Substituted polycyclic compound

Abstract

The present application relates to a substituted polycyclic compound having biological inhibitory activity as represented by formula (I), and a preparation method therefor and a use thereof. The use includes a use of the compound of formula (I) in the preparation of a drug for treating MLL-related diseases. In the preparation process, the compound of the present invention is obtained by means of a series of reactions such as substitution, cyclization, reduction, and deprotection.

Description

Substituted polycyclic compounds

[0001] Cross-referencing

[0002] This application claims priority to Chinese Patent Application No. 202510104922.5, filed on January 22, 2025, entitled “Substituted Polycyclic Compounds,” the entire disclosure of which is incorporated herein by reference. Technical Field

[0003] This invention generally relates to novel substituted heterocyclic compounds having Menin-MLL interaction inhibitory activity, methods of their preparation, pharmaceutical compositions thereof, and also to the use of such compounds and pharmaceutical compositions thereof in treating diseases that benefit from inhibition of Menin-MLL interactions, such as acute leukemia. Background Technology

[0004] The occurrence of acute leukemia in humans is associated with genetic and epigenetic variations (Figueroa 2010; Rodríguez 2011). Approximately 5%–10% of children and adults with acute leukemia experience a chromosomal translocation in the 11q23 region of chromosome 11, resulting in a KMT2A (MLL1) rearrangement (Bill 2020). The 5-year survival rate for aggressive leukemia caused by MLL1 rearrangement (MLL-r) is only 35% (Dimartino 1999; Marshalek 2011). This translocation occurs in up to 80% of infants with acute leukemia (Brown 2013). MLL-r leukemia generally has a poor prognosis, especially infantile acute lymphoblastic leukemia, and to date, there is a lack of more effective and less toxic treatments.

[0005] MLL1 is a histone lysine-N-methyltransferase responsible for H3K4 methylation modification and essential for the expression of HOX family hematopoietic differentiation regulators. MLL1 rearrangement forms the MLL1 fusion protein, causing transcriptional aberrations that drive the development of acute leukemia (Meyer 2018). The MLL1 fusion protein regulates target gene expression through interactions with chromatin-associated protein complexes. Menin protein is a key MLL chaperone protein in this complex, playing a crucial role in the MLL fusion protein-driven target gene expression process, regulating the expression of MEIS1 and HOX family proteins (Chen 2006). Menin protein has a pocket on its surface that binds to MLL1 with high affinity to the conserved N-terminal sequence of MLL1. This interaction is crucial for the MLL1 fusion protein to regulate the transcription of target genes (Yokoyama 2005; Cassini 2007). Therefore, regulating the Menin-MLL interaction is an important target for small molecule therapy of MLL-r leukemia. Blocking the interaction between Menin and the MLL1 fusion protein using small molecule inhibitors has been shown to be a potential treatment option for MLL-r leukemia.

[0006] Currently, no Menin-MLL inhibitors have been approved for marketing, but six inhibitors have recently entered clinical trials overseas. Syndax's inhibitor SNDX-5613 (revumenib) first entered a Phase I / II clinical trial (NCT04065399) on August 22, 2019, targeting acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), mixed lineage acute leukemia (MLAL), mixed phenotype acute leukemia (MPAL), and unspecified lineage acute leukemia (ALAL) with MLL rearrangements or NPM1 mutations. Following closely behind, Kura's inhibitor KO-539 began a Phase I / II clinical trial on August 26, 2019 (NCT04067336); Janssen began a Phase I clinical trial of JNJ-75276617 on March 21, 2021 (NCT04065399); Daiichi Sankyo's inhibitor DS-1594 began a Phase I clinical trial on March 25, 2021; Biomea's inhibitor BMF-219 began a Phase I clinical trial on January 24, 2022; and Sumitomo's inhibitor DSP-5336 began a Phase I clinical trial on February 28, 2022. Domestically, Yehui Pharmaceuticals' BN104 began a Phase I clinical trial on September 4, 2023. Bayer, Agios, the University of Michigan, and the University of Pennsylvania also have patents related to this.

[0007] Recent findings from the AUGMENT-101 (NCT04065399) study indicate that patients develop acquired resistance to the Menin inhibitor revumenib after treatment (Perner 2023). Next-generation sequencing of bone marrow samples from resistant patients revealed somatic mutations in Menin. Mutation sites include M327V or M327I, T349M, G331R, and S160T, with mutation frequencies ranging from 5.9% to 28.2%. In a PDX model, continuous treatment of mice with the Menin inhibitor VTP-50469 resulted in Menin mutations in 68% of relapsed samples, including those at MEN1. M327V MEN1 M327I MEN1 G331R MEN1 G331D MEN1 T349M and MEN1 S160C The mutation sites are almost identical to those in patients. Crystal structure analysis shows that the three high-frequency mutant residues M327, G331, and T349 are close to the W346 residue, affecting the formation of hydrogen bonds between revumenib and the indole NH of W346, thus reducing the binding activity of revumenib to Menin protein, but not affecting the binding surface of Menin protein to MLL. Cellular and in vitro binding experiments also showed that the binding activity of the Menin mutant protein to revumenib is significantly reduced, thereby preventing the revumenib-mediated regulation of leukemia gene expression by the Menin-MLL1 protein complex.

[0008] Therefore, developing next-generation Menin inhibitors targeting Menin resistance mutations is of great clinical demand and will effectively overcome the problem of drug resistance relapse in patients. Using computer-aided drug design, we discovered structurally novel compounds and, through structural modification, obtained new compounds with strong potency and favorable pharmaceutical properties against Menin resistance mutations. Summary of the Invention

[0009] This invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug thereof.

[0010] in,

[0011] Ring A is a 5-8 member nitrogen-containing heterocycle or a nitrogen-containing heteroaromatic ring, and each X is independently C or N.

[0012] R 1a -C(=O)-NR 2a R 2b , Alternatively, a 5-6 membered heteroaryl group, said heteroaryl group comprising 1-3 nitrogen atoms and optionally comprising a carbonyl group, and said heteroaryl group optionally being surrounded by a 3-6 membered cycloalkyl group or a C-membered ring. 1-4 Alkyl substitution,

[0013] R 2a and R 2b Each independently is either H or C. 1-4 Alkyl group, wherein the alkyl group may optionally be substituted with -OH or -NH2.

[0014] R 1b For F or Cl,

[0015] R2 is selected from H, halogens, and C. 1-4 Alkyl, -OC 1-4 Alkyl and -NR 3a R 3b ,

[0016] R 3a and R 3b Each independently is either H or C. 1-4 alkyl,

[0017] X1 and X2 are each independently either N or CH.

