dichloromethane. The organic layer was washed successively with saturated NaHCO ₃ and aqueous NaCl solution, dried over Na ₂ SO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (methanol: chloroform = 1: 40) to obtain 12.6 mg of ISIR-062 (compound (8)). Yield 50%.
The ¹³ C-NMR data and ESI-MS data of ISIR-062 are shown below.
¹³ C-NMR (CDCl ₃ ): δ 9.6, 20.3, 21.3, 23.3, 26.6, 26.9, 27.8, 28.2, 29.3, 36.0, 37.4, 40.7, 42.0, 42.6, 48.4, 52.4, 56.6, 58.7, 60.0, 65.6, 71.3, 76.4, 76.6, 77.8, 81.0, 105.1, 107.7, 132.9, 135.7, 138.7, 149.6, and 170.1.
ESI-MS: m / z 600.3522 (M + Na ⁺ ).

Production Example 8 Production of Compound (9) (FIG. 2, ISIR-072)
Compound (5) (ISIR-042) (21.4 mg, 40 μmol) synthesized in Production Example 3 (2) was dissolved in 1.0 mL of dichloromethane and immobilized at 133 mg (equivalent to 0.2 mmol) of polystyrene resin at room temperature. N-cyclohexylcarbodiimide (PS-Carbodiimide) and 10 μL (0.160 mmol) of propiolic acid were added and stirred for 13.5 hours. The reaction solution was concentrated. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 1: 1) to obtain 10.0 mg of ISIR-072 (compound (9)). Yield 43%.
The ¹ H-NMR data of ISIR-072 is shown below.
¹ H-NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 7.3 Hz), 0.96 (3H, d, J = 6.8 Hz), 1.07 (3H, d, J = 6.8 Hz), 1.20 (3H, s), 1.26 (1H, s), 1.44-1.82 (8H, m), 1.87-2.00 (2H, m), 1.94 (3H, s), 2.03 (1H, d, J = 7.6 Hz), 2.12 (2H , dd, J = 4.6, 8.3 Hz), 2.64 (1H, t, J = 7.1 Hz), 2.64-2.71 (1H, m), 2.75-2.83 (1H, m), 3.15 (1H, quin), 3.30 ( 2H, d, J = 7.8 Hz), 3.35 (3H, s), 3.38-3.53 (2H, m), 3.67-3.76 (1H, m), 3.77-3.89 (2H, m), 3.92 (1H, d, J = 10.5 Hz), 3.95-4.01 (2H, m), 4.78 (1H, dd, J = 3.9, 6.8 Hz), 4.78 (1H, dd, J = 3.9, 6.8 Hz), 5.07 (1H, d, J = 4.2 Hz), 5.08 (1H, s), and 5.32 (1H, s).

Production Example 9 Production of Compound (10) (FIG. 2, ISIR-082)
(1) Synthesis of SI-09 To a solution of compound (5) (ISIR-042) (19 mg, 36 μmol) synthesized in Preparation Example 3 (2) in dichloromethane (1.0 mL), 10 μL of triethylamine and ice-cooled solution were added. 8.9 mg (40 μmol) of 4-nitrobenzenesulfonyl chloride was added and stirred for 2 hours. The reaction was diluted with water and extracted with dichloromethane. The organic layer was washed with a saturated aqueous NaCl solution, dried over Na ₂ SO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (chloroform: methanol = 30: 1) to obtain 22.7 mg of SI-09. Yield 88%.
The ¹ H-NMR data of SI-09 is shown below.
¹ H NMR (CDCl ₃ ): δ 0.83 (3H, d, J = 7.2 Hz), 0.98 (3H, d, J = 6.9 Hz), 1.08 (3H, d, J = 6.6 Hz), 1.20 (3H, s ), 1.52 (3H, t, J = 1.1 Hz), 1.63-1.80 (5H, m), 1.88-2.00 (2H, m), 2.05 (1H, s), 2.13 (2H, dd, J = 3.5, 4.9 Hz), 2.59 (1H, t, J = 7.0), 2.67 (1H, s), 2.77-2.83 (1H, m), 3.09-3.20 (4H, m), 3.29 (2H, d, J = 7.5 Hz) , 3.35 (3H, s), 3.38-3.47 (1H, m), 3.66-3.77 (2H, m), 3.85 (1H, d, J = 10.4 Hz), 3.93 (1H, d, J = 9.9 Hz), 3.95-4.03 (2H, m), 4.77 (1H, dd, J = 4.0, 7.3 Hz), 5.05 (1H, s), 5.07 (1H, d, J = 4.8 Hz), 5.32 (1H, s), 8.04 (2H, d, J = 9.0 Hz), and 9.03 (2H, d, J = 9.0 Hz).

