HDAC inhibitor as well as preparation method and application thereof

Abstract

The invention discloses an HDAC (histone deacetylase) inhibitor as well as a preparation method and application thereof. Specifically, the invention provides a compound shown as a formula (I), or a stereoisomer, a tautomer, a solvate, a prodrug, an isotope label, a pharmaceutically acceptable salt of the compound, and a pharmaceutical composition containing the compound. The invention also provides a compound which inhibits tumor cell proliferation by inhibiting HDAC and affects the cycle and movement ability of tumor cells, so that the compound has good anti-tumor activity. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry, and specifically relates to an HDAC inhibitor and a preparation method and use thereof. Background Art

[0002] Histone acetylation is an epigenetic regulatory mechanism that is regulated by histone acetyltransferases (HATs) and histone deacetylases (HDACs). When HDACs are overexpressed, DNA and chromatin are tightly bound, which is not conducive to the transcription of specific tumor suppressor genes and promotes the occurrence and development of cancer (Biochem. J. 2003, 370 (3): 737-749).

[0003] To date, a total of five HDAC pan-inhibitors have been approved for marketing, including the hydroxamic acid compound vorinostat (SAHA) for the treatment of cutaneous T-cell lymphoma and the hydroxamic acid compound panobinostat for the treatment of relapsed or refractory multiple myeloma. However, since the HDAC inhibitors that have been marketed can act on multiple subtypes of HDAC, nausea, vomiting, bone marrow suppression and other toxic side effects are common, and HDAC subtype selective inhibitors may avoid these adverse reactions. Therefore, the development and research of HDAC subtype selective inhibitors is of great significance. Structurally, HDAC inhibitors are usually composed of an enzyme surface recognition group (Cap), a linker fragment that fills the catalytic channel, and a zinc ion chelating group (ZBG), such as ACY-1215 and TUB-A.

[0004]

[0005] A large number of studies have shown that phenanthrene compounds have a variety of biological activities, including anti-tumor, anti-inflammatory, anti-malarial, anti-viral, anti-neurological diseases, etc. (J.Med.Chem.2012,55(23):10363-10377). Tylophorine is a phenanthroline alkaloid from Asclepiadaceae. Since it was first isolated from Rhizoma Cibotii, its pharmacological activity and synthesis method have been widely studied (Bioorg.Med.Chem.Lett.2017,27(11):2369-2376). This type of alkaloid not only has very strong anti-tumor activity, but also has good therapeutic effects in anti-inflammatory, anti-microbial and anti-leukemia. Antofine, as a typical phenanthroline alkaloid, has strong anti-cancer activity and has a strong inhibitory effect on the GI tract of A549 lung cancer cells. 50 The value is 0.01 μM. However, the targets of phenanthrene compounds are unclear, and the unknown off-target risks limit their further development.

[0006] The present invention combines the structural characteristics of HDAC inhibitors and the anti-tumor activity of phenanthrene compounds, uses phenanthrene ring derivatives as Cap units, covalently connects them to hydroxamic acid groups via Linkers, and constructs target molecules to provide a new type of selective HDAC inhibitors. Summary of the invention

[0007] In order to overcome the deficiencies of the prior art, the present invention provides an HDAC inhibitor and a preparation method and use thereof. In one aspect, the present invention provides a compound represented by formula (I):

[0008]

[0009] or stereoisomers, tautomers, solvates, prodrugs, isotope labels, and pharmaceutically acceptable salts thereof; wherein R1, R2, R3, A, B and D are as defined herein.

[0010] In another aspect, the present invention provides a pharmaceutical composition comprising a compound of formula (I) or a stereoisomer, tautomer, solvate, prodrug, isotope-labeled substance, pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

[0011] In another aspect, the present invention provides the use of the compound of formula (I) or its stereoisomers, tautomers, solvates, prodrugs, isotope labels, and pharmaceutically acceptable salts and / or pharmaceutical compositions containing the same in the preparation of drugs for preventing and / or treating cancer-related diseases.

[0012] Beneficial effects: Compared with the prior art, the compounds of the present invention not only have better selective HDAC (HDAC1 / HDAC6) inhibitory activity, but also can achieve good anti-tumor effects by inhibiting the invasion and metastasis of tumor cells and blocking the cell cycle. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] The accompanying drawings are used to provide a further understanding of the present invention and constitute a part of the specification. Together with the embodiments of the present invention, they are used to explain the present invention and do not constitute a limitation of the present invention. In the accompanying drawings:

[0014] Figure 1 . Shows the effect of different concentrations of compound HD-1 on the cell cycle of HEL cells.

[0015] Figure 2 . Shows the effects of different concentrations of compound HD-1 on HEL cell migration and invasion. DETAILED DESCRIPTION

[0016] definition

[0017] As used in this specification, the following words and phrases are generally intended to have the meanings set forth below, unless the context in which they are used indicates otherwise.

[0018] As used herein, the term "alkyl" refers to a monovalent group of a straight or branched saturated hydrocarbon chain having 1 to 20 carbon atoms, more typically 1 to 10 carbon atoms, 1 to 8 carbon atoms, or 1 to 6 carbon atoms. This term is exemplified by groups such as methyl, ethyl, 1-propyl (n-propyl), 2-propyl (isopropyl), 1-butyl (n-butyl), 2-methyl-1-propyl (isobutyl), 2-butyl (sec-butyl), 2-methyl-2-propyl (tert-butyl), 1-pentyl (n-pentyl), 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3-methyl-1-butyl, 2-methyl-1-butyl, 1-hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2,3-dimethyl-2-butyl, 3,3-dimethyl-2-butyl, 1-heptyl, 1-octyl, 1-nonyl, 1-decyl, and the like.

[0019] As used herein, the term "alkylene" refers to a divalent group of a straight or branched saturated hydrocarbon chain having 1 to 20 carbon atoms (more typically 1 to 10 carbon atoms, 1 to 8 carbon atoms, or 1 to 6 carbon atoms). The term is exemplified by groups such as methylene, ethylene, propylene, butylene, pentylene, hexylene, and the like.

[0020] As used herein, the term "alkenyl" refers to a linear or branched unsaturated hydrocarbon chain monovalent group having 2 to 20 carbon atoms (more typically 2 to 10 carbon atoms, 2 to 8 carbon atoms, or 2 to 6 carbon atoms) and having carbon-carbon double bonds (e.g., 1, 2 or 3 carbon-carbon double bonds). The term is exemplified by groups such as vinyl (i.e., -CH=CH2), propen-1-yl (i.e., -CH=CHCH3), propen-3-yl (or allyl, i.e., -CH2CH=CH2), propen-2-yl (i.e., -C(CH3)=CH2), butadienyl (including 1,2-butadienyl and 1,3-butadienyl), and the like.

[0021] As used herein, the term "alkenylene" refers to a divalent group of a straight or branched unsaturated hydrocarbon chain having 2 to 20 carbon atoms (more typically 1 to 10 carbon atoms, 1 to 8 carbon atoms, or 1 to 6 carbon atoms) and having carbon-carbon double bonds (e.g., 1, 2 or 3 carbon-carbon double bonds). The term is exemplified by groups such as vinylene, propenylene, butenylene, pentenylene, hexenylene, etc.

[0022] As used herein, the term "alkynyl" refers to a linear or branched unsaturated hydrocarbon chain monovalent group having 2 to 20 carbon atoms (more typically 2 to 10 carbon atoms, 2 to 8 carbon atoms, or 2 to 6 carbon atoms) and having carbon-carbon triple bonds (e.g., 1, 2 or 3 carbon-carbon triple bonds). The term is exemplified by groups such as ethynyl (i.e., -C≡CH), propargyl (i.e., -CH2C≡CH), propynyl (i.e., -C≡CCH3), etc.

[0023] As used herein, the term "aryl" refers to an aromatic carbocyclic group of 6 to 14 carbon atoms (more typically 6 to 10 carbon atoms, or 6 carbon atoms) having a single ring (e.g., phenyl) or multiple rings (e.g., biphenyl) or multiple condensed (fused) rings (e.g., naphthyl, fluorenyl, and anthracenyl). The term is exemplified by groups such as phenyl, fluorenyl, naphthyl, anthracenyl, 1,2,3,4-tetrahydronaphthalene (if the point of attachment is through the aryl group), and the like.

[0024] As used herein, the term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0025] As used herein, the term "alkoxy" refers to an "alkyl-O-" group, wherein alkyl is as defined herein. This term is exemplified by groups such as methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, and the like.

[0026] As used herein, the term "haloalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by a halogen, wherein alkyl is as defined herein. The term is exemplified by the groups trifluoromethyl, difluoromethyl, monofluoromethyl, 2,2,2-trifluoroethyl, 1,1-difluoroethyl, and the like.

[0027] As used herein, the term "carbocycle", "carbocyclic radical" refers to a monocyclic saturated or partially unsaturated group of 3 to 8 yuan or a plurality of thick (fused) rings or bridged rings or spirocycles having 3 to 14 carbon atoms as ring atoms.Carbocycle or carbocyclic radical can be saturated or partially unsaturated, and can be fused with another saturated, partially unsaturated or aromatic carbocyclic ring, provided that the ring atom connected to the target molecule is not aromatic carbon.Examples of carbocycle or carbocyclic radical include, but are not limited to cyclopropane, cyclobutane, cyclopentane, cyclohexane, cyclobutene, cyclopentene, cyclohexene, cycloheptene, cyclopentadiene etc.