[0018] L1 is -CH2-, -CH2-CH2-, or -CH2-CH2-CH2-.

[0019] L2 and L3 are each independently -CH2- or -CH2-CH2-.

[0020] R4 is C 1-6 Alkyl, 3-8 membered cycloalkyl, or 3-8 membered heterocycloalkyl

[0021] R3 is -C 1-6 Alkylene-NR 4a R 4b -C 1-6 Alkylene-C(O)-NR 5a R 5b -C 1-6 alkylene -OH or -C 1-6 Alkylene-NR7-C(O)-OC 1-4 Alkylene-OC(O)-C 1-4 Alkyl group, wherein the alkyl or alkylene group may optionally be converted to a halogen, -CN, -OH or -OC. l-4 Alkyl substitution,

[0022] R 4a and R 4b Each is independently selected from H and C. 1-6 Alkyl, -C(O)-C1-4 Alkyl, -C(O)-OC 1-4 Alkyl groups and -C(O)-NR 8a R 8b The C 1-6 Alkyl groups may optionally be replaced by halogens, -CN, -OH, or -S(O)2-C. 1-4 Alkyl, -OC 1-4 Alkyl, -C(O)-NR 6a R 6b Or -NR 6c -C(O)-C 1-4 Alkyl substitution,

[0023] R 5a R 5b、 R 6a R 6b、 R 6c R7, R 8a and R 8b Each is independently selected from H and C. 1-6 alkyl.

[0024] In some implementations... for or Preferred or

[0025] In some implementations, R 1a -C(=O)-NR 2a R 2b R 2a and R 2b Each independently is either H or C. 1-4 alkyl.

[0026] In some implementations, R 1a for

[0027] In some implementations, R 1b It is F.

[0028] In some implementations, R4 is isopropyl.

[0029] In some implementations, R2 is H or C. 1-4 Alkyl group, preferably H.

[0030] In some implementations, R3 is -C 1-6 Alkylene-NR 4a R 4b R 4a and R 4bEach was independently selected from C 1-6 Alkyl, the C 1-6 Alkyl groups may optionally be -OC 1-4 Alkyl substitution.

[0031] In some implementations, R3 is -C 1-6 Alkylene-N(C) 1-6 alkyl)-C 1-6 Alkylene-OC 1-4 alkyl.

[0032] In some implementations, R3 is

[0033] In some embodiments, the present invention provides a compound of formula (II) or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug thereof.

[0034] in,

[0035] R 2a and R 2b Each can be independently methyl, ethyl, or isopropyl.

[0036] X3 is either N or CR 10 ,

[0037] R 10 It can be F, Cl, or methyl.

[0038] In some implementations, R 2a For ethyl, R 2b It is isopropyl.

[0039] In some implementations, R 10 It can be F or Cl, preferably F.

[0040] In some implementations, X3 is CF.

[0041] In some embodiments, the present invention provides the following compounds or pharmaceutically acceptable salts, solvates, polymorphs, tautomers, metabolites, or prodrugs thereof.

[0042] In another aspect, the present invention provides a pharmaceutical composition comprising a compound of the present invention or a pharmaceutically acceptable salt, solvate, polymorph, or isomer thereof, and optionally comprising a pharmaceutically acceptable carrier.

[0043] On another front, the present invention provides a method for treating diseases associated with MLL activity, comprising administering to a subject a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug of the present invention, or a pharmaceutical composition of the present invention; in some embodiments, the disease associated with MLL activity is cancer, preferably acute leukemia (including MLL acute leukemia, MLL partial tandem repeat acute leukemia, NPM mutant acute leukemia, MOZ acute leukemia, NUP98 acute leukemia, and CALM acute leukemia), chronic lymphocytic leukemia, chronic myeloid leukemia, or myelodysplastic syndromes. Syndrome, polycythemia vera, malignant lymphoma (including B-cell lymphoma), myeloma (including multiple myeloma), brain tumor, head and neck cancer, esophageal cancer, thyroid cancer, small cell lung cancer, non-small cell lung cancer, breast cancer, gastric cancer, gallbladder and bile duct cancer, liver cancer, hepatocellular carcinoma, pancreatic cancer, colon cancer, rectal cancer, anal cancer, choriocarcinoma, endometrial cancer, cervical cancer, ovarian cancer, bladder cancer, urothelial carcinoma, kidney cancer, renal cell carcinoma, prostate cancer, testicular tumor, testicular germ cell tumor, ovarian germ cell tumor, Wilms' tumor, malignant melanoma, neuroblastoma, osteosarcoma, Ewing's sarcoma, chondrosarcoma, soft tissue sarcoma, or skin cancer.

[0044] The compounds of the present invention, or their pharmaceutically acceptable salts, solvates, polymorphs, tautomers, metabolites, or prodrugs, or the pharmaceutical compositions of the present invention, are used in combination with at least one different agent, wherein the different agent is selected from at least one agent selected from antitumor alkylating agents, antitumor antibiotics, plant-derived antitumor drugs, antitumor platinum coordination compounds, antitumor camptothecin derivatives, antitumor tyrosine kinase inhibitors, antitumor serine / threonine kinase inhibitors, antitumor phospholipid kinase inhibitors, antitumor monoclonal antibodies, interferons, biological response modifiers, hormone preparations, angiogenesis inhibitors, immune checkpoint inhibitors, epigenetic-related molecule inhibitors, protein post-translational modification inhibitors, proteasome inhibitors, and other antitumor drugs.

[0045] In another aspect, the present invention provides the use of the compounds of the present invention or pharmaceutically acceptable salts, solvates, polymorphs, tautomers, metabolites or prodrugs thereof, or pharmaceutical compositions of the present invention, in the preparation of a medicament for treating diseases associated with MLL activity; in some embodiments, the diseases associated with MLL activity are cancers, preferably acute leukemia (including MLL acute leukemia, MLL partial tandem repeat acute leukemia, NPM mutant acute leukemia, MOZ acute leukemia, NUP98 acute leukemia, and CALM acute leukemia), chronic lymphocytic leukemia, chronic myeloid leukemia, and myelodysplastic syndromes. Polycythemia vera, malignant lymphoma (including B-cell lymphoma), myeloma (including multiple myeloma), brain tumors, head and neck cancer, esophageal cancer, thyroid cancer, small cell lung cancer, non-small cell lung cancer, breast cancer, gastric cancer, gallbladder and bile duct cancer, liver cancer, hepatocellular carcinoma, pancreatic cancer, colon cancer, rectal cancer, anal cancer, choriocarcinoma, endometrial cancer, cervical cancer, ovarian cancer, bladder cancer, urothelial carcinoma, kidney cancer, renal cell carcinoma, prostate cancer, testicular tumors, testicular germ cell tumors, ovarian germ cell tumors, Wilms' tumor, malignant melanoma, neuroblastoma, osteosarcoma, Ewing's sarcoma, chondrosarcoma, soft tissue sarcoma, or skin cancer.

