WO2019230669A1 - Composition pharmaceutique ciblant une séquence de liaison au runx, et inhibiteur du runx - Google Patents

Composition pharmaceutique ciblant une séquence de liaison au runx, et inhibiteur du runx Download PDF

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WO2019230669A1
WO2019230669A1 PCT/JP2019/020961 JP2019020961W WO2019230669A1 WO 2019230669 A1 WO2019230669 A1 WO 2019230669A1 JP 2019020961 W JP2019020961 W JP 2019020961W WO 2019230669 A1 WO2019230669 A1 WO 2019230669A1
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coh
alkyl group
runx
pyrrole
polyamide
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Japanese (ja)
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弘 杉山
靖彦 上久保
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国立大学法人京都大学
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/74—Synthetic polymeric materials
    • A61K31/785—Polymers containing nitrogen
    • A61K31/787—Polymers containing nitrogen containing heterocyclic rings having nitrogen as a ring hetero atom
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing three or more hetero rings

Definitions

  • the present disclosure relates to an anti-tumor composition, a RUNX inhibitor, and an anti-allergy composition comprising a pyrrole-imidazole polyamide targeting a RUNX binding sequence.
  • RUNX Runt-related transcription factor
  • RUNX family members include RUNX1, RUNX2, and RUNX3.
  • RUNX1 is known to be involved in secondary hematopoiesis
  • RUNX2 is related to bone development
  • RUNX3 is known to be involved in neurogenesis, thymocyte proliferation, and the like.
  • Each member of the RUNX family forms a heterodimeric complex with the core binding factor ⁇ subunit (CBF ⁇ ).
  • CBF ⁇ core binding factor ⁇ subunit
  • RUNX recognizes and binds the consensus binding sequence 5'-TGTGGT-3 ', and very rarely 5'-TGCGGT-3', in the transcriptional regulatory region of the target gene via the runt domain. , Control the expression of the target gene.
  • CBF ⁇ is a non-DNA binding regulatory subunit. CBF ⁇ enhances DNA binding ability of RUNX allosterically.
  • RUNX1 is also known as acute myeloid leukemia 1 protein (AML1) and is considered a tumor suppressor in the onset of leukemia.
  • AML1 acute myeloid leukemia 1 protein
  • recent studies suggest that RUNX1 has tumorigenic properties in the development of acute myeloid leukemia (AML), and research on low molecular weight compounds that inhibit the binding between RUNX1 and CBF ⁇ for the purpose of treating leukemia. has been reported (Non-patent Document 1). However, until now, no attempt has been reported to target the RUNX family binding site on genomic DNA for the treatment of various cancers including leukemia.
  • pyrrole-imidazole polyamide (hereinafter abbreviated as “PI polyamide”) is a synthetic oligomer, and is identified by binding in the minor groove of DNA by a pyrrole (P) and imidazole (I) pair facing each other by hairpin linking. It is known to recognize the sequence of PI polyamides recognize P / I pairs as C ⁇ G base pairs, P / P pairs as A ⁇ T or T ⁇ A base pairs, and I / P pairs as G ⁇ C base pairs, thereby It can specifically bind to a double-stranded DNA sequence. Thus, designing different orders of P / I pairs allows in vivo delivery of PI polyamides to target sites on the desired genome.
  • PI polyamide is permeable to the cell membrane, is localized in the cell nucleus, and affects endogenous gene expression at the nanomolar level.
  • PI polyamides bound to target genes have been studied as gene switches that inhibit transcription factor binding to DNA and control specific gene expression.
  • recent studies by the inventors have reported a PI polyamide targeting the RUNX family, a conjugate of the PI polyamide with a compound such as an alkylating agent, and an antitumor agent using the conjugate ( Patent Document 1).
  • the present disclosure aims to develop a pharmaceutical composition having excellent efficacy, particularly an antitumor composition and an antiallergic composition, and a RUNX inhibitor targeting the RUNX binding sequence.
  • the inventors surprisingly replaced at least one pyrrole unit constituting PI polyamide with ⁇ -alanine when synthesizing PI polyamide targeting the RUNX consensus binding site on the genome.
  • the binding property to the target sequence is improved.
  • complex of this PI polyamide and an alkylating agent has the inhibitory effect with respect to the tumor originating in various organs.
  • An antitumor composition comprising a pyrrole-imidazole polyamide capable of binding to a RUNX binding sequence, wherein at least one of the pyrrole units constituting the pyrrole-imidazole polyamide is substituted with ⁇ -alanine
  • the antitumor composition according to [1] comprising a complex of the pyrrole-imidazole polyamide and an agent, [3]
  • the complex is represented by the formula I: [In Formula I, X 1 represents CH, COH or N, X 2 represents CH, COH or N, X 3 represents CH, COH or N, X 4 represents CH, COH or N, X 5 represents CH, COH or N Or N, X 6 represents CH, COH or N, X 7 represents CH, COH or N, X 8 represents CH, COH or N, R 1 represents H or an alkyl group, R 2 represents H or an alkyl group, R 3
  • each of Z in the plurality of Ym-Z may be the same as or different from the other Z, and each of them may be independently selected.
  • the following formula: (In the formula, X is any one of X 1 to X 8 and represents CH or COH, and R ′ is any one of R 1 to R 8 and represents H or an alkyl group.)
  • the complex is represented by formula II: And Formula III: [In Formula II and Formula III, X 1 represents CH, COH or N, X 2 represents CH, COH or N, X 3 represents CH, COH or N, X 4 represents CH, COH or N, X 5 represents CH, COH or N Or N, X 6 represents CH, COH or N, X 7 represents CH, COH or N, X
  • the complex has the formula: The antitumor composition according to [7], which is represented by [9] The antitumor according to any one of [1] to [8], wherein the pyrrole-imidazole polyamide binds to a RUNX binding sequence on DNA and inhibits binding of a RUNX family member to the binding sequence.
  • Composition [10] The antitumor composition according to [9], which inhibits the binding of all members of the RUNX family to the RUNX binding sequence.