[0028] As used herein, the term "heteroaromatic ring", "heteroaryl" refers to an aromatic ring group containing a monocyclic or multiple condensed (fused) ring (e.g., containing 2 or 3 rings) of 5 to 14 ring atoms in the ring, wherein in addition to carbon atoms, the ring atoms also contain at least one heteroatom selected from oxygen, nitrogen and / or sulfur. If the ring is aromatic, the sulfur and nitrogen atoms can also exist in oxidized form. Multiple condensed (fused) ring heteroaryl is a monocyclic heteroaryl as defined above fused with one or more rings selected from the following to form a multiple condensed ring system: heteroaryl (to form, for example, naphthyridinyl, such as 1,8-naphthyridinyl), heterocyclic (for example, 1,2,3,4-tetrahydronaphthyridinyl, such as 1,2,3,4-tetrahydro-1,8-naphthyridinyl), carbocyclic (to form, for example, 5,6,7,8-tetrahydroquinolinyl) and aryl (to form, for example, indazolyl). Such multiple fused ring systems can be optionally substituted with one or more (e.g., 1, 2, 3, or 4) oxo groups on the carbocyclic or heterocyclic portions of the fused rings. When valence requirements permit, the rings of the multiple fused ring systems can be interconnected by fusion, spirocyclic, and bridging bonds. It should be understood that the individual rings of the multiple fused ring systems can be connected to each other in any order. It should also be understood that the attachment point of the multiple fused ring system can be at any position of the multiple fused ring system, including heteroaryl, heterocyclic, aryl, or carbocyclic portions of the multiple fused system. It should also be understood that the attachment point of the heteroaryl group can be on any suitable atom of the heteroaryl group, including carbon atoms and heteroatoms (e.g., nitrogen). Exemplary heteroaryl groups include, but are not limited to, pyridinyl, pyrrolyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrazolyl, thienyl, indolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, furanyl, oxadiazolyl, thiadiazolyl, quinolinyl, isoquinolinyl, benzothiazolyl, benzoxazolyl, indazolyl, quinoxalinyl, quinazolinyl, 5,6,7,8-tetrahydroisoquinolinyl, benzofuranyl, benzimidazolyl, thiaindenyl, pyrrolo[2,3-b]pyridinyl, quinazolinyl-4(3H)-one, triazolyl, 4,5,6,7-tetrahydro-1H-indazolyl, and 3b,4,4a,5-tetrahydro-1H-cyclopropane[3,4]cyclopenta[1,2-c]pyrazolyl.

[0029] As used herein, the terms "heterocycle", "heterocyclyl" and "heterocyclic group" refer to a monoradical saturated or partially unsaturated group having a 5- to 8-membered monocyclic ring or multiple condensed (fused) rings or bridged rings having 5 to 14 ring atoms in the ring, wherein in addition to carbon atoms, the ring atoms also contain at least one heteroatom selected from oxygen, nitrogen and / or sulfur. Examples of heterocyclyl groups include, but are not limited to, a tetrahydropyrrole ring, a piperidine ring, an azepane ring, an azirocine ring, an oxirane ring, an oxetane ring, a tetrahydrofuran ring, a tetrahydropyrane ring, an oxepane ring, an oxocane ring, a thiirane ring, a thietane ring, a tetrahydrothiophene ring, a tetrahydrothiopyran ring, a thipane ring, a thiocane ring, a tetrahydroimidazole ring, a tetrahydropyrazole ring, a tetrahydrooxazole ring, a tetrahydroisoxazole ring, a tetrahydrothiazole ring, a tetrahydroisothiazole ring, a piperazine ring, a morpholine ring, a dioxane ring, a thioxane ring, a dithiane ring, a dihydropyridyl group, 4,5,6,7-tetrahydro-1H-benzo[d]imidazole, 4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine, and the like.

[0030] As used herein, the term "therapeutically effective amount" refers to an amount sufficient to affect treatment as defined below when administered to a mammal in need of such treatment. The therapeutically effective amount will vary with the subject and disease condition being treated, the weight and age of the subject, the severity of the disease condition, the mode of administration, etc., and can be readily determined by one of ordinary skill in the art.

[0031] As used herein, the term "stereoisomer" refers to a compound having the same chemical composition and connectivity, but whose atoms have different orientations in space that cannot be interchanged by single bond rotation. "Stereoisomer" includes "diastereomers" and "enantiomers". "Diastereomers" refers to stereoisomers with two or more chiral centers and whose molecules are not mirror images of each other. Diastereomers have different physical properties, such as melting points, boiling points, spectral characteristics and reactivity. Diastereomeric mixtures can be separated under high-resolution analytical procedures (such as crystallization, electrophoresis and chromatography). "Enantiomers" refers to two stereoisomers that are non-overlapping mirror images of each other.

[0032] As used herein, the term "tautomer" refers to the coexistence of two (or more) compounds that differ only in the position and electron distribution of one (or more) mobile atoms, such as keto-enol tautomers.

[0033] As used herein, the term "pharmaceutically acceptable salt" refers to salts that retain the biological effectiveness and properties of a given compound and are not biologically or otherwise undesirable. Pharmaceutically acceptable salts can be acid addition salts and / or base addition salts. Acid addition salts can be prepared from inorganic acids and organic acids. Salts derived from inorganic acids include hydrochlorides, hydrobromides, sulfates, nitrates, phosphates, carbonates, bisulfates, hydrogenphosphates, dihydrogenphosphates, bicarbonates, etc.; salts derived from organic acids include formates, acetates, propionates, glycolates, pyruvates, oxalates, malates, malonates, succinates, maleates, fumarates, tartrates, citrates, benzoates, cinnamates, mandelates, methanesulfonates, ethanesulfonates, p-toluenesulfonates, salicylates, lactates, nicotinates, lauryl sulfates, naphthylsulfonates, camphorsulfonates, gluconates, glucuronates, oleates, palmitates, stearates, pamoates, trifluoroacetates, etc. Base addition salts can be formed with inorganic or organic bases. Salts derived from inorganic bases include sodium, potassium, ammonium, calcium, magnesium, iron, zinc, copper, lithium, barium, aluminum salts and the like; salts derived from organic bases include salts formed with various primary, secondary and tertiary amines, for example, ethylamine, diethylamine, n-propylamine, isopropylamine, diethanolamine, meglumine, lysine, piperazine, piperidine, morpholine, tromethamine, choline and the like.

[0034] As used herein, the term "pharmaceutically acceptable" means that the substance or composition must be chemically and / or toxicologically compatible with the other ingredients comprising the formulation and / or the mammal to be treated therewith.

[0035] As used herein, the term "solvate" refers to an association or complex of one or more solvent molecules with a compound of the invention. Examples of solvents that form solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine. The term "hydrate" refers to a complex in which the solvent molecule is water.

[0036] As used herein, the term "prodrug" refers to those compounds that readily undergo chemical changes under physiological conditions to provide compounds of the present invention. In addition, prodrugs can be converted into compounds of the present invention by chemical or biochemical methods in an ex vivo environment. For example, when a prodrug is placed in a transdermal patch reservoir together with a suitable enzyme or chemical reagent, the prodrug can be slowly converted into a compound of the present invention.

[0037] Any formula or structure given herein, including Formula I or any formula disclosed herein, is also intended to represent unlabeled forms of the compound as well as isotopically labeled forms. These forms of the compound may also be referred to as "isotopically labeled" or "isotopically enriched analogs." An isotopically labeled compound has a structure as depicted herein except that one or more atoms are replaced by an atom having a selected atomic mass or mass number. Examples of isotopes that can be incorporated into the compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine, chlorine, and iodine, such as, but not limited to, 2 H (deuterium, D), 3 H (tritium), 11 C. 13 C. 14 C. 13 N. 15 N. 15 O. 17 O. 18 O. 31 P. 32 P. 35 S. 18 F. 36 Cl, 123 I and 125 I. Various isotope-labeled compounds of the present invention, for example, compounds into which a radioactive isotope (e.g. 3 H. 13 C and 14 C). Such compounds are synthesized by means well known in the art, for example by employing starting materials in which one or more hydrogens have been replaced with deuterium.

[0038] Compound

[0039] In one embodiment, the compound of the present invention is a compound represented by formula (I):

[0040]

[0041] or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein

[0042] R1 is selected from hydroxyl, C 1-20 Alkoxy, mercapto, C 1-20 Alkylthio, C 1-20 Alkanoyloxy, C 2-20 Alkoxyalkoxy, C 1-20 Haloalkoxy, NR a R b , where R a With R b are each independently selected from hydrogen or C 1-20 alkyl;

[0043] R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl, C 2-20 Alkenyl, C 2-20 Alkynyl; or R2 and R3 are linked to form 1,3-dioxolane or 1,4-dioxanyl;

[0044] A is selected from NR c or a heterocyclic group having 5 to 14 ring atoms, wherein R c Select from H or C 1-20 alkyl;

[0045] B is selected from C1-C 20 Alkylene, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic or -XC 2-20 Alkenylene-, wherein X is selected from C6-C 14 Aryl, heteroaryl having 5 to 14 ring atoms, C3-C 14 a carbocyclic group or a heterocyclic group having 5 to 14 ring atoms; 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms or C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy;

[0046] D is selected from -C(O)NHR d , where R d Selected from hydroxyl, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic group, wherein the C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, hydroxy, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

[0047] In one embodiment, the compound of the present invention is a compound represented by formula (I) or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein

[0048] R1 is selected from hydroxyl, C 1-20 Alkoxy, mercapto, C 1-20 Alkylthio, C 1-20 Haloalkoxy;