[0046] Invention Details

[0047] Exemplary embodiments utilizing the principles of the invention are set forth in the following detailed description of the invention. The features and advantages of the invention can be better understood by referring to the following summary of the invention.

[0048] It should be understood that the scope of protection of each aspect of the present invention is determined by the claims, and the methods and structures within the scope of these claims, as well as their equivalents, are all within the scope of these claims.

[0049] Unless otherwise defined, all technical terms used herein have the same meanings as commonly understood by one of ordinary skill in the art to which the subject matter of the claims pertains. Unless otherwise stated, all patents, patent applications, and publications cited in this document are incorporated herein in their entirety through reference.

[0050] It should be understood that the above summary and the following detailed description are exemplary and explanatory, and not intended to limit any subject matter of the invention. Unless otherwise specified, the singular form includes the plural form. Unless otherwise specified, the use of "or" or "or" means "and / or". Furthermore, the use of the term "comprising" and other forms such as "including," "containing," and "containing" are not limiting.

[0051] Some chemical terms

[0052] The terms “optional,” “optional,” or “optionally” mean that the event or condition described below may or may not occur, including both the occurrence and non-occurrence of the event or condition. For example, “optionally substituted alkyl” means “unsubstituted alkyl” or “substituted alkyl.” Furthermore, the optionally substituted group can be unsubstituted (e.g., -CH2CH3), fully substituted (e.g., -CF2CF3), monosubstituted (e.g., -CH2CH2F), or any level between monosubstituted and fully substituted (e.g., -CH2CHF2, -CF2CH3, -CFHCHF2, etc.). Those skilled in the art will understand that for any group containing one or more substituents, no substitution or substitution mode that is spatially impossible and / or cannot be synthesized is introduced.

[0053] Unless otherwise stated, conventional methods within the scope of the art, such as mass spectrometry, nuclear magnetic resonance, high-performance liquid chromatography, infrared and ultraviolet / visible spectroscopy, and pharmacological methods, are employed. Unless specifically defined herein, the terminology, experimental procedures, and techniques used herein in analytical chemistry, organic synthetic chemistry, and pharmaceutical and medicinal chemistry are known in the art. Standard techniques can be used in chemical synthesis, chemical analysis, drug preparation, formulation and delivery, and patient treatment. For example, reactions and purifications can be carried out using the manufacturer's instructions for use of reagent kits, or in accordance with methods known in the art or the description of this invention. The techniques and methods described herein are generally carried out according to conventional methods well known in the art, based on descriptions in several summary and more specific documents cited and discussed herein. In this specification, groups and their substituents can be selected by those skilled in the art to provide stable structural moieties and compounds.

[0054] When a substituent is described using a conventional chemical formula written from left to right, it also includes chemically equivalent substituents obtained when the structural formula is written from right to left. For example, -CH2O- is equivalent to -OCH2-.

[0055] As used in this article, the terms "group" and "chemical group" refer to a specific part or functional group of a molecule. Chemical groups are often considered as chemical entities that are embedded in or attached to a molecule.

[0056] Some chemical groups named herein may be indicated by abbreviations to represent the total number of carbon atoms. For example, C1-C6 alkyl describes an alkyl group having a total of 1 to 6 carbon atoms, as defined below. The total number of carbon atoms indicated by the abbreviations does not include carbon atoms on possible substituents.

[0057] The terms “halogen,” “halogenated,” or “halide,” used alone or in combination in this article, refer to bromine, chlorine, fluorine, or iodine.

[0058] As used alone or in combination herein, the terms "aromatic," "aromatic ring," "aromatic ring," "aromatic," "aromatic," and "aromatic ring" refer to a planar ring or ring portion of one or more rings having a delocalized electronic conjugated system containing 4n+2 electrons, where n is an integer. An aromatic ring can be formed from 5, 6, 7, 8, 9, or more than 9 atoms. Aromatic compounds can be optionally substituted and can be monocyclic or polycyclic with fused rings. The term aromatic compound includes all carbocyclic rings (such as benzene rings) and rings containing one or more heteroatoms (such as pyridine).

[0059] The term "heteroaromatic ring" as used alone or in combination in this article refers to an aromatic ring containing heteroatoms.

[0060] As used individually or in combination herein, the term "heteroatom" or "hetero" refers to an atom other than carbon and hydrogen. Heteroatoms are independently selected from, but not limited to, oxygen, nitrogen, sulfur, phosphorus, silicon, selenium, and tin. In embodiments where two or more heteroatoms are present, the two or more heteroatoms may be identical to each other, or some or all of the two or more heteroatoms may be different from each other.

[0061] The term “dense” or “dense ring” as used alone or in combination in this article refers to a ring structure in which two or more rings share one or more bonds.

[0062] The term “spiral” or “spiral ring” as used alone or in combination in this article refers to a ring structure in which two or more rings share one or more atoms.

[0063] The term "alkyl" as used alone or in combination herein refers to a monovalent saturated hydrocarbon group, either a straight-chain or a branched chain with optional substitution, having 1-12 carbon atoms, preferably 1-8 carbon atoms, more preferably 1-6 carbon atoms, and connected to other parts of the molecule by single bonds, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, n-octyl, n-nonyl, n-decyl, etc.

[0064] The term "alkylene" as used alone or in combination herein refers to a divalent saturated hydrocarbon group of optional substituted straight or optional branched chain having 1-12 carbon atoms, preferably 1-8 carbon atoms, more preferably 1-6 carbon atoms.

[0065] As used alone or in combination herein, the term "aryl" refers to a fully carbon monocyclic or fused ring having a fully conjugated π-electron system, having 6-14 carbon atoms, preferably 6-12 carbon atoms, and most preferably 6 carbon atoms. The aryl group can be unsubstituted or substituted with one or more substituents, examples of which include, but are not limited to, alkyl, alkyloxy, aryl, aralkyl, amino, halogen, hydroxyl, sulfonyl, sulfinyl, phosphoryl, and heterocyclic groups. Non-limiting examples of unsubstituted aryl groups include, but are not limited to, phenyl, naphthyl, and anthraceneyl.