(2) Synthesis of SI-10 Dissolve 7.5 mg (10 μmol) of SI-09 in 0.9 mL of dimethylformamide, and under ice cooling, add 2.8 mg of potassium carbonate and 0.7 μL (11 μmol) of methyl iodide. The mixture was further stirred for 4.5 hours. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic layer was washed successively with saturated NaHCO ₃ and aqueous NaCl solution, dried over Na ₂ SO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (ethyl acetate: hexane = 1: 1) to obtain 7.3 mg of SI-10. Yield 99%.
The ¹ H-NMR data of SI-10 is shown below.
¹ H-NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 7.1 Hz), 0.97 (3H, d, J = 6.7 Hz), 1.07 (3H, d, J = 6.7 Hz), 1.20 (3H, s), 1.51 (7H, t, J = 1.0 Hz), 1.60-1.84 (5H, m), 1.88-2.00 (2H, m), 2.05 (1H, s), 2.12 (2H, dd, J = 4.3, 8.3 Hz), 2.64 (2H, t, J = 6.5), 2.78 (4H, s), 2.98-3.27 (1H, m), 3.29 (2H, d, J = 8.1 Hz), 3.35 (3H, s), 3.40-3.48 (1H, m), 3.68-3.78 (1H, m), 3.80-3.89 (2H, m), 3.92 (1H, d, J = 10.0 Hz), 3.94-4.03 (2H, m), 4.79 ( 1H, dd, J = 4.2, 8.2 Hz), 5.07 (1H, d, J = 4.1 Hz), 5.12 (1H, s), 5.32 (1H, s), 7.97 (2H, d, J = 8.7 Hz), and 8.38 (2H, d, J = 8.9 Hz).

(3) Synthesis of ISIR-082 (Compound (10)) 7.3 mg (10 μmol) of the above SI-10 was dissolved in 0.7 mL of dimethylformamide, and ice-cooled with 4.1 mg of potassium carbonate and 1.2 μL (12 μmol). After adding thiophenol, the mixture was stirred at 60 ° C. for 48 hours. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic layer was washed with a saturated aqueous NaCl solution, dried over Na ₂ SO ₄ and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (methanol: chloroform = 1: 40) to obtain 3.8 mg of ISIR-082 (compound (10)). Yield 69%.

Production Example 10 Production of Compound (11) (FIG. 3, ISIR-051)
(1) Synthesis of SI-11 Reagent A (FIG. 3: reagent A) (20 mg, 20 mg, synthesized separately in compound (4) (ISIR-041) (18.0 mg, 34 μmol) synthesized in Production Example 2 (2) 54 μmol) of THF solution was added, and the mixture was stirred at room temperature for 22 hours and concentrated under reduced pressure. The reaction was diluted with water and extracted with dichloromethane. The crude product was separated and purified by silica gel chromatography (ethyl acetate: hexane = 1: 2) to obtain 28.0 mg of SI-11. Yield 99%.