[0049] R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl, C 2-20 Alkenyl, C 2-20 Alkynyl; or R2 and R3 are linked to form 1,3-dioxolane or 1,4-dioxanyl;

[0050] A is selected from NRc or a heterocyclic group having 5 to 14 ring atoms, wherein Rc is selected from H or C 1-20 alkyl;

[0051] B is selected from C1-C 20 Alkylene, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic or Wherein X is selected from C6-C 14 Aryl, heteroaryl having 5 to 14 ring atoms, C3-C 14 a carbocyclic group or a heterocyclic group having 5 to 14 ring atoms; 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms or C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy;

[0052] D is selected from -C(O)NHR d , where R d Selected from hydroxyl, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic group, wherein the C6-C 14aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, hydroxy, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

[0053] In one embodiment, the compound of the present invention is a compound represented by formula (I) or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein

[0054] R1 is selected from hydroxyl, C 1-20 Alkoxy, C 1-20 Haloalkoxy;

[0055] R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl; or R2 and R3 are connected to form 1,3-dioxolane or 1,4-dioxanyl;

[0056] A is selected from NR c or a 5-membered or 6-membered heterocyclic group, wherein R c Select from H or C 1-20 alkyl;

[0057] B is selected from C1-C 20 alkylene, phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclic group, C5-C6 carbocyclic group or wherein X is selected from phenyl, 5-membered or 6-membered heteroaryl, C5-C6 carbocyclyl or 5-membered or 6-membered heterocyclyl, which is optionally substituted by one or more groups selected from the following: halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy;

[0058] D is selected from -C(O)NHR d , where R d Selected from hydroxyl, phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclyl, C5-C6 carbocyclyl, wherein the phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclyl, C5-C6 carbocyclyl is optionally substituted by one or more groups selected from the following: halogen, hydroxyl, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

[0059] In one embodiment, the compound of the present invention is a compound represented by formula (I) or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein

[0060] R1 is selected from hydroxyl, methoxy;

[0061] R2 and R3 are each independently selected from hydrogen, methyl; or R2 and R3 are connected to form

[0062] A is selected from NH,

[0063] B is selected from C1-C 20 Alkylene, phenyl, furyl, thienyl, or Wherein X is selected from phenyl, furanyl, thienyl;

[0064] D is selected from -C(O)NHR d , where R d Selected from hydroxyl, phenyl; wherein the phenyl is optionally substituted by one or more groups selected from the following: halogen, hydroxyl, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

[0065] In one embodiment, the compound of the present invention is a compound represented by formula (II)

[0066]

[0067] or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein

[0068] R1, R2, R3, A, B and D are as defined herein.

[0069] In one embodiment, the compound of the present invention is a compound represented by formula (III) or formula (IV):

[0070]

[0071] or its stereoisomers, tautomers, solvates, prodrugs, isotope labels, and pharmaceutically acceptable salts, wherein R4 is selected from hydrogen or C 1-20 Alkyl, A, B and D are as defined herein.

[0072] In one embodiment, the compounds described in the present invention include, but are not limited to:

[0073]

[0074]

[0075] or stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts thereof.

[0076] Pharmaceutical compositions and administration

[0077] The pharmaceutical composition provided by the present invention comprises the compound of the present invention or its stereoisomer, tautomer, solvate, prodrug, isotope label, pharmaceutically acceptable salt, and at least one pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers are known to those skilled in the art, including diluents, lubricants, disintegrants, adhesives, buffers, preservatives, stabilizers, wetting agents, glidants, emulsifiers, colorants, flavoring agents, sweeteners, etc. According to the different routes of administration of the drug, such as oral administration, parenteral administration and rectal administration, the pharmaceutical composition of the present invention can be made in solid form (including but not limited to tablets, capsules, pills, granules, powders, powders, suppositories) or liquid form (including but not limited to solutions, suspensions, emulsions, tinctures, syrups). When the pharmaceutical composition of the present invention is in solid form, the pharmaceutically acceptable carrier generally includes one or more of the following: a) diluents, such as lactose, glucose, sucrose, mannitol, sorbitol, cellulose, etc.; b) lubricants, such as silicon dioxide, talc, stearic acid, polyethylene glycol, etc.; c) binders, such as magnesium aluminosilicate, gelatinized starch, gelatin, tragacanth gum, methylcellulose, sodium carboxymethylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, etc.; d) disintegrants, such as starch, alginic acid, agar, corn starch; e) stabilizers, such as antioxidants such as ascorbic acid; f) glidants, such as silicon dioxide; g) flavoring agents, such as mint, methyl salicylate; sweeteners, such as sucrose, saccharin. When the pharmaceutical composition of the present invention is in liquid form, the pharmaceutically acceptable carrier generally includes one or more of the following: a) a diluent, such as water for injection, physiological saline, Ringer's solution, polyethylene glycol, glycerol, propylene glycol, etc.; b) an antioxidant, such as ascorbic acid or sodium bisulfite; c) a buffer, such as acetate, phosphate, etc.

[0078] The effective dose of the compounds of the present invention depends at least on the nature, extent, delivery method and pharmaceutical dosage form of the condition being treated, and will be ultimately determined by the clinician. It can be expected that about 0.0001 to about 100 mg per kilogram of body weight per day; generally about 0.01 to about 10 mg per kilogram of body weight per day; more typically, about 0.01 to about 5 mg per kilogram of body weight per day; most typically about 0.05 to about 0.5 mg per kilogram of body weight per day. For example, the daily candidate dose for an adult of about 70 kg body weight will be in the range of 1 mg to 1000 mg, preferably in the range of 5 mg to 500 mg, and can be administered in the form of a single dose or multiple doses.

[0079] Indications

[0080] The compounds of the present invention have significant anti-cancer activity and can be used to treat or prevent cancer-related diseases, including but not limited to hematological tumors, lung cancer, liver cancer, colon cancer or multiple myeloma.

[0081] General synthetic method

[0082] Compounds of the present invention can be prepared using methods disclosed herein and their modified approaches and methods well known in the art. Typical embodiments of compounds according to the present invention can be synthesized using the following reaction schemes. It is apparent from the description herein that corresponding different products can be obtained by replacing reaction raw materials with other materials having similar structures. Reaction raw materials are typically obtained from commercial sources or synthesized using disclosed methods.

[0083] Example 1 Synthesis of Compound 6-Hydroxy-N-((7-Hydroxyamino)7-oxoheptyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-1)

[0084]

[0085] Step 1: Synthesis of (E)-2-(4-acetoxyphenyl)-3-(3,4-dimethoxyphenyl)acrylic acid (3):

[0086]

[0087] Add p-hydroxyphenylacetic acid (1,2.0 g, 13.14 mmol) and 3,4-dimethoxybenzaldehyde (2,2.18 g, 13.14 mmol) to a 100 mL dry round-bottom flask, dissolve with 10 mL acetic anhydride, add triethylamine (1.83 mL, 13.14 mmol) under stirring at room temperature, heat to 120 ° C and reflux for 24 h. TLC (V (petroleum ether): V (ethyl acetate) = 1:1) to monitor the reaction progress. After the reaction is complete, cool to room temperature, add 20 mL of water, precipitate a yellow solid, filter, wash the filter cake with ice ethanol (10 mL × 3), and dry at 50 ° C for 3 h to obtain 3.56 g of a yellow solid with a yield of 76.3%. 1H NMR (600MHz, DMSO-d6) δ12.58(s,1H,-COOH),7.72(s,1H,Ar-CH=),7.24(d,J=8.4Hz,2H,Ar-H),7.18(d,J=8.4Hz,2H,Ar- H),6.85-6.91(m,2H,Ar-H),6.42(d,J=1.3Hz,1H,Ar-H),3.72(s,3H,-OCH3),3.31(s,3H,-OCH3),2.29(s,3H,-CH3); HRMS C 19 H 18 O6[MH] - : Theoretical value 341.1025, measured value 341.1027.

[0088] Step 2: Synthesis of methyl (E)-3-(3,4-dimethoxyphenyl)-2-(4-hydroxyphenyl)acrylate (4)

[0089]

[0090] Compound 3 (3.0 g, 8.42 mmol) and 15 mL of methanol were added to a 100 mL round-bottom flask. After stirring and dissolving, the temperature was lowered to 0°C, and then 1.34 mL of concentrated sulfuric acid (25.25 mmol) was slowly added dropwise. After the addition was completed, the temperature was raised to 70°C and refluxed for 5 to 6 hours. The reaction progress was monitored by TLC (V (petroleum ether): V (ethyl acetate) = 1:1). After the reaction was complete, the mixture was concentrated under reduced pressure, 25 mL of water was added, and the solid was precipitated. The solid was filtered under reduced pressure and the filter cake was dried (50°C, 3 hours) to obtain 2.56 g of a light yellow solid with a yield of 97.4%. 1 H NMR(600MHz,DMSO-d6)δ9.56(s,1H,Ar-OH),7.67(s,1H,Ar-CH=),6.99(d,J=8.4Hz,2H,Ar-H),6.86-6.80(m HRMS C 18 H 18 O5[MH] - : Theoretical value 313.1076, measured value 313.1073.