[0066] As used alone or in combination herein, the term "heteroaryl" refers to a monocyclic or fused ring of 5-12 ring atoms, having 5, 6, 7, 8, 9, 10, 11, or 12 ring atoms, of which 1, 2, 3, or 4 are selected from N, O, and S, and the remaining ring atoms are C, and having a fully conjugated π-electron system. Heteroaryl groups can be unsubstituted or substituted, and the substituents include, but are not limited to, alkyl, alkyloxy, aryl, aralkyl, amino, halogen, hydroxyl, cyano, nitro, carbonyl, and heterocyclic groups. Non-limiting examples of unsubstituted heteroaryl groups include, but are not limited to, pyrrole, furanyl, thiophene, imidazolyl, oxazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrazolyl, and triazineyl.

[0067] The term "cycloalkyl" as used alone or in combination herein refers to a stable, monovalent, non-aromatic monocyclic or polycyclic hydrocarbon group containing only carbon and hydrogen atoms. It may include fused ring, spirocyclic, or bridged ring systems, containing 3-15 cyclic carbon atoms, preferably 3-10 cyclic carbon atoms, more preferably 3-8 cyclic carbon atoms, and may be saturated or unsaturated, linked to other parts of the molecule by single bonds. Non-limiting examples of "cycloalkyl" include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl.

[0068] As used individually or in combination herein, the terms "heterocyclic group," "heterocyclic alkyl group," and "heterocycle" refer to a stable 3-18 member monovalent non-aromatic ring comprising 2-12 carbon atoms and 1-6 heteroatoms selected from nitrogen, oxygen, and sulfur. Unless otherwise stated, the heterocyclic group can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which may contain fused rings, spirocyclic, or bridged ring systems. The nitrogen, carbon, or sulfur on the heterocyclic group may be selectively oxidized, the nitrogen atom may be selectively quaternized, and the heterocyclic group may be partially or completely saturated. A heterocyclic group can be connected to the rest of the molecule via a single bond through a carbon atom or heteroatom on the ring. Heterocyclic groups containing fused rings may contain one or more aromatic or heteroaromatic rings, provided that the atoms connected to the rest of the molecule are atoms from non-aromatic rings. For the purposes of this application, the heterocyclic group is preferably a stable 4-11 monovalent non-aromatic monocyclic or bicyclic ring containing 1-3 heteroatoms selected from nitrogen, oxygen, and sulfur; more preferably, it is a stable 4-8 monovalent non-aromatic monocyclic ring containing 1-3 heteroatoms selected from nitrogen, oxygen, and sulfur. Non-limiting examples of heterocyclic groups include azirheptanyl, azirheptanyl, decahydroisoquinolinyl, dihydrofuranyl, dihydroindolyl, dioxopentyl, 1,1-dioxo-thiomorpholinyl, imidazoalkyl, imidazolinyl, isothiazolyl, isoxazolyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, oxazinyl, piperazinyl, piperidinyl, 4-piperidinoneyl, pyranyl, pyrazolyl, pyrrolidinyl, quinazinyl, quininecycloyl, tetrahydrofuranyl, tetrahydropyranyl, etc.

[0069] The terms "polymorph" or "polymorphism (phenomenon)" as used alone or in combination herein refer to compounds of the present invention having multiple crystal lattice forms. Some compounds of the present invention may have more than one crystal form, and the present invention covers all polymorphs or mixtures thereof.

[0070] Intermediate compounds and polymorphs of the compounds of this invention are also within the scope of this invention.

[0071] Unless otherwise specified, the olefin double bonds contained in the compounds of this invention include E and Z isomers.

[0072] The compounds of this invention include compounds having one or more isotopic substitutions, and references to a specific element include all isotopes of that element within their scope. For example, references to hydrogen include... 1 H, 2 H(D) and 3 H(T). Similarly, references to carbon and oxygen within their scope include, respectively, 12 C 13 C and 14 C and 16 O and 18 O.

[0073] It should be understood that the compounds of the present invention may contain asymmetric centers. These asymmetric centers may independently be R or S configurations. Some compounds of the present invention may also exhibit cis-trans isomerism, which will be apparent to those skilled in the art. It should be understood that the compounds of the present invention include their individual geometric isomers and stereoisomers, as well as mixtures thereof, including racemic mixtures. These isomers can be isolated from mixtures thereof by implementing or modifying known methods, such as chromatography and recrystallization techniques, or they can be prepared separately from suitable isomers of their intermediates.

[0074] The term “pharmaceutically acceptable salt” as used alone or in combination in this article includes both salts with added acid salts and salts with added alkali salts.

[0075] The term "pharmaceutically acceptable salt" as used alone or in combination herein refers to salts that retain the biological potency and properties of the free base of a compound, are not biologically or otherwise undesirable, and are formed with inorganic acids, such as, but not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc., or organic acids, such as, but not limited to, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, decanoic acid, hexanoic acid, carbonic acid, cinnamic acid, citric acid, etc. "Pharmaceutically acceptable base salt" refers to salts that retain the biological potency and properties of the free acid of a compound, and are not biologically or otherwise undesirable. These salts are prepared by reacting a free acid with an inorganic or organic base. Salts formed by reacting with an inorganic base include, but are not limited to, sodium salts, potassium salts, lithium salts, ammonium salts, calcium salts, magnesium salts, iron salts, zinc salts, copper salts, manganese salts, aluminum salts, etc. Preferred inorganic salts are ammonium salts, sodium salts, potassium salts, calcium salts, and manganese salts.

[0076] Organic bases that form salts include, but are not limited to, primary amines, secondary amines, tertiary amines, and cyclic amines, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, ethanolamine, dicyclohexylamine, ethylenediamine, purines, piperazine, piperidine, choline, and caffeine. Particularly preferred organic bases are isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine.

[0077] Crystallization often produces solvates of the compounds of this invention. As used herein, the term "solvate" refers to a combination of one or more molecules of the compounds of this invention and one or more solvent molecules.

[0078] The solvent can be water, in which case the solvate is a hydrate. Alternatively, it can be an organic solvent. Therefore, the compounds of this invention can exist as hydrates, including monohydrates, dihydrates, hemihydrates, trihydrates, tetrahydrates, etc., and the corresponding solvated forms. The compounds of this invention can be true solvates, but in other cases, they may simply retain water or a mixture of water and some other solvents by chance. The compounds of this invention can react in a solvent or precipitate or crystallize in a solvent. The solvates of the compounds of this invention are also included within the scope of this invention.

[0079] The term "pharmaceutical composition" as used alone or in combination herein refers to a formulation containing the compounds of the present invention and a medium generally accepted in the art for delivering biologically active compounds to mammals, such as humans. Such a medium includes all pharmaceutically acceptable carriers.