  • a RUNX inhibitor comprising a pyrrole-imidazole polyamide capable of binding to a RUNX binding sequence, wherein at least one of the pyrrole units constituting the pyrrole-imidazole polyamide is substituted with ⁇ -alanine
  • An antiallergic composition comprising a pyrrole-imidazole polyamide capable of binding to a RUNX binding sequence, wherein at least one of the pyrrole units constituting
  • the present disclosure provides a pharmaceutical composition, particularly an antitumor composition and an antiallergic composition, and a RUNX inhibitor having superior efficacy by utilizing PI polyamide having excellent binding properties to a target.
  • a PI polyamide designed to recognize a RUNX binding sequence on DNA, wherein at least one pyrrole unit of a pyrrole-imidazole pair constituting the PI polyamide is substituted with ⁇ -alanine.
  • polyamide an antitumor composition and a RUNX inhibitor having excellent efficacy can be obtained.
  • the antitumor composition has an antitumor effect against various types of cancer including leukemia.
  • the anti-tumor composition is also effective against tumors that are resistant to other molecular targeted therapeutics.
  • the RUNX inhibitor can also be used as an antiallergic composition.
  • a complex comprising PI polyamide has been shown to have a significantly higher binding affinity for the target sequence than a complex comprising PI polyamide in which P is not substituted with ⁇ -alanine, and further has a strong anti-resistance. It was shown to exert a tumor effect.
  • FIG. 1A shows the SPR sensorgram of the Chb-M ′ hydroxy substituent of Example 2. From above, SPR sensorgrams of hydroxy-substituted Chb-M 'at concentrations of 1250, 625.0, 312.5 and 156.3 nM are shown.
  • FIG. 1B shows the SPR sensorgram of the Chb-M hydroxy-substituted product of Example 2. From the top, SPR sensorgrams of hydroxy substitution of Chb-M at concentrations of 625.0, 312.5, 156.3, 78.13 and 39.06 nM are shown.
  • FIG. 2A shows the results of PAGE analysis for the upper strand carried out in Example 3.
  • FIG. 2B shows the results of PAGE analysis for the lower strand carried out in Example 3.
  • FIG. 2C shows a schematic diagram of the alkylation site of each compound used in Example 3.
  • the arrow indicates the alkylation site corresponding to the site shown in FIGS. 3A and B.
  • FIG. 3 is a graph showing the antitumor effect of Chb-M on the medulloblastoma cell line Daoy of Example 5.
  • the horizontal axis indicates the drug concentration ( ⁇ M), and the vertical axis indicates the cell viability (%).
  • FIG. 4A is a graph showing the antitumor effect of Chb-M on malignant glioma strain A172 of Example 5.
  • the horizontal axis indicates the drug concentration ( ⁇ M), and the vertical axis indicates the cell viability (%).
  • FIG. 4B is a graph showing the antitumor effect of Chb-M on malignant glioma strain KALS-1 of Example 4.
  • the horizontal axis indicates the drug concentration ( ⁇ M), and the vertical axis indicates the cell viability (%).
  • FIG. 5 is a graph showing the antitumor effect of Chb-M on the malignant rhabdoid tumor lines MP-MRT-AN and KP-MRT-NS of Example 5.
  • FIG. 6 is a graph showing the antitumor effect of Chb-M on prostate cancer strain DU145 of Example 6.
  • the horizontal axis indicates the drug concentration ( ⁇ M), and the vertical axis indicates the cell viability (%).
  • FIG. 7 is a graph showing the antitumor effect of Chb-M against leukemia strains MOLM13, MV4-11 (p53 wild type) and MV4-11NR (p53 mutant type) of Example 7, and a table showing IC 50 values.
  • the horizontal axis of the graph indicates the drug concentration ( ⁇ M), and the vertical axis indicates the cell viability (%).
  • FIG. 8 is a graph showing the antitumor effect of Chb-M against leukemia strain NB of Example 7 and a table showing IC 50 values.
  • FIG. 9 is a graph showing the antitumor effect of Chb-M against osteosarcoma strain MG63 of Example 8.
  • the horizontal axis indicates the drug concentration ( ⁇ M), and the vertical axis indicates the
  • PI polyamide PI polyamide is generally a polyamide containing an N-methylpyrrole unit (P), an N-methylimidazole unit (I), and a ⁇ -aminobutyric acid moiety. Linked to each other by an amide bond (—C ( ⁇ O) —NH—) (Trauger et al, Nature, 382, 559-61 (1996); White et al, Chem. Biol., 4,569-78 (1997); And Dervan, Bioorg. Med. Chem., 9, 2215-35 (2001)).
  • PI polyamide a ⁇ -aminobutyric acid moiety becomes a linker (hereinafter referred to as “ ⁇ -linker”) and the whole is folded into a U-shaped conformation (hairpin type).
  • ⁇ -linker a linker
  • two chains containing P and I are arranged in parallel across the ⁇ -linker.
  • pyrrole-imidazole pair the pair consisting of P and / or I of the two chains facing each other
  • P PI polyamides bind with high affinity to specific base pairs in DNA.
  • a P / I pair can bind to a C ⁇ G base pair and an I / P pair can bind to a G ⁇ C base pair.
  • the P / P pair can bind to both A ⁇ T base pairs and T ⁇ A base pairs.
  • PI polyamide may contain 3-hydroxypyrrole (Hp) or ⁇ -alanine, and P can be replaced with Hp or ⁇ -alanine.
  • Hp / P pairs can bind to TA base pairs (White at al., Nature, 391, 468-71 (1998)).
  • a ⁇ -alanine / ⁇ -alanine pair can bind to a T • A base pair or an A • T base pair.
  • a ⁇ -alanine / I pair can bind to a CG base pair, and an I / ⁇ alanine pair can bind to a G ⁇ C base pair.
  • the ⁇ -alanine / P pair and the P / ⁇ -alanine pair can bind to a T • A base pair or an A • T base pair.
  • the ⁇ -linker moiety and the ⁇ alanine moiety added to the N-terminus of PI polyamide can also be bound to T • A base pairs or A • T base pairs.
  • the methyl group on the 1-position nitrogen of P and I constituting the PI polyamide may be replaced with hydrogen or an alkyl group other than the methyl group.