[0091] Step 3: Synthesis of methyl 6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxylate (5)

[0092]

[0093] Compound 4 (2.56 g, 8.15 mmol) and 10 mL of trifluoroacetic acid were added to a 100 mL round-bottom flask. After complete dissolution, manganese dioxide (1.42 g, 16.30 mmol) was added three times. After the addition was complete, the reaction was allowed to react at room temperature for 12 h, and the reaction progress was monitored by TLC (V (petroleum ether): V (ethyl acetate) = 1:1). After the reaction was complete, 15 mL of acetic anhydride was added and the temperature was raised to 40 ° C and stirred for 2-3 h. The reaction was detected by TLC (V (petroleum ether): V (ethyl acetate) = 1:1). 40 mL of water was added and the temperature was raised to 70 ° C and the reaction was allowed to react for 2-3 h. The reaction was detected by TLC (V (petroleum ether): V (ethyl acetate) = 1:1). After cooling to room temperature, the reaction solution was extracted with dichloromethane (20 mL×5), and the organic phase was washed with saturated sodium bicarbonate solution (20 mL×2) and saturated sodium chloride solution (20 mL×2), dried over anhydrous Na2SO4, concentrated under reduced pressure, and purified by silica gel column chromatography (V(petroleum ether): V(ethyl acetate)=1:1) to obtain 1.57 g of yellow solid with a yield of 62.6%. 1 H NMR (600MHz, DMSO-d6) δ9.94(s,1H,Ar-OH),8.69(d,J=9.1Hz,1H,Ar-H),8.27(s,1H,Ar-H),8.05(d,J=2.2Hz,1H,Ar-H),7.95(s,1 H,Ar-H),7.58(s,1H,Ar-H),7.21(dd,J=9.1,2.3Hz,1H,Ar-H),4.05(s,3H,-OCH3),3.95(s,3H,-OCH3),3.93(s,3H,-COOCH3); HRMS C 18 H 16 O5[M+H] + : Calculated value 313.1076, measured value 313.1072.

[0094] Step 4: Synthesis of 6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxylic acid (6)

[0095]

[0096] Compound 5 (0.6 g, 2 mmol), 20 mL of methanol, and 10% NaOH (0.64 g, 16.05 mmol) aqueous solution were added to a 100 mL round-bottom flask and reacted at 75 °C for 4-5 h. TLC (V (petroleum ether): V (ethyl acetate) = 1:1) monitored the reaction to be complete. The reaction solvent was removed by rotary evaporation, and 2 mol / L hydrochloric acid solution was added dropwise at 0 °C for acidification. The mixture was filtered under reduced pressure and dried at 50 °C for 4 h to obtain 0.48 g of a brown solid with a yield of 83.8%. 1H NMR(400MHz,DMSO-d6)δ8.82(d,J=8.9Hz,1H,Ar-H),8.19(s,1H,Ar-H),8.10(s,1H,Ar-H),7.96(s,1H HRMS C 17 H 14 O5[MH] - Theoretical value 297.0763, measured value 297.0762.

[0097] Step 5: Synthesis of methyl 7-(6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamido)heptanoate (7)

[0098]

[0099] 0.48g of compound 6 was added to a 50mL round-bottom flask, and 15mL of N,N-dimethylformamide was added at room temperature. After stirring and dissolving, HATU (0.74g, 1.93mmol), HOBt (0.44g, 3.22mmol) and N-methylmorpholine (0.49g, 4.83mmol) were added. After reacting for 20min, 7-aminoheptanoic acid methyl ester hydrochloride (0.38g, 1.93mmol) was added. The reaction was continued for 2h. After TLC (V (dichloromethane): V (methanol) = 20:1) monitoring, the reaction was complete. The organic phase was extracted with dichloromethane (20mL×2), washed with 2mol / L dilute hydrochloric acid solution (20mL×2), saturated sodium bicarbonate solution (20mL×2) and saturated sodium chloride solution (20mL×2), dried over anhydrous sodium sulfate for 2h, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain 0.32g of white solid.

[0100] Step 6: Synthesis of 6-hydroxy-N-((7-hydroxyamino)7-oxoheptyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-1)

[0101]

[0102] Sodium hydroxide (0.23 g, 5.75 mmol) and 0.5 mL of hydroxylamine solution were added to a 50 mL round-bottom flask, stirred at room temperature until dissolved, and 10 mL of a mixed solution of V (tetrahydrofuran): V (methanol) = 1:1 was added. The reaction was continued at room temperature for 1 h. TLC (V (dichloromethane): V (methanol) = 10:1) detected that the reaction was complete. The solvent was evaporated under reduced pressure, 15 mL of water was added, and the pH of the solution was adjusted to 6-7 with 2 mol / L hydrochloric acid. The precipitate was filtered, dried, and purified by silica gel column chromatography to obtain 0.12 g of a light yellow solid with a yield of 37.5%. 1 H NMR(400MHz,DMSO-d6)δ10.35(s,1H,NH-OH),9.81(s,1H,Ar-OH),8.66(s,1H,NH-OH),8.48(t,J=5.7Hz, 1H,CO-NH),8.05(d,J=8.9Hz,1H,Ar-H),7.98(d,J=2.5Hz,1H,Ar-H),7.92(s,1H,Ar-H),7.61(s,1H,Ar- H),7.47(s,1H,Ar-H),7.11(dd,J=8.9,2.4Hz,1H,Ar-H),4.02(s,3H,-OCH3),3.92(s,3H,-OCH3),1.98( t,J=7.2Hz,2H,-CH2-),1.54(m,J=15.3,7.2Hz,5H,-CH2-),1.35(m,J=19.7,6.8,6.1Hz,5H,-CH2-); HRMS C 24 H 28 N2O6[M+H] + : Theoretical value 441.2026, measured value 441.2021.

[0103] Example 2 Synthesis of Compound 6-Hydroxy-N-((6-Hydroxyamino)6-oxohexyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-2)

[0104]

[0105] HD-2 (Example 2)

[0106] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced by 6-aminohexanoic acid methyl ester hydrochloride to obtain 0.11 g of HD-2 (Example 2) as an off-white solid with a yield of 34.8%. 1H NMR (400MHz, DMSO-d6) δ10.34(s,1H,NH-OH),8.49(t,J=5.6Hz,1H,CO-NH),8.04(d,J=9.1Hz,1H ,Ar-H),8.02(d,J=2.3Hz,1H,Ar-H),7.93(s,1H,Ar-H),7.60(s,1H,Ar-H),7.47(s,1H,Ar-H),7 .12(dd,J=8.9,2.3Hz,1H,Ar-H),4.02(s,3H,-OCH3),3.92(s,3H,-OCH3),3.18(s,1H,-CH2-),2 .00(t,J=7.4Hz,2H,-CH2-),1.74(s,1H,-CH2-),1.57(m,4H,-CH2-),1.36(m,2H,-CH2-); HRMSC 23 H 26 N2O6[M+H] + : Theoretical value 427.1869, measured value 427.1862.

[0107] Example 3 Synthesis of compound 6-hydroxy-N-((4-hydroxyamino)4-oxobutyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-3):

[0108]

[0109] HD-3 (Example 3)

[0110] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced by 4-aminobutyric acid methyl ester hydrochloride to obtain 0.10 g of HD-3 (Example 3) as an off-white solid with a yield of 33.6%. 1 H NMR (400MHz, DMSO-d6) δ10.44(s,1H,NH-OH),9.84(s,1H,Ar-OH),8.75(s,1H,NH-OH),8.53(t,J= 5.6Hz,1H,CO-NH),8.06(d,J=8.9Hz,1H,Ar-H),7.99(d,J=2.4Hz,1H,Ar-H),7.93(s,1H,Ar-H),7. 64(s,1H,Ar-H),7.46(s,1H,Ar-H),7.12(dd,J=9.0,2.4Hz,1H,Ar-H),4.02(s,3H,-OCH3),3.92(s ,3H,-OCH3),3.31(t,J=6.5Hz,2H,-CH2-),2.10(t,J=7.5Hz,2H,-CH2-),1.81(m,2H,-CH2-); HRMS C21 H 22 N2O6[M+Na] + : Theoretical value 421.1375, measured value 421.1376.

[0111] Example 4 Synthesis of Compound 6-Hydroxy-N-((4-hydroxycarbamoyl)benzyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-4)

[0112]

[0113] HD-4 (Example 4)

[0114] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced by 4-aminomethylbenzoic acid methyl ester hydrochloride to obtain 0.14 g of HD-4 (Example 4) as a brown solid with a yield of 42.7%. 1 H NMR (600MHz, DMSO-d6) δ11.21(s,1H,NH-OH),9.84(s,1H,Ar-OH),9.12(t,J=6.1Hz,1H,CO-NH),9. 02(s,1H,NH-OH),8.08(d,J=8.8Hz,1H,Ar-H),7.99(d,J=2.3Hz,1H,Ar-H),7.93(s,1H,Ar-H),7.8 0-7.74(m,2H,Ar-H),7.72(s,1H,Ar-H),7.49(d,J=2.5Hz,2H,Ar-H),7.48(s,1H,Ar-H),7.11(m,J =8.9,2.0Hz,1H,Ar-H),4.57(d,J=6.0Hz,2H,-CH2-),4.02(s,3H,-OCH3),3.91(s,3H,-OCH3); HRMS C 19 H 18 O6[M+H] + : Theoretical value 447.1556, measured value 447.1547.

[0115] Example 5 Synthesis of Compound N-hydroxy-5-((6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide)methyl)furan-2-carboxamide (HD-5)

[0116]

[0117] HD-5 (Example 5)

[0118] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced by 5-(aminomethyl)-2-furoic acid methyl ester hydrochloride to obtain 0.10 g of HD-5 (Example 5) as a light yellow solid with a yield of 32.7%. 1 H NMR (400MHz, DMSO-d6) δ9.07 (t, J=5.7Hz, 1H, CO-NH), 8.08 (d, J=8.9Hz, 1H, Ar-H), 8.02(d,J=2.4Hz,1H,Ar-H),7.93(s,1H,Ar-H),7.69(s,1H,Ar-H),7.48(s,1H,Ar-H ),7.14(dd,J=9.0,2.2Hz,1H,Ar-H),7.05-6.95(m,1H,Ar-H),6.49(d,J=3.3Hz,1H ,Ar-H),4.57(d,J=5.6Hz,2H,-CH2-),4.02(s,3H,-OCH3),3.92(s,3H,-OCH3); HRMS C 23 H 20 N2O7[M+H] + : Theoretical value 437.1349, measured value 437.1342.