[0080] As used in this article, the term "acceptable" in relation to formulations, compositions, or ingredients means that it does not have a lasting harmful effect on the overall health of the treated subject.

[0081] The term “pharmaceutically acceptable” as used alone or in combination herein means a substance (such as a carrier or diluent) that does not affect the biological activity or properties of the compounds of the present invention and is relatively non-toxic, i.e., that the substance can be administered to an individual without causing an adverse biological reaction or interacting adversely with any component contained in the composition.

[0082] The term “pharmaceutically acceptable carriers” as used alone or in combination herein includes, but is not limited to, adjuvants, carriers, excipients, auxiliaries, deodorants, diluents, preservatives, dyes / colorants, flavor enhancers, surfactants and wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers that have been approved by the relevant government authorities for use in humans and domesticated animals.

[0083] As used individually or in combination herein, the terms “subject,” “patient,” “object,” or “individual” refer to an individual suffering from a disease, disorder, or symptom, including both mammals and non-mammals. Examples of mammals include, but are not limited to, any member of the class Mammalia: humans; non-human primates (e.g., chimpanzees and other apes and monkeys); livestock such as cattle, horses, sheep, goats, and pigs; domesticated animals such as rabbits, dogs, and cats; and laboratory animals, including rodents such as rats, mice, and guinea pigs. Examples of non-human mammals include, but are not limited to, birds and fish. In one embodiment of the methods and compositions provided herein, the mammal is a human.

[0084] The term "treatment" as used in this article refers to the treatment of relevant diseases or conditions in mammals, particularly humans, including...

[0085] (i) To prevent mammals, especially those previously exposed to a disease or condition but not yet diagnosed with it, from developing the corresponding disease or condition.

[0086] (ii) To suppress a disease or symptom, that is, to control its development;

[0087] (iii) To alleviate the disease or symptom, that is, to make the disease or symptom subside;

[0088] (iv) Relieve symptoms caused by disease or illness.

[0089] The terms “disease” and “symptom” used in this article may be used interchangeably or have different meanings, because some specific diseases or symptoms do not yet have known causative factors (so the cause of the disease is still unclear), so they cannot be recognized as diseases but can only be regarded as unwanted conditions or syndromes. These syndromes have more or less some specific symptoms that have been confirmed by clinical researchers.

[0090] As used herein, the terms "effective amount," "therapeutic effective amount," or "pharmaceutical effective amount" refer to an amount of at least one drug or compound that, when taken, is sufficient to alleviate, to some extent, one or more symptoms of the disease or condition being treated. The result may be a reduction and / or relief of signs, symptoms, or causes, or any other desired change in a biological system. For example, an "effective amount" for treatment is the amount of a composition containing the compounds disclosed herein that is clinically necessary to provide significant symptom relief. Effective amounts suitable for any individual case can be determined using techniques such as dose escalation testing.

[0091] As used herein, the terms “administration,” “application,” “dosage,” etc., refer to methods that deliver a compound or composition to the desired site for biological action. These methods include, but are not limited to, oral administration, duodenal administration, parenteral administration (including intravenous, subcutaneous, intraperitoneal, intramuscular, intra-arterial injection or infusion), local administration, and rectal administration. In a preferred embodiment, the compounds and compositions discussed herein are administered orally.

[0092] Preparation of the compounds of the present invention

[0093] The specific embodiments described below are intended to enable those skilled in the art to better understand and implement the present invention. They should not be considered as limiting the scope of the invention, but merely as exemplary illustrations and typical representatives. Those skilled in the art should understand that there are other synthetic routes for forming the compounds of the present invention; the examples provided below are non-limiting.

[0094] All operations involving easily oxidized or hydrolyzed raw materials are performed under nitrogen protection. Unless otherwise stated, the raw materials used in this invention are commercially available and used directly without further purification.

[0095] Column chromatography used silica gel (200-300 mesh) manufactured by Qingdao Chemical Co., Ltd. Thin-layer chromatography used pre-prepared plates (60PF silica gel) manufactured by E. Merck. 254 Chiral compound separation and enantiomeric excess (ee) determination were performed using an Agilent LC 1200 series (column: CHIRALPAK AD-H, 0.25 mm). Millimeters, 5 micrometers, 30°C). Chiral compound separation and preparation were performed using an IC-5 chiral preparative column (mm, 5 μm, 30°C). The flow rate was 4.7 mL / min at 25 °C (mm, 5 μm). Nuclear magnetic resonance (NMR) chromatography was performed using a Varian VNMRS-400 NMR spectrometer; liquid chromatography-mass spectrometry (LC / MS) was performed using a FINNIGAN Thermo LCQ Advantage MAX, Agilent LC 1200 series (column: Waters Symmetry C18). (millimeter, 5 micrometer, 35℃), using ESI(+) ion mode.

[0096] Experimental Section

[0097] Intermediate 1: (R)-N 1 -(2-Methoxyethyl)-N 1 5-Dimethylhexane-1,4-diamine hydrochloride

[0098] The target compound was synthesized according to the method of intermediate 231 in patent WO2022262796.

[0099] Intermediate 2: (R)-2-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-2-azaspiro[3.5]nonane-7-one

[0100] Step 1: (R)-4-(8,11-dioxa-2-azaspiro[3.2.4] 7 .2 4 (tetran-2-yl)-N-(2-methoxyethyl)-N,5-dimethylhexane-1-amine

[0101] (1,4-dioxane[4.5]decane-8,8-diyl)diethanol (2.02 g) and diisopropylethylamine (5.16 g) were added to dry acetonitrile (50 mL), cooled to -30 °C, and trifluoromethanesulfonic anhydride (6.21 g) was added dropwise. The mixture was stirred at -30 °C for 1 hour. Intermediate 1 (3.30 g) and diisopropylethylamine (5.16 g) were added, and the mixture was heated under reflux for 24 hours. The mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, and extracted with ethyl acetate (50 mL * 3) after adding saturated sodium bicarbonate aqueous solution (100 mL). The extract was washed with water and saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (dichloromethane / methanol, 15:1, with 2% concentrated ammonia) to give the target compound (2.40 g). MS m / z [LC-MS]: 369.31 [M+1].

[0102] Step 2: (R)-2-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-2-azaspiro[3.5]nonane-7-one

[0103] The compound obtained in step 1 (1.85 g) and p-toluenesulfonic acid (2.15 g) were added to an acetone-water mixture (3:1, 20 mL), and the mixture was heated under reflux for 16 hours. After cooling to room temperature, the pH was adjusted to 12 with 1 M sodium hydroxide solution. The mixture was extracted with ethyl acetate (30 mL * 3), washed with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the target compound. MS m / z [LC-MS]: 325.29 [M+1].