  • the alkyl group other than the methyl group include a linear, branched or cyclic saturated or unsaturated alkyl group having 2 to 10 carbon atoms, preferably a linear or branched chain having 2 to 5 carbon atoms.
  • Examples include linear, branched, or cyclic saturated or unsaturated alkyl groups such as ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, and the like.
  • alkyl groups including methyl groups may be substituted.
  • methylene in the alkyl group may be substituted with oxygen or the like.
  • the 3-position of P constituting the PI polyamide may be substituted with a hydroxy group.
  • P or “pyrrole unit” and “I” or “imidazole unit” refer to N-substituted or N-unsubstituted pyrrole units, 3-hydroxypyrrole units, and N-substituted units as described above. Or N unsubstituted imidazole units.
  • the ⁇ -linker constituting the PI polyamide may have a side chain at the ⁇ -position or the ⁇ -position.
  • a ⁇ -linker in which the ⁇ -position or ⁇ -position is substituted with an amino group that is, a linker consisting of an N- ⁇ -N- ⁇ -diaminobutyric acid residue or an N- ⁇ -N- ⁇ -diaminobutyric acid residue.
  • the amino group may be modified with a molecule such as a fluorescent group or biotin.
  • various functional groups may be added to the N-terminal and C-terminal of the PI polyamide.
  • Those skilled in the art can appropriately determine functional groups or molecules to be added to the N-terminal and C-terminal of PI polyamide.
  • various functional groups can be added via an amide bond.
  • Examples of such functional groups include, but are not limited to, carboxyl groups such as ⁇ -alanine residues and ⁇ -aminobutyric acid residues, acetyl groups, amino groups, and the like.
  • an acetyl group may be added to the N-terminus.
  • a dimethylaminopropylamino group may be added to the C-terminus.
  • the N terminal and C terminal of PI polyamide may be modified with molecules such as a fluorescent group, biotin, and isophthalic acid.
  • the fluorescent group is not limited, and examples thereof include fluorescein, rhodamine dyes, cyanine dyes, ATTO dyes, Alexa Fluor dyes, and BODIPY.
  • Fluorescein also includes fluorescein derivatives (eg, fluorescein isothiocyanate).
  • the design method and production method of PI polyamide are known (see, for example, Japanese Patent No. 3045706, Japanese Patent Laid-Open No. 2001-136974, WO 03/000683, Japanese Patent Laid-Open No. 2013-234135, Japanese Patent Laid-Open No. 2014-173032).
  • it can be easily produced by an automatic synthesis method using a solid phase synthesis method (Fmoc solid phase synthesis method) using Fmoc (9-fluorenylmethoxycarbonyl). It can also be produced by a liquid phase synthesis method.
  • the PI polyamide may be a modified PI polyamide that is modified so as to maintain or improve the binding ability to DNA.
  • the modified PI polyamide include, for example, a modified product in which an amino group is added to the ⁇ -position or ⁇ -position of the ⁇ -linker of PI polyamide, that is, N- ⁇ -N- ⁇ -diaminobutyric acid residue or N- ⁇ - A modified product having a ⁇ -linker composed of N- ⁇ -diaminobutyric acid residue, a modified product obtained by modifying the amino group of the modified product with a molecule such as a fluorescent group or biotin, a fluorescent group or biotin at the N-terminus of PI polyamide, etc. And a modified product obtained by modifying the C-terminus of PI polyamide with a molecule such as isophthalic acid.
  • the PI polyamide used in the present disclosure is a PI polyamide designed to recognize a consensus sequence of the RUNX binding site on the genome (also referred to as “RUNX consensus binding sequence” or “RUNX binding sequence”), and therefore A PI polyamide capable of binding to the RUNX consensus binding sequence.
  • the RUNX consensus sequence is known to be 5'-TGTGGT-3 'or 5'-TGCGGT-3'. Therefore, the combination of the pyrrole-imidazole pair constituting the PI polyamide may be determined based on the RUNX consensus binding sequence.
  • the term “pyrrole-imidazole pair” includes a pair consisting of any combination of P, I, Hp, and ⁇ -alanine.
  • the PI polyamide used in the present disclosure comprises a pyrrole-imidazole pair containing at least one ⁇ -alanine. That is, in the PI polyamide used in the present disclosure, at least one of the pyrrole units is substituted with ⁇ -alanine.
  • ⁇ -alanine is represented by the following formula: Represented by
  • the number of ⁇ -alanine contained in the PI polyamide used in the present disclosure is not particularly limited as long as it is 1 or more. Depending on the total number of P and I constituting the PI polyamide, for example, 1 to 3, preferably 1 or 2, ⁇ -alanine is included.
  • the position of ⁇ -alanine contained in the PI polyamide used in the present invention is a ⁇ linker, a position adjacent to the C-terminal or N-terminal, or a ⁇ -linker
  • the position may be near the C-terminal or N-terminal, or may be other positions, for example, near or in the middle between the ⁇ linker and the C-terminal or N-terminal. Preferably, it is near or in the middle between the ⁇ linker and the C-terminal or N-terminal.
  • the PI polyamide in the present disclosure has high affinity for the target RUNX binding sequence.
  • the PI polyamide has a higher affinity for the targeted RUNX binding sequence compared to PI polyamide in which the pyrrole unit is not replaced by ⁇ -alanine.
  • the PI polyamides in this disclosure can compete with members of the RUNX family for binding to RUNX binding sequences on DNA and inhibit RUNX family binding to RUNX binding sequences on the genome.
  • the PI polyamide that recognizes the RUNX binding sequence may form a complex with the agent.
  • the “agent” is a substance that affects DNA and the surrounding chromatin state, and examples thereof include, but are not limited to, an alkylating agent, a chromatin-modifying enzyme control agent, and the like.
  • chromatin-modifying enzyme regulators include, but are not limited to, histone acetylase (HAT) regulators, such as HAT inhibitors (eg, C646), or HAT activators (eg, N- ( 4-chloro-3- (trifluoromethyl) phenyl) -2-ethoxybenzamide (CTB), etc., histone deacetylase (HDAC) regulator, eg, HDAC inhibitor (eg, suberoylanilide hydroxamic acid, etc.) Or an HDAC activator, a histone methylase regulator, a histone demethylase regulator, and the like.