[0119] Example 6 Synthesis of Compound N-hydroxy-5-((6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide)methyl)thiophene-2-carboxamide (HD-6)

[0120]

[0121] HD-6 (Example 6)

[0122] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced by 5-(aminomethyl)thiophene-2-carboxylic acid methyl ester hydrochloride to obtain 0.11 g of HD-6 (Example 6) as a light red solid with a yield of 33.5%. 1H NMR (400MHz, DMSO-d6) δ9.93 (s, 1H, Ar-OH), 9.25 (t, J = 6.0Hz, 1H, CO-NH), 8.10 (d, J =8.9Hz,1H,Ar-H),8.01(d,J=2.4Hz,1H,Ar-H),7.94(s,1H,Ar-H),7.67(s,1H,Ar-H) ,7.49(d,J=6.3Hz,2H,Ar-H),7.12(dd,J=8.9,2.3Hz,1H,Ar-H),7.08(d,J=3.7Hz,1 H, Ar-H), 4.69 (d, J = 5.9Hz, 2H, -CH2-), 4.03 (s, 3H, -OCH3), 3.92 (s, 3H, -OCH3); HRMS C 23 H 20 N2O6S[M+Na] + : Theoretical value 475.0940, measured value 475.0947.

[0123] Example 7 Synthesis of Compound N-Hydroxy-5-(4-(6-Hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide)piperazine-1-yl)pentanamide (HD-7)

[0124]

[0125] HD-7 (Example 7)

[0126] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced with 5-(piperazine-1-yl)pentanoic acid methyl ester hydrochloride to obtain 0.11 g of HD-7 (Example 7) as a white solid with a yield of 30.8%. NMR (400MHz, DMSO-d6) δ10.34(s,1H,NH-OH),9.92(s,1H,Ar-OH),8.65(s,1H,NH-OH),8.01(d,J=2.4Hz,1H,Ar-H),7.93(s,1 H,Ar-H),7.60(d,J=8.8Hz,1H,Ar-H),7.45(s,1H,Ar-H),7.44(s,1H,Ar-H),7.15(dd,J=8.8,2.3Hz,1H,Ar-H),4.01(s,3H,-O CH3),3.91(s,3H,-OCH3),3.77(t,J=17.0Hz,2H,-CH2-),3.16(d,J=19.8Hz,2H,-CH2-),1.99(s,2H,-CH2-),1.95(t,J=7.2H z,2H,-CH2-),1.91(s,2H,-CH2-),1.52-1.45(m,2H,-CH2-),1.40(t,J=7.5Hz,2H,-CH2-),1.18(t,J=7.2Hz,2H,-CH2-); HRMS C 26 H 31 N3O6[M+H] + : Theoretical value 482.2291, measured value 482.2280.

[0127] Example 8 Synthesis of Compound (E)-6-Hydroxy-N-((5-(3-(Hydroxyamino)-3-oxopropen-1-yl)thiophen-2-yl)methyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-8)

[0128]

[0129] HD-8 (Example 8)

[0130] Referring to the synthesis method of step 5 in Example 1, 7-aminoheptanoic acid methyl ester hydrochloride was replaced with (E)-3-(5-(aminomethyl)thiophen-2-yl)acrylate methyl ester hydrochloride to obtain 0.13 g of HD-8 (Example 8) as a light red solid with a yield of 37.4%. 1H NMR (400MHz, DMSO-d6) δ10.69(s,1H,NH-OH),9.85(s,1H,Ar-OH),9.23(t,J=6.0Hz,1H,CO-NH),9.04(s,1H,NH-OH), 8.11(d,J=8.9Hz,1H,Ar-H),8.00(d,J=2.4Hz,1H,Ar-H),7.93(s,1H,Ar-H),7.67(s,1H,Ar-H),7.58(d,J=15.5Hz,1 H,-CH=CH-),7.49(s,1H,Ar-H),7.26(d,J=3.6Hz,1H,Ar-H),7.13(dd,J=8.9,2.3Hz,1H,Ar-H),7.06(d,J=3.6Hz,1H HRMS C 25 H 22 N2O6S[M+H] + : Theoretical value 479.1277, measured value 479.1285.

[0131] Example 9 Synthesis of Compound (E)-6-Hydroxy-N-((5-(3-(Hydroxyamino)-3-oxopropen-1-yl)furan-2-yl)methyl)-2,3-dimethoxyphenanthrene-9-carboxamide (HD-9)

[0132]

[0133] HD-9 (Example 9)

[0134] Referring to the synthesis method of Example 1, 7-aminoheptanoic acid methyl ester hydrochloride in step 5 was replaced with (E)-3-(5-(aminomethyl)furan-2-yl)acrylate methyl ester hydrochloride to obtain 0.11 g of HD-9 (Example 9) as a light red solid with a yield of 33.7%. 1H NMR(400MHz,DMSO-d6)δ9.09(t,J=5.7Hz,1H,CO-NH),8.11-8.00(m,2H,Ar-H),7.93(s,1H, Ar-H),7.67(s,1H,Ar-H),7.49(s,1H,Ar-H),7.22(d,J=15.5Hz,1H,-CH=CH-),7.14(d,J=8 .9Hz,1H,Ar-H),6.72(d,J=3.2Hz,1H,Ar-H),6.48(d,J=3.3Hz,1H,-Ar-H),6.30(d,J=15.6 Hz,1H,-CH=CH-),4.57(d,J=5.6Hz,2H,-CH2),4.02(s,3H,-OCH3),3.91(s,3H,-OCH3); C 25 H 22 N2O7[M+H] + : Theoretical value 463.1505, measured value 463.1501.

[0135] Example 10 Synthesis of Compound N-(4-((2-aminophenyl)carbamoyl)benzyl)-6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide (HD-10)

[0136] Step 1: Synthesis of methyl 4-((6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamido)methyl)benzoate (1)

[0137]

[0138] At room temperature, the intermediate 6 of Example 1 (0.45 g, 1.01 mmol) was dissolved in 15 mL of N,N-dimethylformamide, and HATU (0.39 g, 1.01 mmol), HOBt (0.27 g, 2.02 mmol) and N-methylmorpholine (0.31 g, 3.03 mmol) were added. After reacting for 20 min, methyl 4-aminomethylbenzoate (0.20 g, 1.21 mmol) was added, and the reaction was continued for 2 h. TLC (V (dichloromethane): V (methanol) = 20: 1) monitored the reaction to be complete. The mixture was extracted with dichloromethane (20 mL × 2), and the organic phase was washed with 2 mol / L dilute hydrochloric acid solution (20 mL × 2), saturated sodium bicarbonate solution (20 mL × 2) and saturated sodium chloride solution (20 mL × 2), dried over anhydrous sodium sulfate for 2 h, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain 0.19 g of a white solid with a yield of 42.3%.

[0139] Step 2: Synthesis of N-(4-((2-aminophenyl)carbamoyl)benzyl)-6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide (HD-10)

[0140]

[0141] At room temperature, the intermediate 1 of Example 10 (0.45 g, 1.01 mmol) was dissolved in 15 mL of N,N-dimethylformamide, and HATU (0.39 g, 1.01 mmol), HOBt (0.27 g, 2.02 mmol) and N-methylmorpholine (0.31 g, 3.03 mmol) were added. After reacting for 20 min, o-phenylenediamine (0.13 g, 1.21 mmol) was added, and the reaction was continued for 2 h. TLC (V (dichloromethane): V (methanol) = 20: 1) monitored the reaction to be complete. The mixture was extracted with dichloromethane (20 mL × 2), and the organic phase was washed with 2 mol / L dilute hydrochloric acid solution (20 mL × 2), saturated sodium bicarbonate solution (20 mL × 2) and saturated sodium chloride solution (20 mL × 2), dried over anhydrous sodium sulfate for 2 h, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain 0.19 g of a white solid with a yield of 36.1%. 1 H NMR(600MHz,DMSO-d6)δ9.86(s,1H,NH-OH),9.67(s,1H,Ar-OH),9.18(t,J=6.1Hz,1H,CO-NH),8.10(d,J=8.9Hz,1H,Ar- H),8.06-7.98(m,3H,Ar-H),7.94(s,1H,Ar-H),7.74(s,1H,Ar-H),7.56(d,J=7.9Hz,2H,Ar-H),7.50(s,1H,Ar-H),7.20 (d,J=7.8Hz,1H,Ar-H),7.13(dd,J=9.0,2.4Hz,1H,Ar-H),6.98(t,J=7.4Hz,1H,Ar-H),6.80(d,J=8.0Hz,1H,Ar-H),6.6 HRMS C 31 H 27 N3O5[M+H] + : Theoretical value 522.2029, measured value 522.2028.

