Heterocyclic compounds as BCL-2 inhibitors

Abstract

This invention relates to compounds, pharmaceutical compositions containing the same, methods of preparing the same, and their use as BCL-2 inhibitors. The compounds are those represented by formula (I), or isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts thereof. The invention also relates to the use of the compounds for the treatment or prevention of BCL-2-mediated diseases such as tumors, and methods of using the compounds to treat said diseases.

Description

Technical Field

[0001] This invention relates to compounds, pharmaceutical compositions containing the same, and their use as inhibitors of B-cell lymphoma-2 (BCL-2). More specifically, the invention provides novel compounds as BCL-2 inhibitors, pharmaceutical compositions containing such compounds, and the use of said compounds for the treatment or prevention of BCL-2-mediated diseases and functional impairments, such as tumors. The invention also relates to methods for preparing said compounds. Background Technology

[0002] The BCL-2 protein family is one of the core regulatory mechanisms of apoptosis (also known as programmed cell death). It receives and transmits intrinsic intracellular signals or external environmental stress signals, such as nutrient or hypoxic stress, DNA damage, overactivation of oncogenes, and endoplasmic reticulum stress. It plays a dominant role in the intrinsic apoptotic pathway. The BCL-2 (B-cell lymphoma-2) protein was first discovered in 1986 and is expressed by the BCL-2 gene. The BCL-2 gene is a proto-oncogene, and the proteins it expresses are called BCL-2 family proteins. There are 27 BCL-2 family proteins in the human body, which can be divided into three subclasses based on function and sequence analysis. The first subclass antagonizes apoptosis and includes BCL-XL, BCL-2, BCL-W, MCL-1, and BFL-1. These are mainly located on mitochondria, protecting them from environmental damage. The other two subclasses promote apoptosis; one subclass is the final executor of mitochondrial damage, including BAX and BAK. The rest belong to the BH3 subclass and can directly sense various cellular stress signals. The dynamic balance of interactions between antagonistic and apoptosis-promoting proteins determines the life and death of cells. The apoptosis-antagonistic BCL-2 protein is closely related to tumors; approximately 50% of tumors (such as leukemia, rectal cancer, and prostate cancer) show abnormal overexpression of BCL-2 family proteins, with abnormal BCL-2 activity being widespread in hematological malignancies. Multiple signaling pathways, including JAK-STAT, NFkB, and UPP (ubiquitin-proteasome), can induce overexpression of apoptosis-antagonistic BCL-2 proteins.

[0003] High expression of BCL-2 family anti-apoptotic proteins is associated with drug resistance in various tumors. For example, overexpression of BCL-2 anti-apoptotic proteins can enable tumor cells to evade apoptosis induced by antitumor drugs, thereby leading to drug resistance. Studies have shown that inhibiting BCL-2 family proteins can inhibit tumor angiogenesis, thereby inhibiting tumor metastasis (Benjamin, D.; Isaac, J. et al. J. Clin. Oncol. 2008, 26(25), 4180). Therefore, targeted inhibition of BCL-2 family anti-apoptotic proteins can inhibit tumor occurrence, development, and drug resistance.

[0004] Although more than 20 small molecule inhibitors targeting the BCL-2 family have been reported, very few have entered clinical trials. Teva's Obatoclax only achieved a partial response in 1 of 26 patients with chronic lymphocytic leukemia (CLL) and had strong neurotoxicity, leading to the termination of its development in 2013. AbbVie's Navitoclax (ABT-263) showed a good response rate of 50% in a phase I dose escalation trial in patients with relapsed or refractory lymphomas, but it also exhibited very strong BCL-XL targeting toxicity, such as thrombocytopenia and severe anemia. Venetoclax (ABT-199), jointly developed by AbbVie and Roche, is a highly selective BCL-2 inhibitor (Andrew, J.; Joel, D. et al. Nature Medicine, 2013, 19(2), 202). In the treatment of relapsed / refractory chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MCL), and multiple myeloma (MM), its objective response rate (ORR) and complete response rate (CR) have been greatly improved when used in combination with ibrutinib (Valentin, R.; Grablow, S. et al. Blood, 2018, 132(12), 1248). However, it still has toxic side effects such as leukopenia and thrombocytopenia, anemia, diarrhea, dizziness, fatigue, and increased susceptibility to infection. Severe side effects include pneumonia, anemia, and high fever. Therefore, it is necessary to develop highly active, low-toxicity selective BCL-2 inhibitors. Summary of the Invention

[0005] This invention provides a compound of formula (I) that can be used as a selective inhibitor of BCL-2, including its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts.

[0006]

[0007] in:

[0008] X 1The substituent is selected from a 6-membered saturated heterocyclic group containing one or two heteroatoms selected from N, O, and S, wherein the optional substituent is selected from a 4-membered saturated heterocyclic group containing one or two heteroatoms selected from N, O, and S; preferably, X 1 Selected from optional substituted 6-membered saturated heterocyclic groups containing one or two heteroatoms selected from N and O, wherein the optional substituents are selected from oxetane; more preferably, X 1 Selected from 1,4-dioxane, tetrahydropyranyl, N-oxetane-butylpiperidinyl, and N-oxetane-butylmorpholinyl; most preferably, X 1 Selected from (S)-1,4-dioxane-2-yl, (R)-1,4-dioxane-2-yl, tetrahydropyran-4-yl, 1-(oxetane-3-yl)piperidin-4-yl, and (S)-4-(oxetane-3-yl)morpholin-2-yl;

[0009] X 2 Selected from 5-6 membered heterocyclic groups containing one or two nitrogen atoms; wherein the ring may optionally be substituted by one or two C1-C4 alkyl groups; preferably, X 2 Selected from 6-membered heterocyclic groups containing one or two nitrogen atoms; wherein the ring may optionally be substituted by one or two C1-C4 alkyl groups; more preferably, X 2 Selected from The ring may optionally be replaced by a C1-C4 alkyl group; most preferably, X 2 Selected from

[0010] R 0 Selected from hydrogen and halogens; preferably, R 0 Selected from hydrogen, fluorine, and chlorine; most preferably, R 0 Selected from hydrogen and fluorine;

[0011] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen and C1-C6 alkyl groups, wherein R 1 and R 2 Or R 3 and R 4 It can form a 3- to 6-membered cycloalkyl group together with the attached carbon atom; preferably, R 1 R 2 R 3 R 4 Each is independently selected from hydrogen and C1-C4 alkyl groups, wherein R 1 and R 2 Or R 3 and R 4It can form a 3- to 4-membered cycloalkyl group together with the attached carbon atom; more preferably, R 1 R 2 R 3 R 4 Each is independently selected from hydrogen or methyl and R 1 R 2 R 3 R 4 It's not entirely hydrogen, of which R 1 and R 2 Or R 3 and R 4 It can form a cyclopropyl group together with the attached carbon atom; most preferably, R 1 R 2 R 3 R 4 Each is independently selected from hydrogen or methyl and R 1 R 2 R 3 R 4 It's not entirely hydrogen, of which R 3 and R 4 It can form a cyclopropyl group together with the carbon atom it is attached to.

[0012] Preferably, the present invention relates to the compound of formula (I) above, its isomers, prodrugs, solvates, stable isotopic derivatives or pharmaceutically acceptable salts, wherein:

[0013] X 1 Selected from optional substituted 6-membered saturated heterocyclic groups containing one or two heteroatoms selected from N or O, wherein the optional substituents are selected from oxobutyric groups;

[0014] X 2 It is selected from a 6-membered heterocyclic group containing one or two nitrogen atoms; wherein the ring may optionally be substituted by one or two C1 to C4 alkyl groups;

[0015] R 0 Selected from hydrogen and halogens;

[0016] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen and C1-C4 alkyl groups, wherein R 1 and R 2 Or R 3 and R 4 It can form 3- to 4-membered cycloalkyl groups together with the carbon atoms it is attached to.

[0017] More preferably, the present invention relates to the aforementioned compound of formula (I), its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts, wherein:

[0018] X 1 Selected from optional substituted 6-membered saturated heterocyclic groups containing one or two heteroatoms selected from N or O, wherein the optional substituents are selected from oxobutyric groups;

[0019] X 2 for In this embodiment, the ring may be optionally replaced by a C1 to C4 alkyl group;

[0020] R 0 Selected from hydrogen, fluorine, and chlorine;

[0021] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen and C1-C4 alkyl groups, wherein R 1 and R 2 Or R 3 and R 4 It can form 3- to 4-membered cycloalkyl groups together with the carbon atoms it is attached to.

[0022] More preferably, the present invention relates to the aforementioned compound of formula (I), its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts, wherein:

[0023] X 1 Selected from 1,4-dioxane, tetrahydropyranyl, N-oxetanebutylpiperidinyl, and N-oxetanebutylmorpholinyl;

[0024] X 2 for In this embodiment, the ring may be optionally replaced by a C1 to C4 alkyl group;

[0025] R 0 Selected from hydrogen, fluorine, and chlorine;

[0026] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen or methyl and R 1 R 2 R 3 R 4 It's not entirely hydrogen, of which R 1 and R 2 Or R 3 and R 4 It can form a cyclopropyl group together with the carbon atom it is attached to.

[0027] More preferably, the present invention relates to the aforementioned compound of formula (I), its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts, wherein:

[0028] X 1 Selected from 1,4-dioxane, tetrahydropyranyl, N-oxetanebutylpiperidinyl, and N-oxetanebutylmorpholinyl;

[0029] X 2 Selected from

[0030] R 0 Selected from hydrogen and fluorine;

[0031] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen or methyl and R 1 R 2 R 3 R 4 It's not entirely hydrogen, of which R 1 and R 2 Or R 3 and R 4 It can form a cyclopropyl group together with the carbon atom it is attached to.