  • HAT histone acetylase
  • HAT inhibitors eg, C646
  • HAT activators eg, N- ( 4-chloro-3- (trifluoromethyl) phenyl) -2-ethoxybenzamide (CTB), etc.
  • HDAC histone deacetylase
  • HDAC histone deacetylase
  • alkylating agent is a compound having a functional group that forms a covalent bond with DNA. Although it does not specifically limit as an alkylating agent used in this indication, A thing with low or no cytotoxicity is preferable in consideration of the use in the pharmaceutical composition mentioned later.
  • alkylating agents include, but are not limited to, chlorambucil, duocarmycin, seco-CBI (1-chloromethyl-5-hydroxy-1,2-dihydro-3H-benzo [ e] indole), CBI (1,2,9,9a-tetrahydrocyclopropane [c] benzo [e] indole-4-one), pyrrolobenzodiazepine, nitrogen mustard and the like.
  • an indole may be inserted between PI polyamide and seco-CBI or CBI.
  • Chlorambucil, seco-CBI, and CBI mentioned as examples of the alkylating agent are represented by the following chemical formula.
  • a composite is synthesized by combining the agent and the PI polyamide (hereinafter also referred to as “conjugate”).
  • the complex is also referred to as “conjugate”.
  • the synthesis method can be performed by a known method (for example, see J. Am. Chem. SOC. 1995, 117, 2479-2490).
  • the agent is attached to either the N-terminus, the C-terminus, or the ⁇ linker site of the PI polyamide, or a combination thereof.
  • the agent is bound to the N-terminus or C-terminus of PI polyamide.
  • the agent bonded to each position may be the same or different.
  • the “bonding” mode between the PI polyamide and the agent may be directly bonded or may be bonded via a linker.
  • the linker is not particularly limited as long as it does not interfere with the action of the agent and does not interfere with recognition of the RUNX binding site.
  • an amide bond, phosphodisulfide bond, ester bond, coordination bond, or ether bond examples thereof include molecules containing functional groups that form themselves or one or more of these bonds.
  • “Molecules containing functional groups that form one or more of these bonds” include amide bonds, phosphodisulfide bonds, ester bonds, coordination bonds, ether bonds, etc., together with the terminal portion of the PI polyamide and / or agent.
  • a molecule containing a functional group that forms one or more bonds selected from the group consisting of: “Molecules containing functional groups that form one or more of these bonds” also include one or more bonds selected from the group consisting of amide bonds, phosphodisulfide bonds, ester bonds, coordination bonds, ether bonds, and the like. May be a molecule containing one or more.
  • Preferred linkers include amide bonds or molecules containing functional groups that form amide bonds.
  • Examples of composites of agents and PI polyamides in this disclosure include those of formula I: [In Formula I, X 1 represents CH, COH or N, X 2 represents CH, COH or N, X 3 represents CH, COH or N, X 4 represents CH, COH or N, X 5 represents CH, COH or N Or X, X 6 represents CH, COH or N, X 7 represents CH, COH or N, X 8 represents CH, COH or N, wherein X 1 to X 8 represent PI polyamide Are selected in combinations that allow recognition of the RUNX consensus sequence, R 1 represents H or an alkyl group, R 2 represents H or an alkyl group, R 3 represents H or an alkyl group, R 4 represents H or an alkyl group, R 5 represents H or an alkyl group, R 6 represents H or an alkyl group, R 7 represents H or an alkyl group, R 8 represents H or an alkyl group, R 9 represents H, —NHR 11 , or —Ym—Z;
  • R 9 , R 10 , A, and B is -Ym-Z;
  • each of Ym in the plurality of Ym-Z may be the same as or different from the other Ym.
  • each of Z in the plurality of Ym-Z may be the same as or different from the other Z, and each of them may be independently selected.
  • A is [Wherein the integer n is any value from 0 to 5] Or represents -Ym-Z, wherein -Ym-Z represents the following formula: Any of the groups represented by B represents -Ym-Z, wherein -Ym-Z represents the following formula: [In the formula, the integer n is any value from 0 to 5, preferably any value from 0 to 2.] Any of the groups represented by
  • agents and PI polyamide composites in the present disclosure include Formula II: Or Formula III: [In Formula II and Formula III, X 1 represents CH, COH or N, X 2 represents CH, COH or N, X 3 represents CH, COH or N, X 4 represents CH, COH or N, X 5 represents CH, COH or N Or X, X 6 represents CH, COH or N, X 7 represents CH, COH or N, X 8 represents CH, COH or N, wherein X 1 to X 8 represent PI polyamide Are selected in combinations that allow recognition of the RUNX consensus sequence, R 1 represents H or an alkyl group, R 2 represents H or an alkyl group, R 3 represents H or an alkyl group, R 4 represents H or an alkyl group, R 5 represents H or an alkyl group, R 6 represents H or an alkyl group, R 7 represents H or an alkyl group, R 8 represents H or an alkyl group, R 9 represents H or —NHR 11 ; R 10 represents H
  • X is any one of X 1 to X 8 and represents CH or COH
  • R ′ is any one of R 1 to R 8 and represents H or an alkyl group.
  • Ym represents, for example, an amide bond, a phosphodisulfide bond, an ester bond, a coordination bond, an ether bond, or the like, or a moiety containing a functional group that forms one or more of these bonds.
  • a moiety containing a functional group that forms one or more of these bonds refers to an amide bond, a phosphodisulfide bond, an ester bond, a coordination bond, and a terminal part of PI polyamide and / or an agent. It is a part containing a functional group that forms one or more types of bonds selected from the group consisting of ether bonds and the like.
  • the “part containing a functional group forming one or more of these bonds” is one or more selected from the group consisting of an amide bond, a phosphodisulfide bond, an ester bond, a coordination bond, and an ether bond. One or more bonds may be included.
  • Ym is “a moiety containing a functional group that forms one or more of these bonds”, an example of which is represented by Formula IV: [Where: Q is carbonyl [—C ( ⁇ O) —] or imino (—NH—); Q ′ is an ether bond (—O—) or imino (—NH—) or methylimino [—N (—CH 3 ) —],
  • the integers g and k are each independently any value from 1 to 3
  • the integers h and j are each independently any value from 0 to 5,
  • the integer i is any value between 0 and 2]
  • the structure shown by is mentioned.