The ¹ H-NMR and ¹³ C-NMR data of SI-11 are shown below.
¹ H-NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 6.8 Hz), 0.97 (3H, d, J = 7.0 Hz), 1.08 (3H, d, J = 6.5 Hz), 1.21 (3H, s), 1.46-1.61 (4H, m), 1.65-1.72 (1H, m), 1.73-1.79 (1H, m), 1.89-2.00 (3H, m), 2.12 (2H, q, J = 4.9, 8.6 Hz), 2.62-2.68 (1H, m), 2.69 (1H, t, J = 7.0), 2.80 (1H, t, J = 6.0 Hz), 3.17 (1H, quin), 3.30 (2H, d, J = 7.5 Hz), 3.35 (3H, s), 3.50-3.68 (4H, m), 3.69-3.75 (1H, m), 3.83 (2H, d, J = 2.3), 3.92 (1H, d, J = 9.9 Hz ), 3.97 (2H, ddd, J = 4.1, 10.8, 17.9 Hz), 4.79 (1H, tt), 5.04 (1H, d, J = 4.2), 5.12 (3H, s), 5.17 (2H, d, J = 0.9), 5.32 (1H, s), 7.29 (2H, d, J = 7.1 Hz), 7.33 (2H, t, J = 7.1 Hz), 7.37 (6H, quin), and 8.55 (1H, t, J = 4.7 Hz).
¹³ C-NMR (CDCl ₃ ): δ 9.5, 14.1, 20.3, 21.0, 21.2, 26.6, 26.9, 27.7, 28.1, 35.9, 40.6, 40.8, 41.9, 42.6, 48.4, 52.3, 56.3, 58.6, 60.4, 65.2, 67.1, 68.1, 71.4, 76.2, 76.3, 76.4, 77.7, 80.8, 105.0, 107.2, 108.1, 127.9, 128.1, 128.4, 128.4, 128.6, 128.8, 132.8, 134.6, 135.4, 136.7, 138.5, 149.7, 153.7, 156.0, and 163.6.

(2) Synthesis of ISIR-051 The above SI-11 (28.0 mg, 34 μmol) was dissolved in 1.0 ml of methanol, 39.6 mg of palladium carbon was added, and the mixture was stirred under a hydrogen atmosphere for 2 hours. The crude product was filtered and concentrated under reduced pressure to obtain 15.6 mg of ISIR-051. Yield 81%.

The ¹ H-NMR data of ISIR-051 is shown below.
¹ H-NMR (CDCl ₃ : shows only characteristic signal): δ 0.78 (3H, d), 0.95 (3H, d), 1.06 (3H, d), 1.19 (3H, s), 3.34 (3H, s ), 4.99 (1H, d), 5.28 (1H, s), 5.29 (1H, s), 5.69 (1H, br), 7.04 (2H, br), and 7.66 (1H, br).

Production Example 11 Production of Compound (12) (FIG. 3, ISIR-052)
ISIR-052 was synthesized via SI-12 by the same method as in Production Example 10 using Compound (5) (ISIR-042) (76.0 mg, 14.2 μmol) synthesized in Production Example 3 (2). Two-stage yield 65%.
The ¹ H-NMR data of ISIR-052 is shown below.
¹ H-NMR (CDCl ₃ : shows only characteristic signal): δ 0.78 (3H, d), 0.94 (3H, d), 1.05 (3H, d), 1.19 (3H, s), 3.34 (3H, s ), 4.95 (1H, d), 5.10 (1H, s), 6.36 (1H, t), and 7.62 (2H, d).

Test Example 1 Measurement of cell differentiation inducing activity
(1) Cells and cell culture Suspend human acute myeloid leukemia cells (HL-60 cells) or human monocytic leukemia cells (U937 cells) in RPMI-1640 medium supplemented with 10% fetal bovine serum. The cells were cultured in humidified air at 37 ° C. containing 5% CO ₂ .