[0104] Intermediate 3: (R)-N-(2-methoxyethyl)-N,5-dimethyl-4-(4'-methyl-5',7',8',9'-tetrahydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-1-yl)hexyl-1-amine

[0105] Under nitrogen protection, 2.34 g of 4-iodo-5-methylpyridin-3-amine, 3.24 g of intermediate 2, 225 mg of palladium acetate, and 3.38 g of 1,4-diazabicyclo[2.2.2]octane were added to 25 mL of N,N-dimethylformamide and heated to 120 °C for 12 hours. The reaction mixture was cooled to room temperature, poured into 200 mL of water, and extracted three times with 50 mL of ethyl acetate. The extracts were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (dichloromethane / methanol, 20:1, with 1% concentrated ammonia) to give the target compound (2.88 g). MS m / z [LC-MS]: 413.33 [M+1].

[0106] Intermediate 4: (R)-3,5-difluoro-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzoic acid

[0107] Under nitrogen protection, intermediate 1 (4.13 g), 3,5-difluoro-2-iodobenzoic acid (3.41 g), copper powder (635 mg), and cesium carbonate (19.0 g) were added to isopropyl acetate (50 mL), and the mixture was heated to 90 °C and stirred for 12 hours. The reaction solution was cooled to room temperature, filtered, and 25 mL of 4M ethyl hydrochloride solution was added to the filtrate. After filtration, the filter cake was added to ethyl acetate (200 mL), and 40 mL of concentrated ammonia was added while stirring. Then, sodium chloride (3.41 g) was added, and stirring was continued for 20 minutes. The organic phase was separated, and the aqueous phase was extracted with ethyl acetate (50 mL x 3). The extract and organic phase were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the target compound (4.56 g). MS m / z [LC-MS]: 569.33 [M+1].

[0108] Intermediates 5 to 7 in the table below are synthesized using the same method as intermediate 4.

[0109] Example 1: (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide diphenylsulfonate

[0110] Example 2: (S,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide diphenylsulfonate

[0111] Step 1: N-Ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide

[0112] Intermediate 4 (568 mg), N-ethylpropyl-2-amine (174 mg), diisopropylethylamine (387 mg), and 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (456 mg) were added to acetonitrile (10 mL) and heated to 60 °C for 4 hours. The solvent was removed by concentration under reduced pressure, and the residue was dissolved in dichloromethane (20 mL). The mixture was washed with water, concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (dichloromethane / methanol, 15:1, containing 1% concentrated ammonia) to give the target compound (580 mg). MS m / z [LC-MS]: 638.42 [M+1].

[0113] Step 2: (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide and (S,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide

[0114] The product obtained in step 1 was separated using an IC-5 chiral preparative column (5 μm, 10 x 250 mm) with hexane / ethanol / diethylamine 75:25:0.1 as the elution solvent, at a flow rate of 4.7 mL / min and 25 °C, yielding (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide (195 mg). R = 31.1 minutes. MS m / z [LC-MS]: 638.42 [M+1]. Simultaneously, (S,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide (190 mg) was obtained. R = 18.6 minutes. MS m / z [LC-MS]: 638.42 [M+1].

[0115] Step 3: (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide dibenzamide sulfonate and (S,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide dibenzamide sulfonate

[0116] The (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide (195 mg) obtained in step 2 was dissolved in methanol (5 mL), and benzenesulfonic acid (97 mg) was added. The mixture was stirred at room temperature for 30 minutes. The mixture was concentrated under pressure, and isopropyl ether (10 mL) was added to the residue. The mixture was sonicated for 30 minutes and filtered to obtain (R,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide diphenylsulfonate (265 mg). MS m / z [LC-MS]: 638.42 [M+1]. 1 H NMR (400MHz, CD3OD), δ = 8.49 (s, 0.7H), 8.47 (s, 0.3H), 7.97 (s, 1H), 7.79-7.82 (m, 4H), 7.37-7.50 (m, 7 H),7.27-7.35(m,1H),4.10-4.38(m,4H),3.76-3.90(m,1H),3.40-3.74(m,5H),3.04-3.39(m,9H),2.82 -2.99(m,4H),2.68(s,3H),2.40-2.62(m,1H),2.07-2.32(m,3H),1.80-1.99(m,2H),1.56-1.76(m,2H) ,1.10-1.18(m,3H),1.07(d,J=6.8Hz,3H),0.99(d,J=6.8Hz,3H),0.81-0.93(m,3H),0.48-0.70(m,3H).

[0117] The same method yielded (S,R)-N-ethyl-3,5-difluoro-N-isopropyl-2-(1-(6-((2-methoxyethyl)(methyl)amino)-2-methylhexane-3-yl)-4'-methyl-7',8'-dihydrospiro[azacyclobutane-3,6'-pyrido[3,4-b]indole]-9'(5'H)-yl)benzamide diphenylsulfonate (255 mg). MS m / z [LC-MS]: 638.42 [M+1]. 1 H NMR (400MHz, CD3OD), δ = 8.78 (s, 0.5H), 8.66 (s, 0.2H), 8.56 (s, 0.3H), 7.96-8.08 (m, 1H), 7.79-7.82 (m, 4H ),7.47-7.54(m,1H),7.38-7.45(m,6H),7.32-7.36(m,1H),4.06-4.45(m,4H),3.62-3.94(m,4H),3.34-3.6 0(m,6H),3.08-3.30(m,5H),2.83-3.02(m,4H),2.73(s,3H),2.52-2.70(m,1H),2.08-2.45(m,3H),1.81-2 .02(m,2H),1.54-1.77(m,2H),1.04-1.21(m,6H),0.96-1.03(m,3H),0.82-0.94(m,2H),0.48-0.73(m,4H).

[0118] Examples 3 to 12 in the table below were synthesized using the methods described in Examples 1 and 2.