[0142] Embodiment 11

[0143] Synthesis of compound N-(4-((2-amino)fluorophenyl)benzyl)-6-hydroxy-2,3-dimethoxyphenanthrene-9-carboxamide (HD-11):

[0144]

[0145] HD-11 (Example 11)

[0146] Referring to the synthesis method of Example 10, the o-phenylenediamine in step 2 was replaced with 2-BOC-amino-4-fluoroaniline to obtain 0.15 g of HD-11 (Example 11), a white solid, with a yield of 27.7%. 1 H NMR (400MHz, DMSO-d6) δ9.98(s,1H,Ar-OH),9.64(s,1H,CO-NH),9.18(t,J=6.1Hz,1H,CO-NH),8.09(d,J=8. 9Hz,1H,Ar-H),8.02(d,J=2.8Hz,2H,Ar-H),8.01(s,1H,Ar-H),7.94(s,1H,Ar-H),7.73(s,1H,Ar-H),7.55( d,J=8.0Hz,2H,Ar-H),7.49(s,1H,Ar-H),7.19-7.10(m,2H,Ar-H),6.56(dd,J=11.3,2.9Hz,1H,Ar-H),6.37 (m,1H,Ar-H),5.26(s,2H,-NH2),4.62(d,J=6.0Hz,2H,-CH2-),4.03(s,3H,-OCH3),3.93(s,3H,-OCH3); C 31 H 26 FN3O5[M+H] + : Theoretical value 540.1935, measured value 540.1949.

[0147] Example 12 Synthesis of Compound N-(4-(Hydroxycarbamoyl)benzyl)-2,3,6-trimethoxyphenanthrene-9-carboxamide (HD-12):

[0148] Step 1 Synthesis of compound 2,3,6-trimethoxyphenanthrene-9-carboxylic acid methyl ester (intermediate 1)

[0149]

[0150] The intermediate 5 (0.3 g, 1.01 mmol) of Example 1, sodium hydride (73 mg, 3.03 mmol), and 10 mL of N,N-dimethylformamide were added to a 50 mL round-bottom flask, stirred at room temperature for 0.5 h, and then iodomethane (314 mg, 2.21 mmol) was added, and the reaction was continued at room temperature for 6 h. TLC (V (petroleum ether): V (ethyl acetate) = 2: 1) detected that the reaction was complete. 10 mL of ethanol was added dropwise to quench the reaction, and the mixture was extracted with dichloromethane (10 mL × 3). The organic phases were combined and washed with 2 mol / L hydrochloric acid solution (20 mL × 2), saturated sodium bicarbonate solution (20 mL × 2), and saturated sodium chloride solution (20 mL × 2) in sequence. The organic phases were dried over anhydrous sodium sulfate for 4 h, filtered, concentrated under reduced pressure, and purified by column chromatography to obtain 0.13 g of intermediate 1 as a white solid with a yield of 39.5%.

[0151] Step 2 Synthesis of compound N-(4-(hydroxycarbamoyl)benzyl)-2,3,6-trimethoxyphenanthrene-9-carboxamide (HD-12):

[0152]

[0153] HD-12 (Example 12)

[0154] Referring to the subsequent synthesis method of Example 4, 0.07 g of HD-12 (Example 12) was obtained as an off-white solid with a yield of 38.8%. 1 H NMR(400MHz, DMSO-d6)δ9.17(t,J=6.1Hz,1H,CO-NH),8.18(d,J=9.1Hz,1H,Ar-H),8.13( d,J=2.6Hz,1H,Ar-H),8.10(s,1H,Ar-H),7.93(d,J=7.8Hz,2H,Ar-H),7.80(s,1H,Ar-H), 7.53(s,1H,Ar-H),7.44(d,J=7.7Hz,2H,Ar-H),7.25(dd,J=9.1,2.5Hz,1H,Ar-H),4.58( d,J=6.0Hz,2H,-CH2-),4.06(s,3H,-OCH3),4.01(s,3H,-OCH3),3.93(s,3H,-OCH3); HRMS C 26 H 25 N2O6[M+H] + : Theoretical value 461.1710, measured value 461.1713.

[0155] Embodiment 13

[0156] Synthesis of Compound N-(7-(Hydroxyamino)-7-oxoheptyl)-2,3,6-trimethoxyphenanthrene-9-carboxamide (HD-13)

[0157]

[0158] HD-13 (Example 13)

[0159] Referring to the synthesis method of Example 12, 4-aminomethylbenzoic acid methyl ester hydrochloride was replaced with 7-aminoheptanoic acid methyl ester hydrochloride to obtain 0.06 g of HD-13 (Example 13), an off-white solid, with a yield of 30.7%. 1H NMR (400MHz, DMSO-d6) δ10.36 (d, J = 1.6 Hz, 1H, NH-OH), 8.67 (d, J = 1.6 Hz, 1H, NH-OH), 8.53 (t, J = 5.6 Hz, 1H, CO-NH), 8.16-8.11 (m, 2H, Ar-H), 8.09 (s, 1H, Ar-H), 7.69 (s, 1H, Ar-H), 7.5 1(s,1H,Ar-H),7.25(dd,J=9.1,2.5Hz,1H,Ar-H),4.06(s,3H,-OCH3),4.01(s,3H,-OCH3), 3.93(s,3H,-OCH3),2.00-1.94(m,2H,-CH2-),1.56(m,5H,-CH2-),1.35(m,5H,-CH2-); HRMS C 25 H 30 N2O6[M+H] + : Theoretical value 455.2182, measured value 455.2174.

[0160] Embodiment 14

[0161] Synthesis of compound 2-hydroxy-N-(4-(hydroxycarbamoyl)benzyl)phenanthroline[2,3-d][1,3]dioxolane-5-carboxamide (HD-14):

[0162] Step 1: Synthesis of compound 2,3-methylenedioxybenzaldehyde (2)

[0163]

[0164] Add 3,4-dihydroxybenzaldehyde (3.0g, 21.7mmol), potassium carbonate (6.0g, 43.5mmol), dichloromethane (7mL, 108.6mmol) and anhydrous N,N-dimethylformamide 15mL to a 100mL round-bottom flask, heat at 150℃ for 9h under nitrogen protection, and monitor the reaction completion by TLC (V(petroleum ether):V(ethyl acetate)=1:1). Add 30mL of water to quench the reaction, extract with dichloromethane several times, combine the organic phases, wash the organic phases with saturated sodium chloride solution (20mL×2), dry with anhydrous sodium sulfate for 4h, filter, concentrate under reduced pressure, and purify by column chromatography to obtain 2.43g of brown liquid with a yield of 74.8%.

[0165] Step 2: Synthesis of compound 2-hydroxy-N-(4-(hydroxycarbamoyl)benzyl)phenanthro[2,3-d][1,3]dioxolane-5-carboxamide (HD-14)

[0166]

[0167] HD-14 (Example 14)

[0168] The synthesis method of Example 4 was subsequently referred to to obtain 0.09 g of HD-14 (Example 14) as a yellow solid with a yield of 35.8%. 1 H NMR(400MHz,DMSO-d6)δ11.21(s,1H,NH-OH),9.89(s,1H,Ar-OH),9.14(t,J=6.0Hz,1H,CO-NH), 9.03(s,1H,NH-OH),8.10-8.00(m,2H,Ar-H),7.90(d,J=2.4Hz,1H,Ar-H),7.78(s,1H,Ar-H),7.7 6(d,J=1.8Hz,1H,Ar-H),7.69(s,1H,Ar-H),7.50(s,1H,Ar-H),7.48(s,1H,Ar-H),7.47(s,1H,A r-H),7.13(dd,J=8.9,2.3Hz,1H,Ar-H),6.21(s,2H,-CH2-,),4.58(d,J=6.0Hz,2H,-CH2-); HRMS C 24 H 18 N2O6[M+H] + : Theoretical value 431.1243, measured value 431.1238.

[0169] Embodiment 15

[0170] Synthesis of Compound 2-Hydroxy-N-(7-(Hydroxyamino)-7-oxoheptyl)phenanthro[2,3-d][1,3]dioxolane-5-carboxamide (HD-15)

[0171]

[0172] HD-15 (Example 15)

[0173] Referring to the synthesis method of Example 12, 4-aminomethylbenzoic acid methyl ester hydrochloride was replaced by 7-aminoheptanoic acid methyl ester hydrochloride to obtain 0.11 g of Example 15 as a yellow solid with a yield of 36.7%. 1 H NMR (400MHz, DMSO-d6) δ8.51(t,J=5.7Hz,1H,CO-NH),8.02-8.00(m,2H,Ar-H),7.92(d,J=2.3Hz,1H,Ar-H),7.56(s,1H,Ar-H),7.45(s,1H,Ar HRMS C 23 H 24 N2O6[M+H] + : Theoretical value 425.1713, measured value 425.1703.

[0174] Biological activity evaluation

[0175] 1. HDAC1 / 6 inhibitory activity test

[0176] 1. Experimental reagents and consumables

[0177] HDAC1, HDAC6 (active motif 31504 22117004);

[0178] SAHA (Selleck S1047 12);

[0179] ACY-1215 (Selleck S8001);

[0180] TUB-A(Selleck S2239)

[0181] OptiPlate-384 F (PerkinElmer 6007270 8240-17211).