(2) Measurement of differentiated cells In order to measure differentiation-inducing activity, leukemia cells (5 × 10 ⁴ cells / mL) were added to test compounds (ISIR-040 (compound (3)), ISIR-042 ( Compound (5)), ISIR-062 (Compound (8)), ISIR-072 (Compound (9)), ISIR-082 (Compound (10), ISIR-005, Cotyrenin A (CN-A)) Alternatively, the cells were cultured in the absence of 12-O-tetradecanoylphorbol-13-acetate (TPA) as a promoter, and the oxidase activity to generate superoxide was measured using the ability to reduce NBT as an indicator. NBT reducing ability was measured by incubating the cells in RPMI-1640 medium containing NBT (1 mg / mL) and TPA (100 ng / ml) for 60 minutes at 37 ° C. After the reaction was completed, the cells were centrifuged. The formazan precipitate generated by the reaction in the cells was dissolved in dimethyl sulfoxide and colorimetric (wavelength 560 nm or The activity was expressed as absorbance per 10 ⁷ cells.

FIG. 4 shows the evaluation results of ISIR-040 (compound (3)) on HL-60 cells, together with the results of cotylenin A (CN-A). FIG. 5 shows the evaluation of ISIR-040 (compound (3)), ISIR-042 (compound (5)), ISIR-062 (compound (8)), and ISIR-082 (compound (9)) on U937 cells. The results are shown together with the results of CN-A and ISIR-005. FIG. 6 shows the evaluation results of ISIR-041 (compound (3)), ISIR-042 (compound (5)), ISIR-043 (compound (6)), and ISIR-044 (compound (7)) on U937 cells. Is shown together with the result of ISIR-005. FIG. 7 shows the evaluation results of ISIR-042 (compound (5)) and ISIR-072 (compound (9)) on U937 cells.

As can be seen from these results, the differentiation-inducing activity of all the compounds targeted by the present invention was stronger than that of Cochirenin A. In addition, ISIR-041 (compound (4)), ISIR-042 (compound (5)), ISIR-072 (compound (9)), and ISIR-082 (compound (10)) are described in Patent Document 3. It showed stronger cell differentiation-inducing activity than ISIR-005.

Test Example 2 Inhibitory effect on cancer cell proliferation by combined use of ISIR-042 and interferon α The cell growth inhibitory effect obtained by the combined use of the compound of the present invention and interferon α (IFNα) was measured. Lung cancer cells (A549) were used as test cells, and these cells were precultured in RPMI 1640 medium. In this cell solution (1 × 10 ⁴ cells / mL), ISIR-042 at a predetermined concentration (0, 3, 4, 5, 6, 7 μg / mL) and a predetermined concentration (0, 100, 200, 400 units / mL) ) IFNα was added and cultured for 7 days to evaluate the number of viable cells. The number of viable cells was evaluated by MTT assay. A PBS solution (1 mg / mL) of MTT was added and incubated for 4 hours. After centrifugation, the precipitate was dissolved in 1 mL of DMSO, and the absorbance at 560 nm was measured. The measurement results are shown in FIG.

The concentration required to inhibit the growth of lung cancer cell A549 cells by 50% is calculated with ISIR-005 alone and in combination with 400 units / mL interferon α. In the former case, the concentration is> 7μg / mL or more. In contrast, the latter was 4.5 μg / mL, indicating a significant synergistic effect.

In addition, when the same evaluation was performed on ISIR-041, ISIR-043, and ISIR-044, the same synergistic effect was confirmed in all cases, but the degree did not reach ISIR-042.

Test Example 3 Inhibitory effect of ISIR-042 (compound (5)) on cancer cell growth under hypoxia (Hypoxia) The cell growth inhibitory action of the compound of the present invention was reduced to normal oxygen concentration (21%) and hypoxia (1%) The measurement was performed under the following conditions. Breast cancer cells (MCF-7) were used as test cells, and these cells were precultured in RPMI 1640 medium. To this cell solution (1 × 10 ⁴ cells / mL), ISIR-042 at a predetermined concentration (0, 0.5, 1.0, 1.5, 2.0, 2.5 μg / mL) was added, cultured for 7 days, and the number of viable cells was evaluated. . The number of viable cells was evaluated by MTT assay. A PBS solution (1 mg / mL) of MTT was added and incubated for 4 hours. After centrifugation, the precipitate was dissolved in 1 mL of DMSO, and the absorbance at 560 nm was measured. FIG. 9 shows the measurement results of ISIR-042 together with the results of CN-A and ISIR-005. The concentration required to inhibit the growth of breast cancer cell MCF-7 by 50% is 1.0 μg / mL for ISIR-042 when measured under hypoxia, while 2.5 μg / mL under normal oxygen concentration. A much higher concentration was required than mL. These effects were not observed with CN-A or ISIR-005. In addition, in the case of gemcitabine, which is a known anticancer agent, the growth inhibitory effect was rather lowered under hypoxia (see FIG. 10 b).