[0119] Biological testing

[0120] Biochemical detection of the compound's inhibition of wild-type Menin binding to MLL

[0121] Wild-type Menin binds with high affinity to the conserved N-terminal sequence of MLL. Compounds bind to Menin by competing with the N-terminus of MLL. FITC-MLL(4-43) (synthesized by GenScript) contains the conserved sequence for MLL-Menin binding. We constructed a method for detecting the binding activity of Menin to MLL using a fluorescence polarization binding assay (FP Assay) and applied it to detect the inhibitory binding activity of compounds. The specific method is as follows:

[0122] The compound was serially diluted 5-fold with 100% DMSO, starting from 1 mM (totaling 8 concentrations). 2 μl of each concentration was added to 48 μl of reaction buffer (50 mM NaCl, 50 mM Tris (pH 7.5), 0.05% Tween-20, 1 mM DTT) and mixed well to prepare 4* compounds (final concentrations of 10000, 2000, 400, 80, 16, 3.2, 0.64, 0 nM). 2* Menin was prepared to a final concentration of 5 nM, and 4* FITC-MLL (4-43) was prepared to a final concentration of 2 nM using reaction buffer. 5 μl of the 4* compound was added to a 384-well plate (OptiPlate-384, purchased from PerkinElmer), followed by 10 μl of 2* Menin. After centrifugation, 5 μl of FITC-MLL (4-43) was added, and the reaction was started by centrifugation. The reaction was incubated at 23°C in the dark for 1 hour. After the reaction, signal values ​​(excitation wavelength 480 nm / emission wavelength 535 nm) were read on an EnVision 2104 Multilabel Reader (purchased from PerkinElmer). The data was processed using GraphPad Prism software to obtain the IC50 values ​​of the compounds. 50 value

[0123] Biochemical detection of the compound's inhibition of binding between mutant Menin and MLL

[0124] Will Menin mut The DNA sequence was constructed into the pET28a vector via enzyme digestion and ligation. The recombinant protein was expressed using the BL21 E. coli system, purified with Ni-NTA, and yielded Menin. mut Menin can bind with high affinity to the conserved N-terminal sequence of MLL. The compound binds to Menin by competing with the N-terminus of MLL. (FITC-MLL) -4-43 (Synthesized by GenScript) Contains a conserved sequence for MLL-Menin binding. We used FITC-MLL through a fluorescence polarization binding assay (FP assay). -4-43 with Menin mut The principle that the molecular weight increases and the fluorescence intensifies after binding is used to detect the compound's effect on Menin. mut Inhibitory effect by binding to MLL. Specific methods are as follows:

[0125] The compound was dissolved in DMSO to a final concentration of 10 mM. The 10 mM compound was then diluted 10-fold with DMSO, followed by 5-fold serial dilutions to obtain eight concentrations. 2 μl of each concentration was added to 48 μl of reaction buffer (50 mM NaCl, 50 mM Tris, pH 7.5, 0.05% Tween-20, 1 mM DTT) and mixed thoroughly to prepare 4x compounds (final concentrations of 10000, 2000, 400, 80, 16, 3.2, 0.64, 0 nM) for later use. 2x Menin was prepared using the reaction buffer. mut The final concentration was 5 nM, 4x FITC-MLL -4-43 The final concentration was 2 nM. 5 μl of the 4x compound was added to a 384-well plate (OptiPlate-384, purchased from PerkinElmer), followed by 10 μl of 2x Menin. After centrifugation, 5 μl of FITC-MLL was added. -4-43 The reaction was initiated by brief centrifugation and incubated at 23°C in the dark for 1 hour. After the reaction, signal values ​​(excitation wavelength 480 nm / emission wavelength 535 nm) were read on an EnVision 2104 Multilabel Reader (purchased from PerkinElmer). The inhibitory effect of each compound was determined at 8 concentrations. Data were processed using GraphPad Prism software to calculate the half-maximal inhibitory concentration (IC50) of each compound for the binding of Menin to MLL. Note: x indicates multiplication, representing a factor.

[0126] Table 1. Inhibitory activity of the compounds in the examples against wild-type and mutant Menin-MLL protein binding.

[0127] Table 1 lists the inhibitory activity (IC50) of the compounds in the embodiments of the present invention against the binding of wild-type and mutant Menin-MLL proteins. 50 Values. Data indicate that the compounds provided by this invention possess excellent binding inhibitory activity against both wild-type and mutant Menin-MLL proteins.

[0128] Assay of the inhibitory activity of the compound on the proliferation of MV-4-11 cells expressing wild-type Menin protein:

[0129] MV-4-11 cells are human acute myeloid leukemia cells containing the MLL-AF4 fusion protein. In this experiment, different concentrations of compounds were incubated with MV-4-11 cells, and the cells were analyzed using Promega's [technology / technology / etc.]. The detection reagents established a screening method for MV-4-11 cell proliferation inhibition.

[0130] MV-4-11 cells were cultured in 1640 medium (Biological Industries, catalog number 01-100-1ACS) containing 10% fetal bovine serum (Biological Industries, catalog number 04-001-1ACS) at 37°C and 5% CO2. Cells were passaged 2-3 times per week. Cells were seeded at 5000 cells / well in 96-well cell culture plates. 195 μL / well was cultured at 37°C in 5% CO2. After 24 hours, the compound was serially diluted 3-fold with 100% DMSO, starting at 10 mM (total of 10 concentrations). 4 μL of each concentration was then added to 96 μL of RPMI-1640 medium for further dilution. 5 μL of each diluted compound was added to the prepared cell suspension, and the compound and cells were incubated together in a cell culture incubator for 72 h (3 days). Then, 35 μL of CellTiter-Blue (Promega, catalog number G8082) reagent was added, and the reaction was carried out at 37°C in 5% CO2 for 4 h. Chemiluminescence values ​​were read using a BMG Clariostar Microplate Reader, and the data were processed using GraphPad Prism software to calculate the IC50 inhibitory effect of the compound on cell proliferation. 50 Value. Note: * indicates multiplication, representing a multiple.

[0131] Assay of the inhibitory activity of the compound on the proliferation of MOLM13 cells expressing mutant Menin protein:

[0132] Will Menin M327I The DNA sequence was constructed into the lentiviral vector pCDHL-CMV-3xFlag-puroR via enzyme digestion and ligation. This vector, along with psPAX2 / pMD2G, was co-transfected into 293T cells. After 48 hours, the virus was harvested and used to infect MOLM13 cells. Pooling and single-clone selection were performed using puromycin, ultimately yielding cells that stably express Menin. M327I Stable cell lines for recombinant proteins.

[0133] MOLM13 cells are human acute myeloid leukemia cells containing the MLL-AF9 fusion protein. This experiment will use different concentrations of the compound with Menin... M327I / MOLM13 cells were incubated for 7 days, and viability was detected using Promega's Cell-Titer Glo.