[0182] 2. Instruments

[0183] Centrifuge (Manufacturer: Eppendorf, Model: 5430)

[0184] Microplate reader (manufacturer: Biotek, model: Synergy2)

[0185] 3. Experimental Methods

[0186] The HDAC pan-inhibitor SAHA and the clinically investigated HDAC6 selective inhibitors ACY-1215 and TUB-A were used as positive controls. The HDAC1 and HDAC6 inhibitory activities of the target molecules were determined using the fluorescence-based HDAC Activity Assay: DMSO solutions of the test compounds were prepared, and the buffer (50 mM Tris pH = 7.5, 0.01% Tween-20, 10 mM NaCl, 0.05% BSA), HDAC1 and HDAC6 enzyme solutions, and substrate / trypsin mixed solutions were prepared in sequence according to the instructions of the kit; the DMSO solutions, enzyme solutions, and mixed solutions of the compounds with different concentration gradients (0.32, 1.6, 8, 40, 200, 1000 nM) were added to the OptiPlate-384 well plate to prepare the catalytic reaction system, control wells were set, and the plates were incubated at room temperature for a period of time. The fluorescence signal values ​​were read using a Synergy microplate reader, and the inhibition rate, IC 50 Fitted by GraphPadPrism 5.

[0187]

[0188]

[0189] The data showed that the compounds of the present invention had good inhibitory activity against both HDAC1 and HDAC6, and some of the compounds were superior to the positive controls SAHA, ACY-1215 and TUB-A.

[0190] 2. Anti-tumor cell proliferation activity test

[0191] The antiproliferative activity of the compounds of the present invention on HCT116, A549, HEL, and MOLT-4 tumor cells was tested by CCK8 assay. HCT116, A549, HEL, and MOLT-4 tumor cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA).

[0192] 1. Experimental Reagents

[0193] Fetal bovine serum (10270-106) (GIBCO, Invitrogen Corporation, NY, USA); penicillin-streptomycin (V900929) (Sigma, St. Louis, MO, USA); McCoy's 5A medium (16600082); RPMI1640 medium (11875093) (GIBCO, Invitrogen Corporation, NY, USA); F-12K medium (30-2004) (ATCC, Rockville, MD, USA); trypsin (KGY0012) (KeyGEN, Nanjin, China); CCK8 kit (C3007) (Beyotime, Shanghai, China).

[0194] 2. Experimental Instruments

[0195] FORMA 700 ultra-low temperature refrigerator, Thermo Company; YC-300L medicine storage cabinet, Zhongke Meiling Cryogenic Technology Co., Ltd.; Direct-Q with pump ultrapure water meter, Millipore Company; SWCJ-2FD ultra-clean workbench: Suzhou Purification Equipment Co., Ltd.; 3K15 low-temperature high-speed centrifuge, Sigma Company; BS224 electronic balance: Beijing Sartorius Instrument System Co., Ltd.; Forma 3111 water-jacketed CO2 incubator: Thermo Electron company; YXQ-LS-50SⅡ vertical pressure steam sterilizer: Shanghai Boxun Industrial Medical Equipment Factory; Berthold LB941 microplate multifunctional microplate reader, Berthold Company.

[0196] 3. Experimental cell lines

[0197] HCT116 cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA);

[0198] A549 cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA);

[0199] HEL cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA);

[0200] MOLT-4 cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA).

[0201] 4. Experimental Methods

[0202] Subculture of HCT116 cells / A549 cells: The culture conditions are McCoy's 5A medium / 10% FBS F12K medium containing penicillin (final concentration of 100U / mL), streptomycin (final concentration of 100μg / mL) and 10% FBS. When the cells are fused to 90%, discard the old medium, wash the cells twice with 2mL PBS, discard PBS, add 2mL 0.25% trypsin-0.02% EDTA mixed digestion solution, observe under a microscope for about 30s, and when the cells become round, quickly add 2mL complete medium to terminate digestion, gently blow and collect cells. Centrifuge at 800rpm, 4℃ for 5min, discard the supernatant, resuspend the cells with complete medium, culture in bottles, and change the medium every other day.

[0203] Subculture of HEL cells / MOLT-4 cells: The culture conditions are PRMI-1640 medium containing penicillin (final concentration of 100U / mL), streptomycin (final concentration of 100μg / mL) and 10% FBS. When the cells are fused to 90%, centrifuge at 800rpm, 4℃ for 5min, discard the supernatant, resuspend the cells with complete medium, culture in bottles, and change the medium every other day.

[0204] HCT116 and A549 cells were collected by centrifugation and 1×10 4 / well were inoculated into 96-well plates, with 3 replicates per group. After the cells adhered to the wall, the corresponding culture medium was replaced according to the grouping situation. After 72 hours, CCK-8 was used for determination: PBS was rinsed 3 times, 100 μL of culture medium containing 10% CCK-8 working solution was added to each well, gently shaken evenly, and placed in a cell culture incubator for incubation. After incubation for 2 hours, the absorbance values ​​of each were detected at a wavelength of 450nm using an ELISA reader, and corresponding comparative analysis was performed.

[0205] HEL and MOLT-4 cells were collected by centrifugation and 1×10 4 / well were inoculated into 96-well plates, with 3 replicate wells in each group. The corresponding culture medium was added according to the grouping situation. After 72 hours, CCK-8 was used for determination: 10 μL CCK-8 solution was added to each well, gently shaken evenly, and placed in a cell culture incubator for incubation. After incubation for 2 hours, the absorbance values ​​of each were detected at a wavelength of 450nm by an ELISA reader, and corresponding comparative analysis was performed.

[0206] Table 1

[0207]

[0208] As shown in Table 1, HD-1 has good antiproliferative activity against various tumor cells, and the antiproliferative activity is even significantly better than that of the positive control SAHA.

[0209] Effects of Compounds on the Cell Cycle of Tumor Cells

[0210] 1. Experimental Reagents

[0211] Fetal bovine serum (SH30070.03) (FBS, Hyclone, Logan, UT, USA); Penicillin-streptomycin (15140148) (Thermo Fisher Scientific, Pittsburgh, PA, USA); RPMI1640 medium (11875093) (GIBCO, Invitrogen Corporation, NY, USA); Trypsin (15090046) (Gibco, Grand Island, NY, USA); Cell cycle kit (KGA512); Matrigel Matrix (354234) (BD Bioscience, USA).

[0212] 2. Experimental Instruments

[0213] FORMA 700 ultra-low temperature refrigerator, Thermo Company; YC-300L medicine storage cabinet, Zhongke Meiling Cryogenic Technology Co., Ltd.; Direct-Q with pump ultrapure water meter, Millipore Company; SWCJ-2FD ultra-clean workbench: Suzhou Purification Equipment Co., Ltd.; 3K15 low-temperature high-speed centrifuge, Sigma Company; BS224 electronic balance: Beijing Sartorius Instrument System Co., Ltd.; Forma 3111 water-jacketed CO2 incubator: Thermo Electron company; YXQ-LS-50S℃ vertical pressure steam sterilizer: Shanghai Boxun Industrial Medical Equipment Factory; Thermo Electron company; Beckman DxFlex flow cytometer, Beckman Company, USA; Olympus inverted phase contrast microscope, Olympus Corporation, Japan.

[0214] 3. Experimental cell lines

[0215] HEL cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA).

[0216] 4. Experimental Plan

[0217] HEL cells were incubated with RPMI1640 medium containing 0, 0.1, 0.5, and 2.5 μM of the target compound for 24 hours; cells from each group in the logarithmic growth phase were taken, the old medium was discarded, 0.5 mL of PBS was added to wash the cells twice, 2 mL of 0.25% trypsin and EDTA-free digestion solution were added after discarding PBS, and then 2 mL of complete medium was added to terminate digestion, and the cells were gently blown to collect them. 2000rpm, 4℃, centrifuge for 5min, discard the supernatant; collect and adjust the cell concentration to 1×106 / mL, take 1mL of single cell suspension; after centrifugation of the prepared single cell suspension, remove the supernatant, use 500μL of cold ethanol with a volume fraction of 70% to resuspend the cells, fix them, seal them with a sealing film, and fix them at 4℃ overnight; collect the fixed cells at 800rpm for 15min, wash them twice with PBS; resuspend the cells with 0.4mL PBS and transfer them to a tube and blow gently (to prevent cell breakage); add 500μLPI / RNase A staining working solution prepared in advance and stain in an ice bath away from light for 30min; filter with a 300-mesh (pore size 40-50μm) nylon mesh and detect on the machine;

[0218] 5. Experimental Results

[0219] The results are as follows Figure 1 As shown, compared with the control group, the cell cycle of HEL cells in each group was arrested at G2 / M

[0220] The effect was concentration-dependent, and the 2.5μM HD-1 group had the best effect.

[0221] IV. Transwell assay to investigate the migration and invasion ability of target compounds on cancer cells

[0222] 1. Experimental Reagents

[0223] Fetal bovine serum (SH30070.03) (FBS, Hyclone, Logan, UT, USA); Penicillin-streptomycin (15140148) (Thermo Fisher Scientific, Pittsburgh, PA, USA); RPMI1640 medium (11875093) (GIBCO, Invitrogen Corporation, NY, USA); Trypsin (15090046) (Gibco, Grand Island, NY, USA); Crystal violet stain (KGA229) (KeyGEN Biotech, NanJing, China); Matrigel Matrix (354234) (BD Bioscience, USA).

[0224] 2. Experimental Instruments

[0225] FORMA 700 ultra-low temperature refrigerator, Thermo Company; YC-300L medicine storage cabinet, Zhongke Meiling Cryogenic Technology Co., Ltd.; Direct-Q with pump ultrapure water meter, Millipore Company; SWCJ-2FD ultra-clean workbench: Suzhou Purification Equipment Co., Ltd.; 3K15 low-temperature high-speed centrifuge, Sigma Company; BS224 electronic balance: Beijing Sartorius Instrument System Co., Ltd.; Forma 3111 water-jacketed CO2 incubator: Thermo Electron company; YXQ-LS-50S℃□ vertical pressure steam sterilizer: Shanghai Boxun Industrial Medical Equipment Factory; Thermo Electron company; Beckman DxFlex flow cytometer, Beckman Company, USA; Olympus inverted phase contrast microscope, Olympus Company, Japan; Transwell chamber, BD Company.