Similar effects were observed in cancer cells other than breast cancer cells (MCF-7), such as pancreatic cancer cells (Panc-1, MiaPaca-2), lung cancer cells (A549, PC14), and ovarian cancer cells (SK-OV3). It was. FIG. 11 shows the results of comparing the effects of ISIR-042 (compound (5)) with the effects of ISIR-005 and CN-A on pancreatic cancer cells (MiaPaca-2), measured in the same manner as described above.

As described above, it has been clarified that ISIR-042 (compound (5)) has an unprecedented unique property of significantly suppressing the growth of cancer cells under hypoxia. FIG. 12 shows the effects of ISIR-042 (compound (5)) and ISIR-041 (compound (4)) on pancreatic cancer cells (Panc-1). As can be seen, ISIR-041 (compound (4)) had the same properties as ISIR-042 (compound (5)), although the effect was slightly inferior.

The compound (1) of the present invention is useful as an active ingredient of pharmaceuticals such as cell differentiation inducers and anticancer agents, and in the fields of fusicoccin derivatives and intermediates of various pharmaceuticals.

Claims (12)

1. Compound represented by the general formula (1) or a salt thereof:
   

   

   (Wherein n is an integer of 0 to 5, Me is a methyl group, R ¹ is an alkyl group having 1 to 6 carbon atoms, —N ₃ group or an amino group represented by the following formula:
   

   

   [Wherein R ² is a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; R ³ is a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, or a p-nitrophenylsulfonyl group. Alternatively, it represents an amidyl group (—CNHNH ₂ ). ]
   Indicates. )
2. In general formula (1),
   n is 0 and R ¹ is a methyl group, or
   n is an integer of 2 to 5, and R ¹ is an amino group represented by the following formula (2):
   

   

   [Wherein R ² represents a hydrogen atom; R ³ represents a hydrogen atom, an alkyl group having 1 to 6 carbon atoms, an acyl group having 1 to 7 carbon atoms, or an amidyl group. ]
   The compound or its salt of Claim 1 which is these.
3. 2. The compound according to claim 1, wherein the compound represented by the general formula (1) is a compound represented by any one of the following formulas (3) to (12):
   

   
4. A pharmaceutical composition comprising the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof as an active ingredient.
5. A cell differentiation inducer comprising the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof as an active ingredient.
6. An antitumor agent against hematological malignancy or solid cancer comprising the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof as an active ingredient.
7. The hematological malignancy is any selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma, and the solid cancer is any selected from the group consisting of breast cancer, pancreatic cancer, lung cancer and ovarian cancer, The antitumor agent according to claim 6.
8. A pharmaceutical comprising a combination of the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt and interferon α.
9. A pharmaceutical product according to claim 8, which is a composition comprising the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt and interferon α.
10. A combination kit in which the drug containing the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt and the drug containing interferon α as an active ingredient are packaged separately. 8. The pharmaceutical product according to 8.
11. The pharmaceutical product according to any one of claims 8 to 10, which is an antitumor agent.
12. The target tumor is any hematological malignancy selected from the group consisting of leukemia, malignant lymphoma and malignant myeloma, or any one selected from the group consisting of breast cancer, pancreatic cancer, lung cancer and ovarian cancer The pharmaceutical product according to claim 11, which is a solid cancer.
    

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