  • the integers h and j are preferably each independently any value from 0 to 3.
  • the position of the ester bond and the position of the ether bond or imino bond represented by Q ′ may be interchanged.
  • the linker moiety represented by the above formula IV for example, the position at the right end is connected to the agent and the position at the left end is connected to PI polyamide, but the connecting position may be reversed.
  • Q is preferably imino.
  • Y represented by formula IV the following formula VII: The structure shown by is mentioned.
  • the linker moiety represented by formula VII is used when the agent is attached to the C-terminal side of the PI polyamide.
  • the right end position is connected to the agent and the left end position is connected to PI polyamide, but the connecting positions may be reversed.
  • the pyrrole unit substituted with ⁇ -alanine is preferably in the second, third or fourth position from the N-terminal or C-terminal.
  • -Ym-Z is preferably [In the formula, the integer n is any value from 1 to 5, preferably any value from 0 to 2.] It is group represented by these.
  • -Ym-Z is preferably It is group represented by these.
  • examples of the alkyl group include linear, branched or cyclic saturated or unsaturated alkyl groups having 1 to 10 carbon atoms, preferably linear or branched groups having 1 to 5 carbon atoms.
  • examples include a straight chain, branched chain or cyclic saturated or unsaturated alkyl group, and examples thereof include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl and the like.
  • the alkyl group may be substituted.
  • methylene in the alkyl group may be substituted with oxygen or the like.
  • exemplary embodiments of the composite of the agent and the PI polyamide further include the following formula: [Where: Z represents an agent, preferably an alkylating agent, more preferably an alkylating agent selected from the group consisting of chlorambucil, duocarmycin, seco-CBI, CBI, pyrrolobenzodiazepine, and nitrogen mustard. Show n represents 0, 1, 2, 3, 4, or 5, preferably 1, 2, or 3, more preferably n represents 1, and At least one of the pyrrole units is substituted with ⁇ -alanine, preferably the second, third or fourth pyrrole unit from the N-terminal or C-terminal, more preferably the second or third from the N-terminal or C-terminal. Of pyrrole units are replaced by ⁇ -alanine] The compound shown by these, and its modification thing are mentioned.
  • n 0, 1, 2, 3, 4, or 5, preferably 1, 2, or 3, more preferably n represents 1, and At least one of the pyrrole units is substituted with ⁇ -alanine, preferably the second, third or fourth pyrrole unit from the N-terminal or C-terminal, more preferably the second or third from the N-terminal or C-terminal. Of pyrrole units are replaced by ⁇ -alanine.
  • specific examples of the composite of chlorambucil and PI polyamide include the following compounds.
  • the complex of PI polyamide and agent may be in the form of a pharmacologically acceptable salt.
  • a pharmacologically acceptable salt for example, inorganic acid salts such as hydrochloride, sulfate, phosphate or hydrobromide, or acetate, fumarate, maleate, oxalate, citrate, methanesulfonate, benzenesulfone
  • Organic acid salts such as acid salts or toluene sulfonates can be mentioned.
  • the complex in the complex, at least one part or molecule of the PI polyamide, the agent, and / or the linker moiety that links the PI polyamide and the agent is present in the form of an enantiomer or diastereomer, or a mixture thereof. obtain.
  • the complex includes a mixture of stereoisomers or each pure or substantially pure isomer.
  • the complex can be separated by a conventional method well known in the art, for example, chromatography or fractional crystallization.
  • the above-described complex may contain an isotope (for example, 3 H, 13 C, and / or a molecule on at least one part or molecule of a linker part that links PI polyamide, an agent, and / or PI polyamide and the agent. 14 C, 15 N, 18 F, 32 P, 35 S, 125 I and the like), or a deuterium converter.
  • an isotope for example, 3 H, 13 C, and / or a molecule on at least one part or molecule of a linker part that links PI polyamide, an agent, and / or PI polyamide and the agent.
  • 14 C, 15 N, 18 F, 32 P, 35 S, 125 I and the like or a deuterium converter.
  • a RUNX inhibitor is a composition comprising the PI polyamide.
  • the RUNX inhibitor of the present invention may preferably contain a complex of the PI polyamide and the agent.
  • a further preferred example of a RUNX inhibitor in the present disclosure includes a complex of the above PI polyamide and an alkylating agent.
  • RUNX consensus binding sequence is a binding sequence common to each member of the RUNX family.
  • a RUNX inhibitor includes the PI polyamide designed to recognize a RUNX consensus binding sequence on the genome, thus inhibiting any activity caused by the binding of each member of the RUNX family to DNA.
  • a RUNX inhibitor in the present disclosure binds to a RUNX consensus binding sequence on the genome and inhibits binding of RUNX family members to the binding sequence.
  • RUNX consensus sequences are present in the regulatory regions of various genes, and RUNX family members control the expression of various target genes by binding to the consensus sequences in the regulatory regions. . Therefore, there are various activities involved in the RUNX family, including, but not limited to, for example, transcriptional control of p53 inhibitors (BCL11, TRIM24, etc.) (ie, in cancer, the RUNX family MLL-AF4 + FLT3-ITD acute myeloid leukemia that transcriptionally regulates and enhances BCR-ABL, the causative protein of Philadelphia chromosome positive acute lymphoblastic leukemia (PhALL) And MLL-AF4 is enhanced in transcription, and oncogene c-Myc is regulated by transcription and enhanced.
  • p53 inhibitors BCL11, TRIM24, etc.
  • the RUNX inhibitor may be the PI polyamide or the complex of the PI polyamide and the agent itself depending on the purpose of use, or in addition to the PI polyamide or the complex,
  • An additive may be included.
  • the carrier and additive include, but are not limited to, water, acetic acid, organic solvent, collagen, polyvinyl alcohol, polyvinyl pyrrolidone, carboxyvinyl polymer, sodium carboxymethylcellulose, sodium polyacrylate, sodium alginate, water-soluble Dextran, sodium carboxymethyl starch, pectin, methylcellulose, ethylcellulose, xanthan gum, gum arabic, casein, agar, polyethylene glycol, diglycerin, glycerin, propylene glycol, petrolatum, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol , Lactose, surfactants and the like.