[0032] Furthermore, preferably, the present invention relates to the aforementioned compound of formula (I), its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts, wherein:

[0033] X 1 Selected from (S)-1,4-dioxane-2-yl, (R)-1,4-dioxane-2-yl, tetrahydropyran-4-yl, 1-(oxetane-3-yl)piperidin-4-yl, and (S)-4-(oxetane-3-yl)morpholin-2-yl;

[0034] X 2 Selected from

[0035] R 0 Selected from hydrogen and fluorine;

[0036] R 1 R 2 R 3 R 4 Each is independently selected from hydrogen or methyl and R 1 R 2 R 3 R 4 It's not entirely hydrogen, of which R 3 and R 4It can form a cyclopropyl group together with the carbon atom it is attached to.

[0037] Most preferably, the present invention relates to the aforementioned compound of formula (I), its isomers, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts selected from:

[0038]

[0039]

[0040]

[0041]

[0042]

[0043] The present invention also relates to the use of compounds of formula (I), isomers thereof, prodrugs, solvates, stable isotopic derivatives thereof or pharmaceutically acceptable salts thereof as described in any embodiment of the present invention in the preparation of medicaments used as BCL-2 inhibitors.

[0044] The present invention also relates to the use of compounds of formula (I) or isomers thereof, prodrugs, solvates, stable isotopic derivatives thereof or pharmaceutically acceptable salts thereof, according to any embodiment of the present invention, in the preparation of medicaments for the treatment or prevention of BCL-2-mediated diseases, such as tumors, selected from hematologic malignancies, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0045] The present invention further relates to a pharmaceutical composition comprising a compound of formula (I) or an isomer thereof as described in any embodiment of the present invention, a prodrug, a solvate, a stable isotope derivative thereof or a pharmaceutically acceptable salt thereof, optionally one or more other BCL-2 inhibitors, and one or more pharmaceutically acceptable carriers, diluents and excipients.

[0046] The present invention also relates to the use of the pharmaceutical composition according to the invention in the preparation of a medicament for treating or preventing diseases mediated by BCL-2, such as tumors, the tumors being selected from malignant hematologic diseases, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0047] The present invention also relates to a method for treating or preventing BCL-2-mediated diseases, comprising administering to a patient in need a therapeutically effective amount of a compound or isomer thereof, prodrug, solvate, stable isotope derivative thereof, or pharmaceutically acceptable salt thereof, as described in any embodiment of the present invention; or a pharmaceutical composition thereof, wherein the related disease is, for example, a tumor, selected from hematologic malignancies, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0048] Another aspect of the present invention relates to compounds, or isomers thereof, prodrugs, solvates, stable isotope derivatives or pharmaceutically acceptable salts, as described in any embodiment of the present invention, for the treatment or prevention of BCL-2-mediated diseases, such as tumors selected from hematologic malignancies, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0049] Another aspect of the present invention relates to a pharmaceutical composition comprising a compound of formula (I) or an isomer thereof as described in any embodiment of the present invention, a prodrug, a solvate, a stable isotopic derivative thereof or a pharmaceutically acceptable salt thereof, optionally one or more other BCL-2 inhibitors, and one or more pharmaceutically acceptable carriers, diluents and excipients, for the treatment or prevention of BCL-2-mediated diseases, such as tumors selected from hematologic malignancies, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0050] Another aspect of the invention relates to compounds of formula (I) or isomers thereof, prodrugs, solvates, stable isotopic derivatives thereof, or pharmaceutically acceptable salts thereof, as a treatment and / or prevention of BCL-2-mediated related diseases. The BCL-2-mediated related diseases include, for example, tumors selected from hematologic malignancies, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0051] According to the present invention, the drug can be any dosage form, including but not limited to tablets, capsules, solutions, lyophilized preparations, and injections.

[0052] The pharmaceutical formulations of the present invention can be administered in dose units containing a predetermined amount of the active ingredient per dose unit. Such units may contain, for example, 0.5 mg to 1 gram, preferably 1 mg to 700 mg, and particularly preferably 5 mg to 300 mg of the compound of the present invention, depending on the condition being treated, the method of administration, and the patient's age, weight, and condition. Alternatively, the pharmaceutical formulation can be administered in dose units containing a predetermined amount of the active ingredient per dose unit. Preferred dose unit formulations are those containing the daily dose or fractional dose, or a corresponding fraction thereof, as indicated above. Furthermore, this type of pharmaceutical formulation can be prepared using methods known in the pharmaceutical industry.

[0053] The pharmaceutical formulations of the present invention are suitable for administration by any desired and suitable method, such as oral (including oral cavity or sublingual), rectal, nasal, local (including oral cavity, sublingual, or percutaneous), vaginal, or parenteral (including subcutaneous, intramuscular, intravenous, or intradermal) administration. Such formulations can be prepared using all methods known in the pharmaceutical field, for example, by combining the active ingredient with one or more excipients or one or more adjuvants.

[0054] Preparation process

[0055] The present invention also provides a method for preparing the compound.

[0056] Process 1

[0057]

[0058] R 0 With X 1 The definition is as stated above;

[0059] first step:

[0060] Compound (I) was dissolved in chlorosulfonic acid and reacted in an oil bath (120-150°C) for 10-20 hours. After cooling to room temperature, the reaction was quenched with ice water, extracted with ethyl acetate, and the crude product obtained by drying and concentrating the organic phase was dissolved in anhydrous tetrahydrofuran. Ammonia was added dropwise at low temperature (-80 to -60°C) and stirring was continued for 1-5 hours. The mixture was then acidified with hydrochloric acid to obtain compound (II).

[0061] Step Two:

[0062] Compound (II) and the corresponding amine are dissolved in a solvent (such as acetonitrile), and a base (such as triethylamine or diisopropylethylamine) is added. The mixture is stirred for 10 to 20 hours under the protection of an inert gas (such as nitrogen or argon) at a temperature of 25 to 60 °C to obtain compound (III).

[0063] Process 2

[0064]

[0065] R 1 R 2 R 3 R 4 The definition is as stated above;

[0066] first step:

[0067] Phosphorus oxychloride was added dropwise to a dichloromethane solution of N,N-dimethylformamide in an ice bath under nitrogen protection. After the addition was complete, the mixture was stirred at room temperature for 30 minutes, then cooled to 0°C, and a dichloromethane solution of compound (IV) was added dropwise. The mixture was reacted at room temperature to 60°C for 10 to 24 hours to obtain compound (V).

[0068] Step Two:

[0069] Under nitrogen protection, compound (V), p-chlorophenylboronic acid, base such as potassium carbonate, phase transfer catalyst such as tetra-n-butylammonium bromide and catalyst such as palladium acetate are added to a solvent such as water. The system is evacuated and replaced with nitrogen three times, and heated to 40-100℃ for 2-10 hours to obtain compound (VI).

[0070] Step 3:

[0071] Compound (VI) was dissolved in solvents such as tetrahydrofuran and methanol, and a reducing agent such as sodium borohydride was added. The mixture was stirred at room temperature for 1 to 5 hours to obtain compound (VII).

[0072] Step 4:

[0073] Compound (VII) was dissolved in a solvent such as dichloromethane, and a chlorination reagent such as thionyl chloride was added. The mixture was stirred at room temperature for 10–24 hours to obtain compound (VIII).

[0074] Process 3

[0075]

[0076] R 0 R 1 R 2 R 3 R 4 With X 1 The definition is as stated above;

[0077] first step:

[0078] Compound (IX) (synthetic reference: WO 2017212431) (0.51 g, 2.00 mmol) and (R)-3-methylpiperazine-1-carboxylic acid tert-butyl ester were dissolved in a solvent (such as dimethyl sulfoxide), and a base such as dipotassium hydrogen phosphate was added at room temperature. The mixture was stirred at 90–120 °C for 12–48 hours to obtain compound (X).

[0079] Step Two:

[0080] Compound (X) was added to a solution of 1,4-dioxane, such as hydrogen chloride, and stirred at room temperature for 1 to 3 hours to obtain compound (XI).

[0081] Step 3:

[0082] Compound (XI) and compound (VIII) are dissolved in a solvent such as acetonitrile, and a base such as N,N-diisopropylethylamine is added. The mixture is heated to 60-90°C and stirred for 2-8 hours to obtain compound (XII).

[0083] Step 4:

[0084] Compound (XII) was dissolved in a solvent such as 2-butanol, a base such as sodium hydroxide was added, the mixture was heated to 90-120°C and stirred for 12-36 hours, and then acidified with an acid such as hydrochloric acid to obtain compound (XIII).

[0085] Step 5:

[0086] Compound (XIII), compound (III), condensing agent such as 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, and base such as 4-dimethylaminopyridine are dissolved in solvent such as dichloromethane and stirred at room temperature for 12 to 36 hours to obtain compound (XIV).

[0087] Process 4

[0088]

[0089] R 0 R 1 R 2 R 3 R 4 With X 1 The definition is as stated above;

[0090] first step:

[0091] Compound (XV) (synthesis reference: Journal of Organic Chemistry, 84(8), 4814-4829; 2019) (0.51 g, 2.00 mmol) and (R)-2-methylpiperazine were dissolved in a solvent (such as dimethyl sulfoxide), and a base such as N,N-diisopropylethylamine was added at room temperature. The mixture was stirred at 50-100 °C for 12-24 hours to obtain compound (XVI).

[0092] Step Two:

[0093] Compounds (XVI) and (VIII) were dissolved in a solvent such as acetonitrile, and a base such as N,N-diisopropylethylamine was added. The mixture was heated to 50–90 °C and stirred for 8–24 hours to obtain compound (XVII).

[0094] Step 3:

[0095] Compound (XVII) was dissolved in solvents such as water, methanol and tetrahydrofuran, and a base such as lithium hydroxide was added. The mixture was heated to 50-90°C and stirred for 1-5 hours. Then it was acidified with an acid such as hydrochloric acid to obtain compound (XVIII).

[0096] Step 4:

[0097] Compound (XVIII), compound (III), condensing agent such as 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, and base such as 4-dimethylaminopyridine are dissolved in a solvent such as dichloromethane and stirred at room temperature for 12 to 36 hours to obtain compound (XIX). Detailed Implementation

[0098] definition

[0099] Unless otherwise stated, the terms used in the specification and claims have the following meanings. Groups not specifically defined in this invention have meanings commonly understood in the art by those skilled in the art.