[0134] MOLM13 cells were cultured in RPMI-1640 medium containing 20% ​​fetal bovine serum. The day before the addition of the compound, 500 cells / well were seeded in 96-well cell culture plates, with 195 μL of medium per well. The compound was dissolved in DMSO to a final concentration of 10 mM. The 10 mM compound was serially diluted 3-fold with DMSO to obtain 10 concentrations. 2 μL of each concentration was added to 48 μL of RPMI-1640 medium for a 25-fold dilution, creating the 40x compound, which was then added to the prepared cell suspension. After incubation for 7 days, 35 μL of Cell-Titer Glo was added, and the cells were incubated at room temperature for 10 minutes. Fluorescence signals were read on a CLARIO starPlus imager. Finally, the IC50 of the compound's inhibitory effect on cell proliferation was calculated using GraphPad Prism 5.0. 50 Value. Note: * indicates multiplication, representing a multiple.

[0135] Table 2. Inhibitory activity of the compounds in the examples against the proliferation of MV-4-11 cells expressing wild-type Menin protein and MOLM13 cells expressing mutant Menin protein.

[0136] Table 2 lists the IC50 inhibitory activity of the compounds from the embodiments of the present invention against the proliferation of MV-4-11 cells expressing wild-type Menin protein and MOLM13 cells expressing mutant Menin protein. 50 Values. Data show that the compounds provided by this invention have good inhibitory activity against the proliferation of cells expressing both wild-type and mutant Menin proteins.

[0137] Determination of pharmacokinetic data of the compound in SD rats

[0138] Male SD rats were obtained from Beijing Vital River Laboratory Animal Technology Co., Ltd. Rats were divided into groups of three and orally administered a suspension of the test sample (10 mg / kg, suspension of 0.5% MC + 0.1% SDS) via gavage. Animals were fasted overnight before the experiment, from 10 hours before administration to 4 hours after administration. Blood samples were collected at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours after administration. After anesthesia with isoflurane using a small animal anesthesia machine, 0.3 mL of whole blood was collected through the fundus venous plexus and placed in heparin anticoagulant tubes. The samples were centrifuged at 4000 rpm for 5 minutes at 4°C. Plasma was transferred to centrifuge tubes and stored at -80°C until analysis. Protein precipitation was used to extract the plasma samples, and the extract was analyzed by LC / MS. Results of some compound detections are shown in Table 3.

[0139] Table 3 Pharmacokinetic parameters of the compounds in the examples

[0140] Table 3 lists the pharmacokinetic data of the compounds of the present invention in SD rats. This indicates that the compounds provided by the present invention have relatively good in vivo metabolic levels.

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug thereof, in, Ring A is a 5-8 member nitrogen-containing heterocycle or a nitrogen-containing heteroaromatic ring, and each X is independently C or N. R 1a -C(=O)-NR 2a R 2b , Alternatively, a 5-6 membered heteroaryl group, said heteroaryl group comprising 1-3 nitrogen atoms and optionally comprising a carbonyl group, and said heteroaryl group optionally being surrounded by a 3-6 membered cycloalkyl group or a C-membered ring. 1-4 Alkyl substitution, R 2a and R 2b Each independently is either H or C. 1-4 Alkyl group, wherein the alkyl group may optionally be substituted with -OH or -NH2. R 1b For F or Cl, R2 is selected from H, halogens, and C. 1-4 Alkyl, -OC 1-4 Alkyl and -NR 3a R 3b , R 3a and R 3b Each independently is either H or C. 1-4 alkyl, X1 and X2 are each independently either N or CH. L1 is -CH2-, -CH2-CH2-, or -CH2-CH2-CH2-. L2 and L3 are each independently -CH2- or -CH2-CH2-. R4 is C 1-6 Alkyl, 3-8 membered cycloalkyl, or 3-8 membered heterocycloalkyl R3 is -C 1-6 Alkylene-NR 4a R 4b -C 1-6 Alkylene-C(O)-NR 5a R 5b -C 1-6 alkylene -OH or -C 1-6 Alkylene-NR7-C(O)-OC 1-4 Alkylene-OC(O)-C 1-4 Alkyl group, wherein the alkyl or alkylene group may optionally be converted to a halogen, -CN, -OH or -OC. l-4 Alkyl substitution, R 4a and R 4b Each is independently selected from H and C. 1-6 Alkyl, -C(O)-C 1-4 Alkyl, -C(O)-OC 1-4 Alkyl groups and -C(O)-NR 8a R 8b The C 1-6 Alkyl groups may optionally be replaced by halogens, -CN, -OH, or -S(O)2-C. 1-4 Alkyl, -OC 1-4 Alkyl, -C(O)-NR 6a R 6b Or -NR 6c -C(O)-C 1-4 Alkyl substitution, R 5a R 5b、 R 6a R 6b、 R 6c R7, R 8a and R 8b Each is independently selected from H and C. 1-6 alkyl.

2. The compound according to claim 1, or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug thereof, wherein... for or R 1a for R 1b R4 is F, R2 is isopropyl, R3 is H, and R4 is F.

3. A compound of formula (II) or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug thereof, in, R 2a and R 2b Each can be independently methyl, ethyl, or isopropyl. X3 is either N or CR 10 , R 10 It can be F, Cl, or methyl.

4. The compound according to claim 3, having the following structure. Or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite, or prodrug.

5. A pharmaceutical composition comprising a compound according to any one of claims 1-4 or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite or prodrug thereof, and optionally comprising a pharmaceutically acceptable carrier.

6. Use of the compound of any one of claims 1-4 or a pharmaceutically acceptable salt, solvate, polymorph, tautomer, metabolite or prodrug thereof, or the pharmaceutical composition of claim 5, in the preparation of a medicament for treating diseases related to MLL activity.

7. The use according to claim 6, wherein the disease associated with MLL activity is cancer.

8. The use according to claim 6, wherein the disease associated with MLL activity is acute leukemia, chronic lymphocytic leukemia, chronic myeloid leukemia, myelodysplastic syndrome, polycythemia vera, malignant lymphoma, myeloma, brain tumor, head and neck cancer, esophageal cancer, thyroid cancer, small cell lung cancer, non-small cell lung cancer, breast cancer, gastric cancer, gallbladder and bile duct cancer, liver cancer, hepatocellular carcinoma, pancreatic cancer, colon cancer, rectal cancer, anal cancer, choriocarcinoma, endometrial cancer, cervical cancer, ovarian cancer, bladder cancer, urothelial carcinoma, kidney cancer, renal cell carcinoma, prostate cancer, testicular tumor, testicular germ cell tumor, ovarian germ cell tumor, Wilms' tumor, malignant melanoma, neuroblastoma, osteosarcoma, Ewing's sarcoma, chondrosarcoma, soft tissue sarcoma, or skin cancer.

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