[0226] 3. Experimental cell lines

[0227] HEL cells were purchased from American Tissue Culture Collection (ATCC, Rockville, MD, USA).

[0228] 4. Experimental plan

[0229] (1) Transwell assay to investigate the ability of target compounds to affect cancer cell migration:

[0230] Implantation of cells: The cells were digested and centrifuged routinely, resuspended in serum-free medium, counted, and the cell density was adjusted to 2.5×105 / mL with serum-free medium; the incubated chamber was removed, the upper chamber liquid was aspirated, 100 μL of single cell suspension was gently added to the upper chamber along the wall, three duplicate wells were set up in each well, 600 μL of medium containing 10% FBS was added to the lower chamber of the 24-well plate, and the plate was placed in a 37°C incubator for routine culture for 24 hours. Staining: After the cells in the chamber were routinely cultured in a 37°C incubator for 24 hours, the chamber was removed, the upper chamber was gently washed 3 times with room temperature PBS, and the lower chamber was immersed in PBS for 5 minutes each time. 700 μL of 4% polymethanol was added to the lower chamber of a new 24-well plate, and the washed chamber was placed in it for fixation for 30 minutes. Discard the methanol, add 700 μL of 0.1% crystal violet to the lower chamber, put the chamber back, let it stand for 15 minutes, rinse the upper chamber gently with water, and gently wipe off the cells that have not passed through the inner layer of the chamber with a wet cotton swab. Do not leave small water droplets to prevent membrane deformation. Invert and dry naturally or dry in a low-temperature oven. Cell counting: Use a slide and place the chamber upright; use an inverted microscope to count the cells that pass through the chamber, and randomly select the counting field of view.

[0231] (2) Transwell assay to investigate the invasive ability of target compounds on cancer cells

[0232] Preparation of glue: The day before the experiment, take the Matrigel glue from the -20℃ refrigerator to a 4℃ refrigerator to melt overnight. The day before the experiment, put the pipette tip in a -20℃ refrigerator to pre-cool. Take it out 30 minutes before the experiment and pre-cool it on ice. Dilute the cooled culture medium and Matrigel stock solution at a ratio of 1:9, gently blow and mix thoroughly, and place on ice. Spreading glue: Add 40μL of diluted Matrigel glue to the upper chamber of the chamber, shake it gently to spread it flat, and let it stand in a 37℃ incubator for 4h. After the Matrigel glue is fully solidified, aspirate the unsolidified liquid in the upper chamber, gently add 70μL of culture medium, and incubate in a 37℃ incubator for 30min to fully hydrate. Implantation of cells: Centrifuge each group of cells routinely, resuspend them in serum-free culture medium, count them, and adjust the cell density to 2.5×10 with serum-free culture medium. 5 / mL; take out the incubated chamber, suck out the liquid in the upper chamber, take 200μL of single cell suspension and gently add it to the upper chamber along the wall, set up three duplicate wells in each well, add 600μL of culture medium containing 10% FBS to the lower chamber of the 24-well plate, and place it in a 37℃ incubator for routine culture for 24h. Staining: After the cells in the chamber are routinely cultured in a 37℃□ incubator for 24h, take out the chamber, gently wash the upper chamber 3 times with room temperature PBS, and soak the lower chamber in PBS for 5min each time. Add 700μL of methanol to the lower chamber of a new 24-well plate, place the washed chamber in it and fix it for 30min. Discard the methanol, add 700μL of 1% crystal violet to the lower chamber, put the chamber back, let it stand for 30min, gently rinse the upper chamber with clean water, and gently wipe off the cells that have not passed through the inner layer of the chamber with a wet cotton swab. Do not leave small water droplets to prevent membrane deformation. Invert and dry naturally or dry in a low-temperature oven. Cell counting: Use a glass slide and place the chamber upright; use an inverted microscope to count the cells that pass through the chamber and randomly select the counting field.

[0233] like Figure 2 As shown, compared with the control group, the migration and invasion abilities of HEL cells in each group decreased, among which 0.01 μM HD-1 had the best effect.

Claims

1. Compound represented by formula (I): or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein R1 is selected from hydroxyl, C 1-20 Alkoxy, mercapto, C 1-20 Alkylthio, C 1-20 Alkanoyloxy, C 2-20 Alkoxyalkoxy, C 1-20 Haloalkoxy, NR a R b , where R a With R b are each independently selected from hydrogen or C 1-20 alkyl; R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl, C 2-20 Alkenyl, C 2-20 Alkynyl; or R2 and R3 are linked to form 1,3-dioxolane or 1,4-dioxanyl; A is selected from NR c or a heterocyclic group having 5 to 14 ring atoms, wherein R c Select from H or C 1-20 alkyl; B is selected from C1-C 20 Alkylene, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic or -XC 2-20 Alkenylene-, wherein X is selected from C6-C 14 Aryl, heteroaryl having 5 to 14 ring atoms, C3-C 14 a carbocyclic group or a heterocyclic group having 5 to 14 ring atoms; 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms or C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy; D is selected from -C(O)NHR d , where R d Selected from hydroxyl, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic group, wherein the C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, hydroxy, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

2. The compound according to claim 1 or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein R1 is selected from hydroxyl, C 1-20 Alkoxy, mercapto, C 1-20 Alkylthio, C 1-20 Haloalkoxy; R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl, C 2-20 Alkenyl, C 2-20 Alkynyl; or R2 and R3 are linked to form 1,3-dioxolane or 1,4-dioxanyl; A is selected from NRc or a heterocyclic group having 5 to 14 ring atoms, wherein Rc is selected from H or C 1-20 alkyl; B is selected from C1-C 20 Alkylene, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic or Wherein X is selected from C6-C 14 Aryl, heteroaryl having 5 to 14 ring atoms, C3-C 14 a carbocyclic group or a heterocyclic group having 5 to 14 ring atoms; 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms or C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy; D is selected from -C(O)NHR d , where R d Selected from hydroxyl, C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 Carbocyclic group, wherein the C6-C 14 aryl, heteroaryl having 5 to 14 ring atoms, heterocyclyl having 5 to 14 ring atoms, C3-C 14 The carbocyclic group is optionally substituted by one or more groups selected from the group consisting of halogen, hydroxy, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

3. The compound according to claim 1 or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein R1 is selected from hydroxyl, C 1-20 Alkoxy, C 1-20 Haloalkoxy; R2 and R3 are each independently selected from hydrogen, C 1-20 Alkyl; or R2 and R3 are connected to form 1,3-dioxolane or 1,4-dioxanyl; A is selected from NR c or a 5-membered or 6-membered heterocyclic group, wherein R c Select from H or C 1-20 alkyl; B is selected from C1-C 20 alkylene, phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclic group, C5-C6 carbocyclic group or wherein X is selected from phenyl, 5-membered or 6-membered heteroaryl, C5-C6 carbocyclyl or 5-membered or 6-membered heterocyclyl, which is optionally substituted by one or more groups selected from the following: halogen, oxo, C1-C 20 Alkyl, C1-C 20 Halogenated alkyl, C1-C 20 Alkoxy, C1-C 20 Alkanoyl, C1-C 20 Alkoxycarbonyl, C1-C 20 Alkanoyloxy, C1-C 20 Alkoxycarbonyloxy; D is selected from -C(O)NHR d , where R d Selected from hydroxyl, phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclyl, C5-C6 carbocyclyl, wherein the phenyl, 5-membered or 6-membered heteroaryl, 5-membered or 6-membered heterocyclyl, C5-C6 carbocyclyl is optionally substituted by one or more groups selected from the following: halogen, hydroxyl, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

4. The compound according to claim 1 or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein R1 is selected from hydroxyl, methoxy; R2 and R3 are each independently selected from hydrogen, methyl; or R2 and R3 are connected to form A is selected from NH, B is selected from C1-C 20 Alkylene, phenyl, furyl, thienyl, or Wherein X is selected from phenyl, furanyl, thienyl; D is selected from -C(O)NHR d , where R d Selected from hydroxyl, phenyl; wherein the phenyl is optionally substituted by one or more groups selected from the following: halogen, hydroxyl, amino, nitro, mercapto, cyano, C1-C6 alkyl, C1-C6 alkoxy.

5. The compound according to claim 1, which is a compound represented by formula (II) or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein R1, R2, R3, A, B and D are as defined according to any one of claims 1-4.

6. The compound according to claim 1, which is a compound represented by formula (III) or formula (IV): or its stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts, wherein, R4 is selected from hydrogen or C 1-20 Alkyl, A, B and D are as defined according to any one of claims 1-4.

7. The compound according to claim 1, which is selected from: or stereoisomers, tautomers, solvates, prodrugs, isotope-labeled substances, and pharmaceutically acceptable salts thereof.

8. A pharmaceutical composition comprising the compound according to any one of claims 1 to 7 or its stereoisomer, tautomer, solvate, prodrug, isotope-labeled substance, or pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

9. Use of a compound according to any one of claims 1 to 7 or its stereoisomer, tautomer, solvate, prodrug, isotope-labeled substance, or a pharmaceutically acceptable salt thereof and / or a pharmaceutical composition according to claim 8 in the preparation of a medicament for preventing and / or treating cancer diseases.

10. The use according to claim 9, wherein the cancer is selected from hematological tumors, lung cancer, liver cancer, colon cancer or multiple myeloma.

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