[0100] The notation “Cx-Cy” used in this invention represents the range of carbon atoms, where x and y are both integers. For example, C3-C8 cycloalkyl represents a cycloalkyl with 3-8 carbon atoms, and -C0-C2 alkyl represents an alkyl with 0-2 carbon atoms, where -C0 alkyl refers to a chemical single bond.

[0101] In this invention, the term "alkyl" refers to a saturated aliphatic hydrocarbon group, including straight-chain and branched groups with 1 to 20 carbon atoms, such as straight-chain and branched groups with 1 to 18 carbon atoms, 1 to 12 carbon atoms, 1 to 8 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, and various branched isomers thereof. The alkyl group may be optionally substituted or unsubstituted.

[0102] In this invention, the term "cycloalkyl" refers to a saturated monocyclic or polycyclic cyclic hydrocarbon group comprising 3 to 12 ring atoms, for example, 3 to 12, 3 to 10, 3 to 8, or 3 to 6 ring atoms, or a 3, 4, 5, or 6-membered ring. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc. The cycloalkyl group may be optionally substituted or unsubstituted.

[0103] In this invention, the term "heterocyclic group" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon group comprising 3 to 20 ring atoms, for example, 3 to 16, 3 to 12, 3 to 10, 3 to 8, or 3 to 6 ring atoms, wherein one or more ring atoms are selected from nitrogen, oxygen, or S(O)m (where m is an integer from 0 to 2), but excluding ring portions of -OO-, -OS-, or -SS-, and the remaining ring atoms are carbon. Preferably, it comprises 3 to 12 ring atoms, wherein 1 to 4 are heteroatoms; more preferably, the heterocyclic group comprises 3 to 10 ring atoms; even more preferably, it comprises 3 to 8 ring atoms; most preferably, it is a 5-membered or 6-membered ring, wherein 1 to 4 are heteroatoms; more preferably, 1 to 3 are heteroatoms; and most preferably, 1 to 2 are heteroatoms. Non-limiting examples of monocyclic heterocyclic groups include oxobutyl, pyrrolyl, piperidinyl, 4-piperidinyl, piperazine, 1,4-dioxane, morpholinyl, 2-morpholinyl, 4-morpholinyl, thiomorpholinyl, pyranyl, tetrahydropyranyl, 4-tetrahydropyranyl, homopiperazine, dioxyl, 2-dioxyl, etc. Polycyclic heterocyclic groups include spirocyclic, fused-ring, and bridged-ring heterocyclic groups. Heterocyclic groups can be optionally substituted or unsubstituted.

[0104] In this invention, the term "hybrid heterocyclic group" refers to a substituted or unsubstituted heterocyclic group having two terminal monovalent cores, which is produced by removing one hydrogen atom from each of the two terminal atoms; the heterocyclic group has the meaning described above. Non-limiting examples of "hybrid heterocyclic group" include pyrrolidinyl, piperidinyl, piperazineyl, morpholinyl, etc.

[0105] In this invention, the term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0106] In this invention, "optional" or "optionally" means that the event or environment described below may but does not have to occur, and the description includes the possibility or absence of the event or environment. For example, "optionally alkyl-substituted heterocyclic group" means that an alkyl group may but does not have to be present, and the description includes cases where the heterocyclic group is substituted with an alkyl group and cases where the heterocyclic group is not substituted with an alkyl group.

[0107] In this invention, "substituted" refers to one or more hydrogen atoms in a group, preferably up to five, and more preferably one to three hydrogen atoms, which are independently substituted by the corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art can determine (through experiment or theory) possible or impossible substitutions without much effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom having an unsaturated bond (such as an alkene).

[0108] The substituents include, but are not limited to, the various groups described above.

[0109] The compounds claimed in this invention include not only the compound itself, but also isomers of the compound, prodrugs, solvates, stable isotopic derivatives or pharmaceutically acceptable salts thereof.

[0110] The term "pharmaceutical composition" as used in this invention refers to a mixture containing one or more isomers of the compounds described in this invention, prodrugs, solvates, stable isotopic derivatives, or pharmaceutically acceptable salts thereof, and other chemical components. Other components include, for example, pharmaceutically acceptable carriers, diluents, and excipients. The purpose of the pharmaceutical composition is to facilitate administration to a living organism, thereby promoting the absorption of the active ingredient and the exertion of its biological activity.

[0111] When used in the specification, the term "comprising" includes "consisting of".

[0112] The "room temperature" mentioned in this invention refers to 15-30℃.

[0113] The "stable isotope derivatives" of this invention include: isotope-substituted derivatives obtained by replacing any hydrogen atom in formula (I) with 1-5 deuterium atoms, isotope-substituted derivatives obtained by replacing any carbon atom in formula (I) with 1-3 carbon-14 atoms, or isotope-substituted derivatives obtained by replacing any oxygen atom in formula (I) with 1-3 oxygen-18 atoms.

[0114] The “pharmaceutically acceptable salts” described in this invention are discussed in Berge, et al., “Pharmaceutically Acceptable Salts”, J. Pharm. Sci., 66, 1-19 (1977), and are obvious to medicinal chemists that the salts are substantially nontoxic and provide the desired pharmacokinetic properties, palatability, absorption, distribution, metabolism, or excretion.

[0115] The pharmaceutically acceptable salts of this invention can be synthesized by conventional chemical methods.

[0116] Generally, salts can be prepared by reacting a free base or acid with an equistoichiometric or excess amount of an acid (inorganic or organic) or base in a suitable solvent or solvent combination.

[0117] The term "prodrug" as used in this invention refers to a compound that is metabolized in vivo and converted into its original active compound. Typically, a prodrug is an inactive substance, or one with less activity than the active parent compound, but it provides convenient handling, administration, or improved metabolic properties.

[0118] The term "isomer" in this invention refers to tautomers, mesosomes, racemates, enantiomers, diastereomers, and mixtures thereof of the compound of formula (I) of this invention. All such isomers, including stereoisomers and geometric isomers, are included in this invention. Geometric isomers include cis-trans isomers.

[0119] The term "solvent" as used in this invention refers to the association of one or more solvent molecules with a compound or salt thereof of this invention. Examples of solvents that form pharmaceutically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, ethyl acetate, acetic acid, etc.

[0120] This invention includes any polymorph of the compound or its salts, as well as any hydrates or other solvates.

[0121] In this invention, the term "patient" generally refers to mammals, especially humans.

[0122] In this invention, the term "tumor" includes both benign and malignant tumors, such as cancer.

[0123] In this invention, the term "cancer" includes various tumors mediated by BCL-2, including but not limited to malignant hematologic diseases, particularly acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, prostate cancer, rectal cancer, pancreatic cancer, and glioma.

[0124] In this invention, the term "therapeutic effective amount" refers to the amount of the compound of this invention that can effectively treat or prevent related diseases mediated by BCL-2.

[0125] Example

[0126] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0127] The structures of all compounds in this invention can be identified by nuclear magnetic resonance (1H NMR) and / or mass spectrometry (MS).

[0128] 1¹H NMR chemical shifts (δ) were recorded in ppm (parts per million). NMR was performed using a Bruker AVANCE III 400 MHz spectrometer. Suitable solvents were selected from deuterated chloroform (CDCl₃), deuterated methanol (CD₃OD), and deuterated dimethyl sulfoxide (DMSO-d). 6 Tetramethylsilane was used as an internal standard (TMS).

[0129] Low-resolution mass spectrometry (MS) was performed using an Agilent 1260 HPLC / 6120 mass spectrometer with an Agilent ZORBAXXDB-C18 lens, 4.6 × 50 mm, 3.5 μm.

[0130] Gradient elution conditions: 0: 95% solvent A1 and 5% solvent B1; 1-2: 5% solvent A1 and 95% solvent B1; 2.01-2.50: 95% solvent A1 and 5% solvent B1. Percentages represent the volume percentage of a specific solvent in the total solvent volume. Solvent A1: 0.01% formic acid aqueous solution; Solvent B1: 0.01% formic acid in acetonitrile solution; percentages represent the volume percentage of the solute in the solution.

[0131] Thin-layer silica gel plates are Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates. Column chromatography generally uses Yantai Huanghai 100-200 or 200-300 mesh silica gel as the support.

[0132] Preparative liquid chromatography (prep-HPLC) was performed using a Waters SQD2 mass-guided high-performance liquid chromatograph, XBridge-C18; 30 x 150 mm preparative column, 5 μm.

[0133] Method 1: Acetonitrile-water (0.2% formic acid), flow rate 25 mL / min; Method 2: Acetonitrile-water (0.8% ammonium bicarbonate), flow rate 25 mL / min;

[0134] The known starting materials of this invention can be synthesized using or according to methods known in the art, or can be purchased from Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc., Shanghai Bid Pharmaceutical, Shanghai Aladdin Chemical, Shanghai Mairui Chemical, Bailingwei Chemical, Anai Chemical, etc. Venetoclax was purchased from WuXi AppTec (Wuhan) Chemical Technology Co., Ltd.

[0135] Unless otherwise specified in the examples, all solvents used in the reaction are anhydrous solvents. Commercially available tetrahydrofuran was used for the anhydrous tetrahydrofuran, sodium bismuth submersible was used as the dehydrating agent, benzophenone was used as the indicator, and the solution was refluxed under argon protection until it turned blue-purple. The solution was collected by distillation and stored at room temperature under argon protection. Other anhydrous solvents were purchased from Anai Chemical and Bailingwei Chemical. Unless otherwise specified, all transfers and uses of anhydrous solvents must be carried out under argon protection.

[0136] Unless otherwise specified in the examples, the reactions were carried out under an argon or nitrogen atmosphere.

[0137] Argon or nitrogen atmosphere refers to a reaction flask connected to an argon or nitrogen gas balloon with a volume of approximately 1L.