  • the present disclosure further provides a method for inhibiting the activity of the RUNX family by using the RUNX inhibitor of the present invention.
  • the RUNX family inhibition method can inhibit not only RUNX1, RUNX2, or RUNX3, but also a gene cluster (target gene group) controlled by all members of the RUNX family at a time.
  • the amount of the RUNX inhibitor used can be appropriately adjusted according to the purpose of use.
  • kits containing the RUNX inhibitor of the present invention are also provided.
  • the kit may contain pharmaceutically acceptable carriers and additives, reagents, adjuvants, dedicated containers, other necessary accessories, instructions, and the like.
  • the kit of the present invention can be used, for example, as a cancer treatment kit or a research reagent kit.
  • the pharmaceutical composition is a composition containing the PI polyamide.
  • the pharmaceutical composition in the present disclosure may preferably contain a complex of the PI polyamide and the agent.
  • a further preferred example of a pharmaceutical composition in the present disclosure includes a complex of the above PI polyamide and an alkylating agent.
  • the pharmaceutical composition in the present disclosure may include a RUNX inhibitor disclosed in the present specification.
  • RUNX consensus sequences are present in the regulatory regions of various genes. RUNX family members control the expression of various target genes by binding to consensus sequences in the regulatory region. Examples of such target genes include, but are not limited to, genes that are highly expressed in CBF leukemia (eg, IL3, CSF2, CSF2RB, etc.), the RUNX family itself (RUNX1, RUNX2, RUNX3), p53 inhibitor ( For example, BCL11, TRIM24, etc.), c-kit gene and the like can be mentioned.
  • CBF leukemia eg, IL3, CSF2, CSF2RB, etc.
  • RUNX1, RUNX2, RUNX3 p53 inhibitor
  • BCL11, TRIM24, etc. c-kit gene and the like can be mentioned.
  • the pharmaceutical composition of the present invention contains the above-mentioned PI polyamide designed to recognize a RUNX consensus binding sequence on the genome, it down-regulates the expression of various genes targeted by each RUNX family member.
  • the pharmaceutical composition of the invention binds to a RUNX consensus binding sequence on the genome and inhibits binding of RUNX family members to the binding sequence.
  • ⁇ ⁇ Various diseases can be treated and prevented by administering the pharmaceutical composition of the present disclosure in vivo.
  • the pharmaceutical composition in the present disclosure should be used for any organism that utilizes double-stranded DNA for biological control, particularly mammals (eg, humans, rats, rabbits, sheep, pigs, cattle, cats, dogs, monkeys, etc.). Can do.
  • target diseases of the pharmaceutical composition according to the present disclosure are all diseases involving RUNX family members.
  • target diseases of the pharmaceutical composition in the present disclosure include tumors, particularly cancer, such as, but not limited to, leukemia (eg, acute myeloid leukemia, acute lymphocytic leukemia, chronic myelogenous leukemia).
  • leukemia eg, acute myeloid leukemia, acute lymphocytic leukemia, chronic myelogenous leukemia.
  • an antitumor composition comprising the PI polyamide or a complex of the PI polyamide and an agent is provided as a pharmaceutical composition in the present disclosure.
  • the antitumor composition in the present disclosure includes a composite of the PI polyamide and an alkylating agent.
  • mast cell tumors and mastocytosis eg, mast cell hypertrophy, severe allergic diseases, atopic dermatitis, anaphylactic shock, severe bronchial asthma attacks Mast cell diseases such as severe drug dermatitis), various allergies, and immune diseases.
  • a mast cell is defined as a cell that expresses on the cell surface both the specific receptor FceRI for an IgE antibody closely related to allergy and c-kit, which is a receptor for the cytokine called stem cell factor (SCF).
  • SCF stem cell factor
  • an antiallergic composition comprising the PI polyamide or a complex of the PI polyamide and an agent is provided as a pharmaceutical composition in the present disclosure.
  • the antiallergic composition of the present invention contains a composite of the above PI polyamide and an alkylating agent.
  • an antiallergic composition in the present disclosure includes a RUNX inhibitor disclosed herein.
  • the inventors have shown that a complex of PI polyamide and alkylating agent targeting the RUNX binding sequence suppresses the expression of c-kit (stem cell factor receptor tyrosine kinase) in mast cells.
  • c-kit stem cell factor receptor tyrosine kinase
  • the pharmaceutical compositions in this disclosure that target the same RUNX binding sequence also inhibit c-kit expression as well. Therefore, the pharmaceutical composition in the present disclosure can suppress, treat, or prevent all symptoms or diseases caused by mast cell activation.
  • the pharmaceutical composition in the present disclosure may be in a dosage form for oral administration or parenteral administration.
  • These dosage forms can be formulated according to a conventional method, and may contain pharmaceutically acceptable carriers and additives.
  • Such carriers and additives include water, acetic acid, pharmaceutically acceptable organic solvents, collagen, polyvinyl alcohol, polyvinyl pyrrolidone, carboxyvinyl polymer, sodium carboxymethylcellulose, sodium polyacrylate, sodium alginate, water soluble dextran, Sodium carboxymethyl starch, pectin, methylcellulose, ethylcellulose, xanthan gum, gum arabic, casein, agar, polyethylene glycol, diglycerin, glycerin, propylene glycol, petrolatum, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol, lactose And surfactants acceptable as pharmaceutical additives.
  • the above additives are selected from the above alone or in combination as appropriate according to the dosage form of the pharmaceutical composition in the present disclosure.
  • tablets, capsules, fine granules, powders, granules, solutions, syrups, sprays, coatings, eye drops, external preparations, etc. or appropriate dosage forms for oral administration Can be administered.
  • parenteral administration examples include an injection form.
  • an injection form it can be administered systemically or locally by intravenous injection such as infusion, subcutaneous injection, intraperitoneal injection, intratumoral injection, and the like.