[0138] A hydrogen atmosphere refers to a reaction flask connected to a hydrogen balloon with a volume of approximately 1L.

[0139] The hydrogenation reaction is usually carried out under vacuum, filled with hydrogen gas, and repeated 3 times.

[0140] Unless otherwise specified in the examples, the reaction temperature is room temperature, and the temperature range is 15℃-30℃.

[0141] In the examples, the reaction process was monitored using thin-layer chromatography (TLC). The developing solvent systems used were A: dichloromethane and methanol; and B: petroleum ether and ethyl acetate. The volume ratio of the solvents was adjusted according to the polarity of the compounds.

[0142] The eluent systems for column chromatography and the developing solvent systems for thin-layer chromatography used to purify the compounds include: A: dichloromethane and methanol system; B: petroleum ether and ethyl acetate system. The volume ratio of the solvents is adjusted according to the polarity of the compounds, and can also be adjusted by adding small amounts of triethylamine and acidic or basic reagents.

[0143] Intermediate 1

[0144] 3-Fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)benzenesulfonamide

[0145]

[0146] first step

[0147] 3,4-Difluoro-5-nitrobenzenesulfonamide

[0148] 1,2-Difluoro-3-nitrobenzene (10.00 g, 62.89 mmol) was dissolved in chlorosulfonic acid (21 mL), and the mixture was heated to 150 °C and stirred under reflux for 10 hours. After cooling to room temperature, a saturated sodium bicarbonate aqueous solution was added to the reaction mixture under ice bath conditions to adjust the pH to approximately 7. The mixture was extracted with dichloromethane (100 mL × 3), the organic phase was washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain the crude product 3,4-difluoro-5-nitrobenzenesulfonyl chloride. Isopropanol (200 mL) and ammonia (5 mL, 37%) were added to a 1000 mL three-necked flask and stirred at -78 °C for 10 minutes. The resulting crude product, 3,4-difluoro-5-nitrobenzenesulfonyl chloride, was dissolved in isopropanol (30 mL) and slowly added dropwise to the isopropanol and ammonia mixture at -78 °C. After the addition was complete, the mixture was stirred at -78 °C for two hours. Dilute hydrochloric acid (1 N) was added to adjust the pH of the system to approximately 6. The mixture was brought to room temperature, and the solution was concentrated under reduced pressure to remove most of the isopropanol solvent. Pure water was added, and a solid precipitated out. The solid crude product was filtered to obtain the target product, 3,4-difluoro-5-nitrobenzenesulfonamide (4.90 g, yellow solid), by slurrying. Yield: 33%.

[0149] 1 H NMR (400MHz, DMSO-d6) δ8.38-8.36(m,1H),8.29-8.26(m,1H),7.84(s,2H).

[0150] Step 2

[0151] 3-Fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)benzenesulfonamide

[0152] Compound 3,4-difluoro-5-nitrobenzenesulfonamide (1.55 g, 6.51 mmol), (tetrahydro-2H-pyran-4-yl)methylamine (0.89 g, 7.73 mmol), N,N-diisopropylethylamine (3.91 g, 30.31 mmol), and acetonitrile (20.0 mL) were mixed. The mixture was stirred at 40 °C for 2 hours. The solvent was evaporated, and the mixture was quenched with 50 mL of water and extracted with ethyl acetate (60 mL × 2). The combined organic phases were washed with saturated brine (50 mL × 3). The organic phases were dried over anhydrous sodium sulfate, filtered to remove the drying agent, and dissolved under reduced pressure to obtain the crude product. Column chromatography (petroleum ether / ethyl acetate = 1:1) was used to purify the target product 3-fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)benzenesulfonamide (1.66 g, yellow solid). Yield: 76%.

[0153] MS m / z(ESI): 334[M+1];

[0154] 1H NMR(400MHz,DMSO-d6)δ8.33-8.30(m,2H),7.76-7.74(m,1H),7.45(s,2H),3.86-3.8 4(m,2H),3.50-3.45(m,2H),3.29-3.23(m,3H),1.59-1.57(m,2H),1.25-1.20(m,2H).

[0155] The synthesis steps for intermediate 2 are the same as those for intermediate 1.

[0156]

[0157] Intermediate 3

[0158] 3-Fluoro-5-nitro-4-(((1-(3-oxetane-butyl)piperidin-4-yl)methyl)amino)benzenesulfonamide

[0159]

[0160] first step

[0161] 3-Fluoro-5-nitro-4-((piperidin-4-ylmethyl)amino)benzenesulfonamide

[0162] Compound 4-(((2-fluoro-6-nitro-4-aminosulfonylphenyl)amino)methyl)piperidine-1-carboxylic acid tert-butyl ester (synthetic reference intermediate 1) (0.49 g, 1.12 mmol) was mixed with trifluoroacetic acid (3 mL) and dichloromethane (9 mL) and stirred at room temperature for 0.5 h. The solvent was evaporated, and the mixture was adjusted to neutral pH with triethylamine and evaporated again to give crude 3-fluoro-5-nitro-4-((piperidine-4-ylmethyl)amino)benzenesulfonamide (0.35 g, yellow oil). This mixture was used directly in the next reaction without purification. MS m / z (ESI): 333 [M+1].

[0163] Step 2

[0164] 3-Fluoro-5-nitro-4-(((1-(3-oxetane-butyl)piperidin-4-yl)methyl)amino)benzenesulfonamide

[0165] Compound 3-fluoro-5-nitro-4-(((piperidin-4-ylmethyl)amino)benzenesulfonamide (0.35 g, 1.05 mmol), oxetane-3-one (0.23 g, 3.15 mmol), sodium cyanoborohydride (0.23 g, 5.23 mmol), and methanol (8 mL) were mixed and stirred at room temperature for 1.5 hours. The mixture was quenched with 10 mL of water and extracted with ethyl acetate (10 mL × 2). The combined organic phases were washed with saturated brine (10 mL). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the drying agent, and evaporated to dryness. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 3:1) to give the target product 3-fluoro-5-nitro-4-(((1-(3-oxetane-4-ylmethyl)amino)benzenesulfonamide (0.32 g, yellow solid). Yield: 78%.

[0166] MS m / z(ESI): 389[M+1];

[0167] 1 H NMR(400MHz,CD3OD)δ8.47-8.46(m,1H),7.72-7.70(m,1H),4.68-4.65(m,2H),4.60-4.59(m,2H) ,3.57-3.55(m,2H),3.49-3.45(m,1H),2.83-2.80(m,2H),1.89-1.78(m,3H),1.42-1.20(m,4H).

[0168] Intermediate 4

[0169] (R)-4'-chloro-6-(chloromethyl)-3-methyl-2,3,4,5-tetrahydro-1,1'-biphenyl

[0170]

[0171] first step

[0172] (R)-2-chloro-4-methylcyclohex-1-ene-1-carboxaldehyde

[0173] N,N-dimethylformamide (0.59 g, 8.00 mmol) was dissolved in dichloromethane (10 mL), cooled to 0 °C in an ice bath, and phosphine oxychloride (0.92 g, 6.00 mmol) was added dropwise. The mixture was kept warm and stirred for 30 minutes, then allowed to rise naturally to room temperature, and stirring was continued for 3 hours. The mixture was then cooled to 0 °C, and a solution of (R)-3-methylcyclohexyl-1-one (synthesis reference: Tetrahedron 73 (2017) 3202-3212) (0.45 g, 4.0 mmol) in dichloromethane (5 mL) was slowly added. The mixture was then slowly raised to room temperature and stirred for 18 hours. The reaction solution was quenched by adding saturated sodium bicarbonate solution (10 mL), concentrated to remove dichloromethane, and then ethyl acetate (25 mL) and water (15 mL) were added. The organic phase was separated, and the aqueous phase was extracted with ethyl acetate (25 mL × 2). The combined organic phases were washed with saturated brine (20 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain the target compound (R)-2-chloro-4-methylcyclohex-1-ene-1-carboxaldehyde (0.45 g, pale yellow liquid), crude product.

[0174] MS m / z(ESI): 159 & 161 [M+1];

[0175] 1 H NMR(400MHz, CDCl3)δ10.20(s,0.8H),10.18(s,0.2H),2.63-2.47(m,2H),2.29-2.26( m,1H),2.24-2.15(m,1H),1.90-1.77(m,2H),1.27-1.21(m,1H),1.03(d,J=6.4Hz,3H).

[0176] Step 2

[0177] (R)-4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-carboxaldehyde

[0178] In a single-necked flask, (R)-2-chloro-4-methylcyclohex-1-ene-1-carboxaldehyde (0.45 g, 2.80 mmol), p-chlorophenylboronic acid (0.44 g, 2.80 mmol), tetrabutylammonium bromide (0.91 g, 2.80 mmol), potassium carbonate (1.17 g, 8.50 mmol), palladium acetate (0.16 g, 0.70 mmol), and water (15 mL) were added sequentially. The reaction system was purged with nitrogen three times and heated to 45 °C for 4 hours. The reaction mixture was cooled to room temperature, and ethyl acetate (25 mL) was added. The organic phase was separated, and the aqueous phase was extracted with ethyl acetate (25 mL). The combined organic phases were washed with saturated brine (15 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 96:4) to give the target compound (R)-4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-carboxaldehyde (0.45 g, pale yellow oil), with a yield of 67.5%.

[0179] MS m / z(ESI):235&237[M+1];

[0180] 1 H NMR(400MHz, CDCl3)δ9.41(s,0.9H),9.37(s,0.1H),7.30-7.24(m,2H),7.10-7.08(m,2H),2.5 4-2.47(m,2H),2.16-2.03(m,2H),1.82-1.75(m,2H),1.21-1.15(m,1H),0.96(d,J=6.4Hz,3H).

[0181] Step 3

[0182] (R)-(4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methanol

[0183] (R)-4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-carboxaldehyde (0.23 g, 1.00 mmol) was dissolved in tetrahydrofuran (2 mL), and sodium borohydride (57 mg, 1.50 mmol) was added at 0 °C. The mixture was stirred at room temperature for 2 hours. The reaction solution was quenched dropwise with saturated ammonium chloride aqueous solution (10 mL), extracted with ethyl acetate (10 mL), and the organic phase was washed with saturated brine (10 mL). The solution was dried, filtered, and the filtrate was concentrated to give crude (R)-(4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methanol (0.24 g, pale yellow oil). The crude product was used directly in the next step without purification. MS m / z (ESI): 237 & 239 [M-17].

[0184] Step 4

[0185] (R)-4'-chloro-6-(chloromethyl)-3-methyl-2,3,4,5-tetrahydro-1,1'-biphenyl

[0186] (R)-(4'-chloro-5-methyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methanol (0.24 g, 1.00 mmol) was dissolved in dichloromethane solution (2 mL), and sulfoxide (0.24 g, 1.50 mmol) was added at room temperature and stirred for 1 hour. The reaction solution was quenched dropwise with saturated sodium bicarbonate aqueous solution (10 mL), extracted with dichloromethane (10 mL), and the organic phase was washed with saturated brine (10 mL), dried, filtered, and the filtrate was concentrated. The reaction solution was then evaporated to dryness to obtain the crude product (R)-4'-chloro-6-(chloromethyl)-3-methyl-2,3,4,5-tetrahydro-1,1'-biphenyl (0.26 g, pale yellow oil). The crude product was used directly in the next step without purification.

[0187] The synthesis steps for intermediates 5-7 are the same as those for intermediate 4.

[0188]

[0189] Intermediate 8

[0190] 5-(chloromethyl)-6-(4-chlorophenyl)-8,8-dimethylspiro[2,5]oct-5-ene

[0191]

[0192] first step

[0193] 7,7-Dimethyl-8-methylene-1,4-dioxane[4.5]decane

[0194] 14.00 g (39.20 mmol) of methyltriphenylphosphine bromide was added to anhydrous tetrahydrofuran (120 mL), cooled to -40 °C, and a solution of n-butyllithium in n-hexane (2.5 M, 39.2 mmol, 15.6 mL) was added dropwise under nitrogen protection. After the addition was complete, the mixture was allowed to warm to room temperature and stirred for 1 hour. Then, 6.00 g (32.60 mmol) of 7,7-dimethyl-1,4-dioxaspiro[4.5]decane-8-one was added dropwise, and the reaction was allowed to proceed overnight at room temperature. The reaction mixture was quenched with saturated ammonium chloride aqueous solution (200 mL), and the aqueous phase was extracted with ethyl acetate (100 mL × 2). The combined organic phases were washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated by silica gel column chromatography (0–10% ethyl acetate / petroleum ether) to give the product 7,7-dimethyl-8-methylene-1,4-dioxane[4.5]decane (3.80 g, colorless oil). Yield: 64%.

[0195] 1 H NMR (400MHz, CDCl3) δ4.73(s,2H),3.98-3.94(m,4H),2.43-2.36(m,2H),1.73-1.68(m,2H),1.62(s,2H),1.16(s,6H).

[0196] Step 2

[0197] 4,4-Dimethyl-7,10-dioxabispirol [2.2.4] 6 .2 3 Dodecane

[0198] Compound 7,7-dimethyl-8-methylene-1,4-dioxaspiro[4.5]decane (0.90 g, 4.94 mmol) was dissolved in dichloromethane (20 mL), and a toluene solution of diethylzinc (2 M, 14.80 mmol, 7.4 mL) was added dropwise under nitrogen protection at 0 °C. The mixture was stirred for 0.5 hours after the addition was complete. Subsequently, chloroiodomethane (5.20 g, 29.60 mmol) was added dropwise, and the mixture was stirred overnight at room temperature. The reaction was quenched by adding saturated ammonium chloride aqueous solution (30 mL), and the organic phase was separated. The aqueous phase was extracted with dichloromethane (30 mL × 2), and the combined organic phases were washed with water (30 mL × 2) and saturated brine (20 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the crude product 4,4-dimethyl-7,10-dioxaspiro[2.2.4]decane. 6 .2 3 Dodecane (1.50 g, yellow oily substance). Crude product.

[0199] 1H NMR (400MHz, CDCl3) δ3.96-3.91(m,4H),1.69-1.62(m,2H),1.55(s,2H),1.44-1.40(m,2H),0.83(s,6H),0.50-0.46(m,2H),0.11-0.08(m,2H).

[0200] Step 3

[0201] 4,4-Dimethylspiro[2,5]octane-6-one

[0202] The compound 4,4-dimethyl-7,10-dioxadispiro[2.2.4] was used. 6 .2 3 Dodecane (1.50 g, crude product, 4.94 mmol) was dissolved in a mixture of tetrahydrofuran hydrochloride (2 M, 14.00 mmol, 7 mL) and stirred at room temperature for 2 hours. The reaction mixture was quenched with saturated sodium bicarbonate aqueous solution (30 mL). The mixture was extracted with ethyl acetate (30 mL), and the organic phase was washed with water (30 mL × 2) and saturated brine (30 mL), respectively. The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the crude product 4,4-dimethylspiro[2.5]octane-6-one (0.70 g, yellow oil). Yield: 93.2%. MS m / z (ESI): 153 [M+1].

[0203] Step 4

[0204] 6-Chloro-8,8-dimethylspiro[2,5]oct-5-ene-5-carboxaldehyde

[0205] Under nitrogen protection, phosphorus oxychloride (1.10 g, 6.90 mmol) was added dropwise to a dichloromethane solution (30 mL) of N,N-dimethylformamide (0.67 g, 9.20 mmol) in an ice bath. After the addition was complete, the mixture was stirred at room temperature for 30 minutes, then cooled to 0 °C, and a dichloromethane solution (5 mL) of 4,4-dimethylspiro[2.5]octane-6-one (0.70 g, 4.60 mmol) was added dropwise. The mixture was allowed to react overnight at room temperature. The reaction was quenched with water (30 mL), and the organic phase was separated. The organic phase was washed with saturated sodium bicarbonate solution (30 mL) and saturated brine (30 mL), respectively. The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (100% petroleum ether) to obtain 6-chloro-8,8-dimethylspiro[2.5]octane-5-en-5-carboxaldehyde (0.34 g, pale yellow oil). Yield: 37.2%. MS m / z(ESI): 199&201[M+1].

[0206] Step 5

[0207] 6-(4-Chlorophenyl)-8,8-dimethylspiro[2,5]oct-5-ene-5-carboxaldehyde

[0208] Under nitrogen protection, 6-chloro-8,8-dimethylspiro[2.5]oct-5-ene-5-carboxaldehyde (0.34 g, 1.70 mmol), p-chlorophenylboronic acid (0.41 g, 2.60 mmol), potassium carbonate (0.70 g, 5.10 mmol), tetrabutylammonium bromide (0.55 g, 1.70 mmol), and palladium acetate (77 mg, 0.34 mmol) were added to water (15 mL). The system was purged with nitrogen three times and heated to 50 °C for 4 hours. After cooling to room temperature, the reaction solution was extracted with ethyl acetate (20 mL × 2), the combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-5% ethyl acetate / petroleum ether) to give 6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-ene-5-carboxaldehyde (0.18 g, pale yellow oil). Yield: 38.2%. MS m / z (ESI): 275 & 277 [M+1].

[0209] Step 6

[0210] (6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methanol

[0211] 6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-carboxaldehyde (0.18 g, 0.66 mmol) was dissolved in a tetrahydrofuran-methanol mixture (v / v = 6 / 1, 7 mL), and sodium borohydride (75 mg, 1.78 mmol) was added. The mixture was stirred at room temperature for 2 hours. The reaction was quenched with saturated ammonium chloride aqueous solution (20 mL). The reaction solution was extracted with ethyl acetate (20 mL × 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give (6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methanol (0.16 g, colorless oil). Yield: 88.2%. MS m / z (ESI): 260 & 262 [M-17].

[0212] Step 7

[0213] 5-(chloromethyl)-6-(4-chlorophenyl)-8,8-dimethylspiro[2,5]oct-5-ene

[0214] (6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methanol (0.16 g, 0.58 mmol) was dissolved in dichloromethane (10 mL), and thionyl chloride (0.21 g, 1.74 mmol) was added. The mixture was stirred overnight at room temperature. The solution was concentrated under reduced pressure to give 5-(chloromethyl)-6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-ene (0.17 g, yellow oil). Crude product.

[0215] 1 H NMR (400MHz, CDCl3) δ7.36-7.29(m,2H),7.20-7.13(m,2H),3.88(s,2H),2.20-2. 11(m,2H),1.64-1.53(m,2H),0.85(s,6H),0.63-0.57(m,2H),0.24-0.16(m,2H).

[0216] Intermediate 9

[0217] 7,7,8,8-Tetramethyl-1,4-dioxaspiro[4.5]decane

[0218]

[0219] 7,7,8,8-Tetramethyl-1,4-dioxaspiro[4.5]decane

[0220] The compound 4,4-dimethyl-7,10-dioxadispiro[2.2.4] was used. 6 .2 3 Dodecane (see intermediate 8, step 2) (4.30 g, 21.9 mmol) and platinum dioxide (1.00 g, 4.40 mmol) were added to acetic acid (30 mL). The reaction system was evacuated and purged with hydrogen, then heated to 40 °C and reacted overnight. After cooling to room temperature, ethyl acetate (30 mL) was added for dilution. The reaction solution was filtered through diatomaceous earth, and the filter cake was washed with ethyl acetate (20 mL). The filtrate was concentrated under reduced pressure to obtain the crude product 7,7,8,8-tetramethyl-1,4-dioxane[4.5]decane (4.50 g, colorless oil). Crude product. MS m / z (ESI): 199 [M+1].

[0221] The synthesis of intermediate 10 follows the same steps as the synthesis of intermediate 8.