  • the pharmaceutical composition of the present disclosure when used as an injectable preparation, is dissolved in a solvent (eg, physiological saline, buffer solution, glucose solution, 0.1% acetic acid, etc.), and appropriate additives (humans) Serum albumin, PEG, mannose modified dendrimer, cyclodextrin conjugate, etc.) can be used.
  • a solvent eg, physiological saline, buffer solution, glucose solution, 0.1% acetic acid, etc.
  • appropriate additives humans
  • it may be freeze-dried to obtain a dosage form that dissolves before use.
  • the freeze-drying excipient for example, sugar alcohols and saccharides such as mannitol and glucose can be used.
  • the dosage of the pharmaceutical composition in the present disclosure varies depending on age, sex, symptom, administration route, administration frequency, and dosage form.
  • the dose is 0.01 to 1000 mg, preferably 0.1 to 100 mg, more preferably 1 to 30 mg per day.
  • the administration method is appropriately selected depending on the age and symptoms of the patient. Administration may be divided, for example, once every several days, once per day, or 2-4 times.
  • the pharmaceutical composition, particularly the antitumor composition in the present disclosure can be used in combination with other antitumor agents.
  • Other anti-tumor agents include any anti-tumor agent used to treat a particular cancer, or p53 inducer.
  • any known antitumor agent may be used, for example, cytarabine, imatinib, gefitinib, PRIMA-1, MIRA-1, Nutlin3, etc. Can be mentioned.
  • the administration ratio between the pharmaceutical composition and the other antitumor agent in the present disclosure is not particularly limited, and may be appropriately determined by those skilled in the art so as to obtain a desired antitumor effect.
  • Example 1 Synthesis of PI Polyamide and Conjugate A PI polyamide that specifically recognizes the RUNX consensus binding sequence of 5'-TGTGGT-3 'by sequential binding of four pyrrole-imidazole pairs, A PI polyamide in which the pyrrole unit was replaced by ⁇ -alanine was designed, and the PI polyamide was synthesized with a chlorambucil conjugate (Chb-M). As a control, a conjugate (Chb-M ′) of PI polyamide and chlorambucil in which the pyrrole unit was not replaced by ⁇ -alanine was synthesized.
  • Chb-M chlorambucil conjugate
  • Chb-M ′ The hydroxy substituted Chb-M ′ (1.8 mg, 1.2 ⁇ mol) of Chb-M ′ was dissolved in a mixture of MeCN / 0.1% TFA (2: 5) and stirred at 40 ° C. for 2 days. The mixture was purified by HPLC and the final weight of the target product was 1.1 mg (0.75 ⁇ l, 65% yield). Retention time was 9.92 minutes (detected at 254 nm with 0.1% TFA containing 0-100% acetonitrile with a linear gradient of 20 minutes at a flow rate of 1.0 mL / min).
  • MALDI-TOFMS spectrum m / z C 71 H 91 N 24 O 13 + [M + H] + calculated value 1487.719, actual value 1487.715
  • Example 2 Binding Affinity of Chb-M to Target DNA
  • the binding affinities of Chb-M and Chb-M ′ were compared using hydroxy substitution of Chb-M and Chb-M ′.
  • Biotinylated hairpin DNA containing the target sequence (5′-TGTGGT-3 ′) was immobilized on the sensor chip by forming a complex of biotin and streptavidin.
  • the binding properties were evaluated by SPR (surface plasmon resonance).
  • the concentration of the PI polyamide conjugate was measured as follows.
  • Biotinylated hairpin DNA (5'-biotin-CGCTTGTGTCTCGTTTCGGACCACAAAGCG-3 ') was purchased from SIGMA-Aldrich. Streptavidin-functionalized SA sensor chip was purchased from Biacore. Biotinylated DNA was immobilized on the sensor chip to obtain the desired immobilization level (increase by about 900 RU).
  • the SPR assay was performed using HBS-EP buffer (10 mM HPES (pH 7.4), 150 mM NaCl, 3 mM EDTA, and 0.005% Surfactant P20) at 25 ° C. with 0.1% DMSO.
  • PI polyamide conjugate solutions with various concentrations were prepared in buffers containing 0.1% DMSO and injected at a flow rate of 20 ⁇ L / min.
  • ka binding rate
  • kd dissociation rate
  • K D dissociation constant
  • Chb-M had a binding affinity for the target 100 times higher than that of Chb-M ′.
  • Example 3 Chb-M DNA alkylation activity
  • the DNA alkylation activity of Chb-M and Chb-M ' was tested by polyacrylamide gel electrophoresis (PAGE) analysis.
  • the DNA fragment was alkylated using Chb-M and Chb-M ′. Alkylation was carried out at room temperature for 18 hours. In order to visualize the alkylation site, the sample was heated at 95 ° C. for 10 minutes and then treated with 1M piperidine at 95 ° C. for 25 minutes. The alkylated DNA was cleaved by heat treatment. Piperidine treatment converts the modified sugar terminus to a 3′-phosphate terminus. A DNA fragment having a 3′-phosphate end was observed on the gel as an alkylated band.
  • Table 2 shows all oligonucleotides (purchased from Sigma-Aldrich) used in this example.
  • Complementary FAM-labeled fragments (30 bp) were prepared by the same procedure using other DNA fragments (ODN5: 5′-FAM-ATATGTGTATATGTGGTTAATGTGGTAAT-3 ′ and ODN6: 5′-ATTACCACATTAACCCACAATTACCACATAT-3 ′).
  • ODN 3 and 4 and ODN 7 and 8 were used as reference DNA. All ODNs except ODN2 and ODN6 were 5′-end FAM labeled.
  • FIG. 2A shows the results of the alkylation activity for the upper chain. Since two bands were observed at the same position as ODN3 and ODN4, Chb-M ′ alkylated two adenines in the target sequence. On the other hand, Chb-M alkylated only one site. A weak alkylated band at the mismatch site was also observed, but Chb-M had higher sequence specificity in the target sequence than Chb-M ′. Further, the density of the band showed that the alkylation activity of Chb-M was superior to that of Chb-M ′. Thus, the introduction of ⁇ -alanine increased both the sequence specificity and alkylation activity of PI polyamide. The alkylation activity results for the lower strand showed no difference in sequence specificity between Chb-M and Chb-M ′ (FIG. 2B).
  • Example 4 Cytotoxicity of Chb-M on lung cancer cell lines
  • 50% inhibitory concentrations (IC 50 ) for various lung cancer cell lines were determined.
  • EGFR mutant lung cancer cell lines PC3, PC9, LU99A
  • EGFR wild type cell lines A549, ABC1, RERFLCMS
  • the molecular target drug gefitinib EGFR inhibitor
  • a non-molecular target drug cisplatin for conventional lung cancer
  • Chb-M ′ were used.
  • Example 5 Cytotoxicity against brain tumor cell line by Chb-M To evaluate the cytotoxic effect of Chb-M on brain tumor cells, 50% inhibitory concentration (IC 50 ) for various brain tumor cell lines was determined.
  • Chb-M ′ is a conjugate of PI polyamide and chlorambucil targeting the 5′-WGGGCCW-3 ′ sequence (W is either A or T) and is the same as in Example 1. The method was synthesized. The chemical formula of Chb-S is shown below.
  • the medulloblastoma cell line Daoy was brought to a density of 1 ⁇ 10 5 cells / mL.
  • Various concentrations of PI polyamide (Chb-M, Chb-M ′, Chb-S) and Chb (chlorambucil) were added to the medium and incubated for 72 hours.
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions.
  • the dose that inhibited growth by 50% was analyzed by a semi-effective method.
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions. The dose that inhibited growth by 50% was analyzed by the median-effect method.
  • FIGS. 4A and B The results are shown in FIGS. 4A and B.
  • the IC 50 of Chb-M for A172 cells 0.531 ⁇ M for 72 hours and 0.421 ⁇ M for 96 hours
  • the control Chb-M ′ 23.066 ⁇ M for 72 hours and 6.677 ⁇ M for 96 hours.
  • the concentration was much lower than that.
  • the IC 50 of Chb-M for KALS-1 cells (0.214 ⁇ M for 72 hours and 0.738 ⁇ M for 96 hours) is compared to the control Chb-M ′ (44.82 ⁇ M for 96 hours). Much lower concentrations.
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions. The dose that inhibited growth by 50% was analyzed by the median-effect method.
  • Example 6 Cytotoxicity against prostate cancer cell line by Chb-M CRPC-Adeno: To assess the antitumor effect of Chb-M on surgical resection refractory prostate cancer (adenocarcinoma), an IC 50 of 72 hours was determined. It was. Surgical resection refractory prostate cancer (adenocarcinoma) cell lines: DU145 cells (DU 145 (ATCC R HTB- 81 TM): ATCC purchased from) was to 1 ⁇ 10 5 cells / mL density. Various concentrations of Chb-M, Chb-M ′, Chb-S, or Chb were added to the medium and incubated for 72 hours.
  • DU145 cells DU 145 (ATCC R HTB- 81 TM): ATCC purchased from
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions. The dose that inhibited growth by 50% was analyzed by the median-effect method.
  • Example 7 Cytotoxicity against leukemia cell line by Chb-M (1) Anti-tumor effect against acute myeloid leukemia
  • MOLM13 Japanese Collection of Research Bioresources, Japan
  • AML cell line of TP53 wild-type cell line MV4-11 and MV4-11NR cells carrying the 53TP53 R248W mutation is described in T.W. Acquired from Dr. Ikezoe (Department of Hematology and Respiratory Medicine, Kochi University, Kochi, Japan). These cells were brought to a density of 1 ⁇ 10 5 cells / mL.
  • Chb-M or Chb-M ′ were added to the medium and incubated for 48 hours.
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions.
  • the dose that inhibited growth by 50% was analyzed by the median-effect method.
  • IC 50 values of 48 hours and 72 hours were determined.
  • the NB4 cell line cell line was obtained from Keio University School of Medicine. NB4 cells were brought to a density of 1 ⁇ 10 5 cells / mL.
  • Various concentrations of Chb-M or Chb-M ′ were added to the medium and incubated for 48 and 72 hours.
  • Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions. The dose that inhibited growth by 50% was analyzed by the median-effect method.
  • Example 8 Cyb-M to osteosarcoma line
  • the osteosarcoma cell MG63 cell line cell line was obtained from the Department of Pediatrics, Kyoto University. MG63 cells were brought to a density of 1 ⁇ 10 5 cells / mL. Various concentrations of Chb-M or Chb-M ′ were added to the medium and incubated for 72 hours. Cell viability was assessed by counting viable cells using Cell Count Reagent SF (nacalai tesque, Inc.) and InfiniteR 200 PRO multimode reader (TECAN) according to the manufacturer's instructions. The dose that inhibited growth by 50% was analyzed by the median-effect method.
  • the antitumor composition of the present invention can target various types of cancer including leukemia by the cluster control of the RUNX family.
  • the antitumor composition of the present invention is also effective against tumors that are resistant to other molecularly targeted therapeutic agents.
  • the anti-tumor composition of the present invention is effective against cancers that are not effective with molecularly targeted therapeutic agents currently used in clinical practice, and is therefore an all-purpose anticancer agent that is also effective for cancers that are said to be difficult to treat. The use as is expected.
  • the RUNX inhibitor of the present invention can be used as an antiallergic composition.
  • SEQ ID NO: 1 Sequence of ODN1 SEQ ID NO: 2: Sequence of ODN2 SEQ ID NO: 3: Sequence of ODN3 SEQ ID NO: 4: Sequence of ODN4 SEQ ID NO: 5: Sequence of ODN5 SEQ ID NO: 6: Sequence of ODN6 SEQ ID NO: 7: Sequence of ODN7 SEQ ID NO: 8: Sequence of ODN8

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Abstract

La présente invention concerne : une composition anti-tumorale et une composition anti-allergène comprenant un polyamide pyrrole-imidazole pouvant se lier à une séquence de liaison au RUNX, un ou plusieurs motifs pyrrole qui constituent le polyamide pyrrole-imidazole qui est substitué par une β-alanine ; et un inhibiteur du RUNX.
PCT/JP2019/020961 2018-05-28 2019-05-27 Composition pharmaceutique ciblant une séquence de liaison au runx, et inhibiteur du runx WO2019230669A1 (fr)

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