[0222]

[0223] Example 1

[0224] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0225]

[0226] first step

[0227] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(3-methylpiperazin-1-yl)benzoate

[0228] Methyl 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-fluorobenzoate (synthesis reference: Journal of Organic Chemistry, 84(8), 4814-4829; 2019) (2.86 g, 10.00 mmol), (R)-2-methylpiperazine (3.00 g, 30.00 mmol), N,N-diisopropylethylamine (3.12 g, 24.18 mmol), and dimethyl sulfoxide (20 mL) were mixed. The mixture was stirred at 60 °C for 16 hours. The mixture was diluted with 50 mL of water and extracted with ethyl acetate (60 mL × 2). The combined organic phases were washed with saturated brine (50 mL × 3). The organic phases were dried over anhydrous sodium sulfate, filtered to remove the drying agent, and dissolved under reduced pressure to obtain the crude product. Column chromatography purification (dichloromethane / methanol = 90:10) yielded the target product (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(3-methylpiperazin-1-yl)benzoate (2.55 g, yellow solid). Yield: 70%.

[0229] MS m / z(ESI): 367[M+1];

[0230] 1H NMR (400MHz, CDCl3) δ9.92 (s, 1H), 8.19 (d, J = 2.4Hz, 1H), 7.91 (d, J = 9.2Hz, 1H) ,7.52(d,J=2.4Hz,1H),7.36-7.35(m,1H),6.66-6.63(m,1H),6.43(d,J=1.6Hz, 1H),6.34(d,J=2.4Hz,1H),3.79(s,3H),3.54-3.48(m,2H),3.05-3.02(m,1H), 2.93-2.84(m,2H),2.77-2.71(m,1H),2.42-2.37(m,1H),1.08(d,J=6.4Hz,3H).

[0231] Step 2

[0232] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)methyl benzoate

[0233] The compound (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(3-methylpiperazin-1-yl)benzoate (34 mg, 0.09 mmol), 4'-chloro-6-(chloromethyl)-3,3-dimethyl-2,3,4,5-tetrahydro-1,1'-biphenyl (synthesis reference: US 20100298323) (30 mg, 0.11 mmol), N,N-diisopropylethylamine (36 mg, 0.28 mmol), and acetonitrile (5.0 mL) were mixed. The mixture was stirred at 60 °C for 16 hours. The mixture was quenched with 10 mL of water and extracted with ethyl acetate (10 mL × 2). The combined organic phases were washed with saturated brine (10 mL × 2). The organic phases were dried over anhydrous sodium sulfate, filtered to remove the drying agent, and dissolved under reduced pressure to obtain the crude product. Purification using preparative stenography (petroleum ether / ethyl acetate = 1:1) yielded the target product, methyl (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)benzoate (28 mg, colorless oily liquid). Yield: 50%. MS m / z (ESI): 599 & 601 [M+1].

[0234] Step 3

[0235] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid

[0236] Compound (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-(((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)benzoate (28 mg, 0.05 mmol), lithium hydroxide (20 mg, 0.83 mmol), methanol (1 mL), tetrahydrofuran (1 mL), and water (1 mL) were mixed and stirred at 60 °C for 1.5 h. The mixture was adjusted to pH neutral with 1 N hydrochloric acid and extracted with ethyl acetate (10 mL × 2). The combined organic phases were washed with saturated brine (10 mL). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the drying agent, and dissolved under reduced pressure to obtain crude (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid (25.6 mg, crude). This mixture was used directly in the next reaction without purification. MS m / z (ESI): 585 & 587 [M+1].

[0237] Step 4

[0238] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0239] Compound (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid (26 mg, 0.04 mmol), 3-fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)benzenesulfonamide (intermediate 1) (15 mg, 0.04 mmol), 4-dimethylaminopyridine (11 mg, 0.09 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (17 mg, 0.09 mmol), and dichloromethane (2.5 mL) were mixed. The mixture was stirred at room temperature for 16 hours. The mixture was quenched with 10 mL of water and extracted with dichloromethane (10 mL × 2). The combined organic phases were washed with saturated brine (10 mL). The organic phase was dried over anhydrous sodium sulfate, filtered to remove the drying agent, and dissolved under reduced pressure to obtain the crude product. Silica gel plate purification (dichloromethane / methanol = 15:1) yielded the target product (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-fluoro-5-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide 1 (15.7 mg, yellow solid). Yield: 38%.

[0240] MS m / z(ESI): 900 & 902 [M+1];

[0241] 1H NMR (400MHz, CD3OD) δ8.49 (s, 1H), 8.08-8.05 (m, 1H), 7.71-7.64 (m, 2H), 7.50 (s, 1H), 7.44 (d, J = 3.6Hz, 1H), 7.36-7.34 ( m,2H),7.09-7.07(m,2H),6.77(d,J=8.0Hz,1H),6.42-6.36(m,2H),4.04-3.93(m,3H),3.71-3.67(m,2H),3.52-3.38(m,5 H),3.28-3.26(m,2H),3.08-3.02(m,1H),2.96-2.93(m,1H),2.65-2.63(m,1H),2.24-2.15(m,2H),2.06-2.04(m,1H),1.9 3-1.89(m,1H),1.85(d,J=4.0Hz,3H),1.70-1.64(m,3H),1.58-1.56(m,2H),1.39-1.28(m,2H),1.01(s,3H),0.90(s,3H).

[0242] The synthesis steps of Examples 2 to 7 are the same as those of Example 1.

[0243]

[0244]

[0245] Example 8

[0246] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0247]

[0248] first step

[0249] (R)-4-(3-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-cyanophenyl)-3-methylpiperazine-1-carboxylic acid tert-butyl ester

[0250] Compound 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-fluorobenzonitrile (synthesis reference: WO2017212431) (0.51 g, 2.00 mmol) and (R)-3-methylpiperazine-1-carboxylic acid tert-butyl ester (1.20 g, 6.00 mmol) were dissolved in dimethyl sulfoxide (15 mL), and dipotassium hydrogen phosphate (1.05 g, 6.00 mmol) was added at room temperature. The mixture was stirred at 110 °C for 36 hours. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate (30 mL × 2). The combined organic phases were washed with saturated brine (30 mL × 2), dried, and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give (R)-4-(3-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-cyanophenyl)-3-methylpiperazine-1-carboxylic acid tert-butyl ester (0.35 g, white solid), yield 41%.

[0251] MS m / z(ESI): 434[M+1];

[0252] 1 H NMR (400MHz, CDCl3) δ9.29 (s, 1H), 8.19 (s, 1H), 7.69 (s, 1H), 7.49 (d, J = 8.8Hz, 1H), 7.42-7.40 (m, 1H), 6.53-6.49 (m,2H),6.08(s,1H),40.5-3.83(m,3H),3.21-3.20(m,2H),3.04-2.95(m,2H),1.44(s,9H),1.02(d,J=6.4Hz,3H).

[0253] Step 2

[0254] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(2-methylpiperazin-1-yl)benzonitrile

[0255] (R)-4-(3-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-cyanophenyl)-3-methylpiperazin-1-carboxylic acid tert-butyl ester (0.35 g, 0.81 mmol) was dissolved in dioxane hydrochloride solution (4 M, 20.00 mmol, 5 mL) and stirred at room temperature for 1 hour. The reaction solution was evaporated to dryness to give crude product (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(2-methylpiperazin-1-yl)benzonitrile (0.38 g, crude product), which was used directly in the next step without purification. MS m / z (ESI): 334 [M+1].

[0256] Step 3

[0257] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)benzonitrile

[0258] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(2-methylpiperazin-1-yl)benzonitrile (0.38 g, crude) and 4'-chloro-6-(chloromethyl)-3,3-dimethyl-2,3,4,5-tetrahydro-1,1'-biphenyl (0.21 g, 0.80 mmol) (synthetic reference: US 20100298323) were dissolved in acetonitrile (10 mL), and N,N-diisopropylethylamine (0.31 g, 2.40 mmol) was added. The mixture was stirred at 80 °C for 4 hours. After cooling to room temperature, the reaction mixture was diluted with water and extracted with ethyl acetate (20 mL × 2). The combined organic phases were washed with saturated brine (20 mL). The organic phase was dried and concentrated to obtain a crude product, which was then purified by silica gel column chromatography (petroleum ether: ethyl acetate = 2:1) to give (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)benzonitrile (0.30 g, white solid), yield 66%.

[0259] MS m / z(ESI): 566 & 568 [M+1];

[0260] 1 H NMR (400MHz, CDCl3) δ9.40(s,1H),8.19(s,1H),7.68(s,1H),7.43(d,J=8.8Hz,1H),7.42-7.40(m,1H ),7.23(d,J=8.4Hz,2H),6.92(d,J=8.4Hz,2H),6.52-6.45(m,2H),6.04(s,1H),3.81-3.78(m,1H),3 .16-3.13(m,1H),2.98-2.95(m,1H),2.69-2.65(m,3H),2.54-2.51(m,1H),2.21-2.14(m,2H),2.04- 1.97(m,2H),1.89-1.76(m,2H),1.43-1.40(m,2H),1.07(d,J=6.4Hz,3H),0.94(s,3H),0.93(s,3H).

[0261] Step 4

[0262] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)benzoic acid

[0263] Compound (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)benzonitrile (0.11 g, 0.20 mmol) was dissolved in 2-butanol (3 mL), and sodium hydroxide (160 mg, 4.00 mmol) was added at 90 °C. The mixture was heated to 105 °C and stirred for 24 hours. The reaction solution was desolvated under reduced pressure, and the residue was extracted with 1N hydrochloric acid aqueous solution (4 mL) and ethyl acetate (10 mL × 2). The combined organic phases were washed with saturated brine (10 mL). The organic phase was dried and concentrated to obtain a crude product, which was then purified using a preparative silica gel plate (dichloromethane:methanol = 10:1) to give (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)benzoic acid (90 mg, white solid), yield 77%. MS m / z (ESI): 585 & 587 [M+1].

[0264] Step 5

[0265] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0266] Following the final step of the synthesis as described in Example 1, the target product (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide 8 was obtained.

[0267] MS m / z(ESI): 882 & 884 [M+1];

[0268] 1 H NMR (400MHz, CDCl3) δ10.13(s,1H),9.29(s,1H),8.88(s,1H),8.53-8.50(m,1H),8.22-8.16(m,2H),7.94(d,J=8.8Hz,1H),7.70(s, 1H),7.47-7.45(m,1H),7.22(d,J=8.0Hz,2H),6.93-6.89(m,3H),6.56-6.49(m,2H),5.94(s,1H),4.05-4.01(m,2H),3.78-3.74(m, 1H),3.45-3.39(m,2H),3.28-3.25(m,2H),3.15-3.12(m,1H),2.96-2.89(m,1H),2.73-2.62(m,3H),2.51-2.48(m,1H),2.20-2.11( m,2H),1.99-1.95(m,3H),1.86-1.83(m,1H),1.79-1.72(m,3H),1.45-1.39(m,4H),1.03(d,J=6.4Hz,3H),0.93(s,3H),0.92(s,3H).

[0269] The synthesis steps of Examples 9 to 13 are the same as those of Example 8.

[0270]

[0271]

[0272] The synthesis steps of Examples 14 to 22 are the same as those of Example 1.

[0273]

[0274]

[0275]

[0276] The synthesis steps of Examples 23 to 26 are the same as those of Example 8.

[0277]

[0278]

[0279] The synthesis steps of Examples 27 to 30 are the same as those in Example 1.

[0280]

[0281]

[0282] Example 31

[0283] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0284]

[0285] first step

[0286] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2,5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)methyl benzoate

[0287] 5-(chloromethyl)-6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-ene (intermediate 8) (0.17 g, 0.58 mmol), (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(3-methylpiperazin-1-yl)benzoate and N,N-diisopropylethylamine (0.37 g, 2.90 mmol) were dissolved in acetonitrile (15 mL) and heated under reflux for 3 hours. After cooling, the mixture was concentrated under reduced pressure. The residue was dissolved in ethyl acetate (30 mL), washed with water (30 mL × 3) and saturated brine (30 mL), respectively. The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (10%–40% ethyl acetate / petroleum ether) to give (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)benzoate (0.10 g, white solid). Yield: 27.7%. MS m / z (ESI): 625 & 627 [M+1].

[0288] Step 2

[0289] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2,5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid

[0290] Methyl (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)benzoate (0.10 g, 0.16 mmol) was dissolved in ethanol (3 mL) and sodium hydroxide aqueous solution (2 N, 6.00 mmol, 3 mL), and the mixture was heated to 70 °C and reacted for 2 hours. After cooling, the mixture was diluted with water (20 mL), and the pH was adjusted to approximately 5 with 1 N dilute hydrochloric acid. The aqueous phase was extracted with ethyl acetate (20 mL × 2). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid (80 mg, white solid). Yield: 81.8%. MS m / z (ESI): 611 & 613 [M+1].

[0291] Step 3

[0292] (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0293] Compounds (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)benzoic acid (20 mg, 0.03 mmol), 3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)benzylsulfonamide (synthetic reference: WO 2018041248 A1) (10 mg, 0.03 mmol), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (31 mg, 0.17 mmol) and 4-dimethylaminopyridine (8 mg, 0.07 mmol) were dissolved in dichloromethane (5 mL) and stirred overnight at room temperature. Dilute with dichloromethane (20 mL), wash the reaction solution with water (20 mL × 3) and saturated brine (20 mL), respectively, dry the organic phase with anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. Purify the residue by preparative thin-layer chromatography (dichloromethane / methanol = 15 / 1) to give the target product (R)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-(4-((6-(4-chlorophenyl)-8,8-dimethylspiro[2.5]oct-5-en-5-yl)methyl)-3-methylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide 31 (16.6 mg, yellow solid), yield: 55.7%.

[0294] MS m / z(ESI): 908 & 910 [M+1];

[0295] 1H NMR (400MHz, CDCl3) δ10.14(brs,1H),9.70(s,1H),8.96-8.82(m,1H),8.57-8.48(m,1H),8.27-8.09(m,2H),8.00-7.9 0(m,1H),7.75-7.63(m,1H),7.50-7.43(m,1H),7.25-7.19(m,2H),7.01-6.85(m,3H),6.59-6.49(m,2H),5.98(d,J=2.0 Hz,1H),4.10-3.95(m,2H),3.48-3.38(m,2H),3.32-3.17(m,4H),3.12-3.03(m,1H),2.87-2.73(m,1H),2.70-2.45(m, 3H),2.18-2.04(m,3H),2.03-1.83(m,3H),1.50-1.19(m,6H),0.93-0.78(m,8H),0.60-0.48(m,2H),0.15-0.02(m,2H).

[0296] The synthesis steps of Examples 32 to 34 are the same as those of Example 31.

[0297]

[0298] The synthesis steps of Examples 35 to 38 are the same as those of Example 1.

[0299]

[0300]

[0301] The synthesis steps of Examples 39 to 42 are the same as those of Example 31.

[0302]

[0303]

[0304] The synthesis steps of Examples 43 to 46 are the same as those of Example 8.

[0305]

[0306]

[0307] The synthesis steps of Examples 47 to 56 are the same as those of Example 1.

[0308]

[0309]

[0310]

[0311]

[0312] Example 57

[0313] 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((4aR,8aR)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)octahydroquinoxalin-1(2H)-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0314]

[0315] Example 57 The synthesis was performed following the steps of Example 1, except that in the second step, (R)-2-methylpiperazine was replaced with (4aR,8aR)-decahydroquinoxaline (synthesis reference: European Journal of Organic Chemistry, 2012(34), 6752-6759; 2012) to obtain the target compound 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((4aR,8aR)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)octahydroquinoxaline-1(2H)-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide 57.

[0316] MS m / z(ESI): 822 & 824 [M+1];

[0317] 1H NMR(400MHz, CDCl3)δ10.21(s,1H),8.93-8.92(m,1H),8.88-8.87(m,1H) ,8.57-8.53(m,1H),8.20-8.18(m,2H),8.01(d,J=8.4Hz,1H),7.71(d,J=1 .6Hz,1H),7.46-7.44(m,1H),7.20(d,J=8.0Hz,2H),6.93(m,1H),6.89(d, J=8.0Hz,2H),6.73-6.70(m,1H),6.57-6.56(m,1H),6.17(d,J=1.6Hz,1H) ,4.06-4.02(m,2H),3.45-3.41(m,3H),3.29-3.26(m,2H),3.19-3.16(m, 1H),3.04-3.01(m,1H),2.81-2.69(m,2H),2.49-2.43(m,2H),2.22-2.19( m,1H),1.98-1.96(m,4H),1.96-1.92(m,4H),1.85-1.82(m,1H),1.76-1.7 3(m,3H),1.47-1.44(m,3H),1.41-1.35(m,4H),0.96(s,3H),0.93(s,3H).

[0318] Example 58

[0319] 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((2R,5R)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2,5-dimethylpiperazin-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide

[0320]

[0321] Example 58 The synthesis was performed following the steps of Example 1, except that in the second step, (R)-2-methylpiperazine was replaced with (2R,5R)-2,5-dimethylpiperazine (synthesis reference: Organic Chemistry Frontiers, 5(23), 3402-3405; 2018) to obtain the target compound 2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((2R,5R)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-2,5-dimethylpiperazine-1-yl)-N-((3-nitro-4-(((tetrahydro-2H-pyran-4-yl)methyl)amino)phenyl)sulfonyl)benzamide 58.

[0322] MS m / z(ESI): 896 & 898 [M+1];

[0323] 1 H NMR (400MHz, CDCl3) δ10.10(s,1H),9.25(s,1H),8.88(d,J=2.2Hz,1H),8.53-8.51(m,1H),8.20(d,J=2.5Hz,1H),8.19-8.02(m,1H),7.94(d,J= 9.2Hz,1H),7.70(d,J=2.4Hz,1H),7.48-7.42(m,1H),7.22(d,J=8.4Hz, 2H),6.99-6.87(m,2H),6.59-6.52(m,1H),6.50-6.40(m,1H),5.95(d,J =2.2Hz,1H),4.10-4.01(m,2H),3.70-3.60(m,2H),3.47-3.37(m,2H),3 .33-3.21(m,2H),3.19-3.03(m,2H),2.59-2.50(m,2H),2.39-2.31(m,2 H),1.97-1.90(m,3H),1.82-1.71(m,2H),1.46-1.33(m,3H),1.28-1.20 (m,3H),1.25-1.15(m,4H),1.02-1.00(m,2H),0.95(s,3H),0.93(s,3H).

[0324] Example 59

[0325] N-((4-((((S)-1,4-dioxane-2-yl)methyl)amino)-3-nitrophenyl)sulfonyl)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((3R,5R)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3,5-dimethylpiperazin-1-yl)benzamide

[0326]

[0327] Example 59 was synthesized following the synthetic steps of Example 1, except that in the second step, (2R,6R)-2,6-dimethylpiperazine was used instead of (R)-2-methylpiperazine to obtain the target compound N-((4-((((S)-1,4-dioxane-2-yl)methyl)amino)-3-nitrophenyl)sulfonyl)-2-((1H-pyrrolo[2,3-b]pyridin-5-yl)oxy)-4-((3R,5R)-4-((4'-chloro-5,5-dimethyl-3,4,5,6-tetrahydro-[1,1'-biphenyl]-2-yl)methyl)-3,5-dimethylpiperazine-1-yl)benzamide 59.

[0328] MS m / z(ESI): 898 & 890 [M+1];

[0329] 1 H NMR (400MHz, CDCl3) δ9.16 (s, 1H), 8.86 (d, J = 2.2Hz, 1H), 8.61 (s, 1H), 8.22-8.13 (m, 2H), 7.91 (d, J = 9.1Hz, 1H), 7.67-7.6 0(m,1H),7.44(s,1H),7.23(d,J=8.3Hz,2H),6.93(d,J=8.3Hz,2H),6.88(d,J=9.2Hz,1H),6.60-6.48(m,2H),5.94-5.90( m,1H),3.97-3.85(m,2H),3.85-3.74(m,2H),3.67-3.60(m,2H),3.55-3.29(m,3H),3.13-3.02(m,2H),2.79-2.70(m,2H), 2.69-2.59(m,2H),2.24-2.15(m,2H),2.12-2.05(m,4H),2.05-1.86(m,2H),0.93-0.91(m,6H),0.69(s,3H),0.67(s,3H).

[0330] Biological experiments

[0331] Tests of BCL-2 bioactivity inhibition

[0332] The effect of the compounds of this invention on the bioactivity of BCL-2 was evaluated using fluorescence polarization assays.

[0333] The experimental methods are summarized as follows:

[0334] An affinity assay based on fluorescence polarization was used to assess the effect of compounds on the bioactivity of BCL-2 by detecting their binding activity to BCL-2 and leukemia pro-apoptotic protein (BIM). The reaction buffer contained the following components: PBS (pH 7.4, 3 mM Na2HPO4, 155 mM NaCl, 1 mM KH2PO4), 1 mM DTT; recombinant human Bcl-2 protein (catalog number 10195-H08E) was purchased from Beijing Yiqiao Shenzhou Biotechnology Co., Ltd., and diluted to 5 nM with the reaction buffer; FITC-labeled BIM peptide was purchased from Nanjing Genscript Biotech Co., Ltd., and diluted to 5 nM with the reaction buffer.

[0335] The compound was dissolved and diluted in 100% DMSO to 0.1, 1, and 10 μM, and then serially diluted 4-fold with DMSO to the lowest concentrations of 0.0061, 0.061, and 0.61 nM. Each concentration point was then diluted 50-fold with reaction buffer.

[0336] Add 3 μL of the compound solution and 12 μL of BCL-2 solution to a black 384-well plate, mix thoroughly, and incubate at room temperature for 15 minutes. Then add 15 μL of FITC-BIM solution, incubate the reaction mixture at room temperature in the dark for 30 minutes, and immediately detect fluorescence polarization on an Envision microplate reader (Perkin Elmer) with excitation wavelength of 480 nm and emission wavelength of 535 nm. In this experiment, the group without BCL-2 protein served as a negative control (100% inhibition), and the group with BCL-2 protein but without the compound served as a positive control (0% inhibition). The percentage of inhibition of BCL-2 affinity by the compound can be calculated using the following formula:

[0337] The IC50 values ​​of the compounds were calculated using the following formula at eight concentration points with XLfit (ID Business Solutions Ltd., UK) software:

[0338] Y=Bottom+(Top-Bottom) / (1+10^((logIC 50 -X)*slope factor))

[0339] Where Y is the inhibition percentage, X is the logarithm of the concentration of the analyte, Bottom is the maximum inhibition percentage, Top is the minimum inhibition percentage, and slope factor is the curve slope coefficient.

[0340] Tests of BCL-XL bioactivity inhibition

[0341] The effect of the compounds of this invention on the bioactivity of BCL-XL was evaluated using fluorescence polarization assays.

[0342] The experimental methods are summarized as follows:

[0343] An affinity assay based on fluorescence polarization was used to assess the effect of compounds on the binding activity of BCL-XL and BIM, thereby evaluating the effect of compounds on the bioactivity of BCL-XL. The reaction buffer contained the following components: PBS (pH 7.4, 3 mM Na2HPO4, 155 mM NaCl, 1 mM KH2PO4), 1 mM DTT; human recombinant Bcl-XL protein (catalog number 10455-H08E) was purchased from Beijing Yiqiao Shenzhou Biotechnology Co., Ltd., and diluted to 10 nM with the reaction buffer; FITC-labeled BIM peptide was purchased from Nanjing Genscript Biotech Co., Ltd., and diluted to 10 nM with the reaction buffer.

[0344] The compound was dissolved and diluted to 1 μM in 100% DMSO, and then serially diluted 4-fold with DMSO to the lowest concentration of 0.061 nM. Each concentration point was then diluted 50-fold with reaction buffer.

[0345] Add 3 μl of the compound solution and 12 μL of BCL-XL solution to a black 384-well detection plate, mix thoroughly, and incubate at room temperature for 15 minutes. Then add 15 μL of FITC-BIM solution, incubate the reaction mixture at room temperature in the dark for 30 minutes, and immediately detect fluorescence polarization on an Envision microplate reader (Perkin Elmer) with excitation wavelength of 480 nm and emission wavelength of 535 nm. In this experiment, the group without BCL-XL protein served as a negative control (100% inhibition), and the group with BCL-XL protein but without the compound served as a positive control (0% inhibition). The percentage of inhibition of BCL-XL affinity by the compound can be calculated using the following formula:

[0346] Compound IC 50 The values ​​were calculated from eight concentration points using XLfit (ID Business Solutions Ltd., UK) software via the following formula:

[0347] Y=Bottom+(Top-Bottom) / (1+10^((logIC 50-X)*slope factor))

[0348] Where Y is the inhibition percentage, X is the logarithm of the concentration of the analyte, Bottom is the maximum inhibition percentage, Top is the minimum inhibition percentage, and slope factor is the curve slope coefficient.

[0349] RS4;11 cells (acute lymphoblastic leukemia cells) IC50 50 Measurement

[0350] The effect of the compounds of the present invention on the proliferation of RS4;11 cells was evaluated using a luminescence cell viability assay.

[0351] The experimental methods are summarized as follows:

[0352] The CellTilter-Glo (CTG) assay kit was used to detect the key indicator of ATP metabolism in living cells by employing a unique, highly sensitive and stable luciferase. The luminescent signal generated in the experiment was directly proportional to the number of viable cells in the culture medium, thereby detecting the cell proliferation status of RS4;11.

[0353] CellTilter-Glo reagent (Promega, G7572) consists of CellTilter-Glo lyophilized powder and CellTilter-Glo buffer. To use, simply dissolve the lyophilized powder in the buffer.

[0354] RS4;11 cells (ATCC, CRL-1873) were cultured in RPMI 1640 complete medium (Thermofisher, 72400-047) containing 10% FBS (GBICO, 10099-141) and 100 units / ml penicillin-streptomycin mixture (Thermofisher, 15140122). When the cell coverage in the culture vessel reached 80-90%, the cells were digested with 0.25% trypsin (containing EDTA) (Thermofisher, 25200056), dispersed, and seeded into white 384-well plates (Thermofisher, 164610). The 384-well plates were then incubated overnight at 37°C with 5% CO2. The compound was dissolved and diluted to 5 mM in 100% DMSO, then serially diluted 4-fold with DMSO to a minimum concentration of 0.061 μM. Each concentration point was then diluted 50-fold using FBS-free RPMI 1640 medium. If the compound IC50... 50The value is very low, allowing for a reduction in the initial concentration of the compound. After overnight incubation, 3 μL of the diluted compound in culture medium was added to each well, and the mixture was gently centrifuged to mix. The group without cells served as a negative control (100% inhibition), while the group with 0.2% DMSO served as a positive control (0% inhibition). The 384-well plate was incubated at 37°C with 5% CO2 for 48 hours. After equilibration to room temperature, 15 μL of CTG reagent was added to each well, and the plate was gently shaken on a shaker for 3 minutes to ensure complete cell lysis. The plate was then left to stand for 10 minutes to allow the cryo-light signal to stabilize, and the cryo-light signal was read using an EnVision (Perkin Elmer) scanner.

[0355] The percentage of inhibition of RS4;11 cell proliferation by the compound can be calculated using the following formula:

[0356] Inhibition percentage = 100 - 100 * (signal compound - signal negative control) / (signal positive control - signal negative control)

[0357] Compound IC 50 The values ​​were calculated from eight concentration points using XLfit (ID Business Solutions Ltd., UK) software via the following formula:

[0358] Y=Bottom+(Top-Bottom) / (1+10^((LogIC 50 -X)*slope factor))

[0359] Where Y represents the inhibition percentage, Bottom is the bottom plateau of the curve, Top is the top plateau of the curve, and X is the logarithm of the concentration of the analyte.

[0360] The results of the above in vitro BCL-2 and BCL-XL protein activity assays are shown in Table 1 below.

[0361] The results of the cell experiments are shown in Table 2.

[0362] Table 1: Results of BCL-2 and BCL-XL protein activity assays

[0363]

[0364]

[0365]

[0366] Table 2: Results of RS4;11 cell viability assay

[0367]

[0368]

[0369]

[0370] The experimental results above show that the compounds in the embodiments of the present invention can effectively and selectively inhibit the activity of BCL-2, with weaker inhibition of BCL-XL. They can be used to treat various cancers caused by abnormal overexpression of BCL-2 family proteins, especially malignant hematological diseases such as acute lymphoblastic leukemia, lung cancer, breast cancer, ovarian cancer, rectal cancer, prostate cancer, pancreatic cancer, and glioma. Furthermore, they avoid the toxic side effects caused by BCL-XL inhibition, such as thrombocytopenia. Some compounds can also effectively inhibit the proliferation of RS4;11 acute lymphoblastic cells. They have a strong inhibitory effect on malignant hematological diseases such as acute lymphoblastic leukemia.

[0371] It will be apparent to those skilled in the art that this disclosure is not limited to the illustrative embodiments described above, and that it may be embodied in other specific forms without departing from the essential characteristics of this disclosure. Therefore, it is intended that these embodiments be considered illustrative and non-limiting in all respects, and that reference be made to the appended claims rather than the foregoing embodiments, and that all variations therein within the equivalent meaning and scope of the claims are included therein.

Claims

1. The following compounds or their pharmaceutically acceptable salts are selected from:

2. Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for the treatment or prevention of BCL-2-mediated diseases, said diseases being selected from acute lymphoblastic leukemia, malignant hematologic diseases, lung cancer, breast cancer, ovarian cancer, rectal cancer, prostate cancer, pancreatic cancer, and glioma.

3. A pharmaceutical composition comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof, optionally one or more other BCL-2 inhibitors, and one or more pharmaceutically acceptable carriers, diluents, and excipients.

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