1,3-disubstituted piperidine derivatives, processes for their preparation and use

Abstract

The application belongs to the technical field of medicines, and particularly relates to a 1,3-disubstituted piperidine derivative and a preparation method and application thereof. The application provides a 1,3-disubstituted piperidine derivative and a preparation method and application thereof, wherein the 1,3-disubstituted piperidine derivative is a non-adenosine DOT1L inhibitor; the use of an adenosine or adenosine-like mother nucleus is avoided in the structure, the metabolic stability is higher, the properties are stable, the 1,3-disubstituted piperidine derivative is easy to store, the preparation route is simple and easy to implement, and the 1,3-disubstituted piperidine derivative is easy to purify.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a 1,3-disubstituted piperidine derivative and its preparation method and application. Background Technology

[0002] Piperidine is a six-membered heterocyclic compound containing a nitrogen atom, and its structure is widely found in natural products and drugs. In medicinal chemistry, introducing piperidine and its derivatives into drug molecules can improve efficacy and bioavailability, facilitating more effective absorption and utilization of drugs. Furthermore, piperidine and its derivatives can be applied to drug delivery systems, enhancing drug solubility, stability, and delivery efficiency through binding. Additionally, as non-adenosine compounds, piperidine and its derivatives exhibit higher metabolic stability in vivo compared to adenosine compounds, resulting in better pharmacokinetic properties. Recent studies have also found that piperidine derivatives demonstrate advantages in tumor research, including inhibiting tumor cell growth, suppressing kinase activity, slowing tumor angiogenesis, and reversing cancer cell drug resistance, making it a promising structural type for anti-tumor drugs.

[0003] DOT1L (Disruptor of Telomeric Silencing 1-Like) is a histone methyltransferase that participates in gene expression regulation by catalyzing methylation on histone H3, particularly at the H3K79 site. DOT1L plays a crucial role in maintaining normal cell function, regulating gene expression, maintaining chromatin structure, repairing DNA damage, and controlling the cell cycle. However, in tumor cells, abnormal regulation and activity of DOT1L can lead to dysregulation of gene expression, thereby promoting tumor development and progression. Furthermore, abnormal expression or activity of DOT1L is also associated with epigenetic regulation in tumor cells, which may lead to drug resistance in tumors.

[0004] Due to its crucial role in various tumor types, DOT1L has become a potential target for cancer treatment. Developing inhibitors or antagonists targeting DOT1L could provide a novel treatment strategy. Research and development of treatments targeting DOT1L will offer new directions for cancer therapy.

[0005] In recent years, studies have discovered various pathological mutations within the functional active domain of the DOT1L protein, with the gain-of-function mutation R231Q being a prime example. The R231Q mutation refers to a mutation at amino acid position 231 in the DOT1L gene, where the native amino acid arginine (R) is replaced by glutamine (Q). This mutation has attracted widespread attention in tumor research. Literature confirms that the R231Q mutation can enhance the substrate binding ability of the DOT1L protein, strengthen its activity, and thus show a stronger effect in promoting tumor cell proliferation, self-renewal, invasion and metastasis, and the development of drug resistance. Compared with wild-type DOT1L protein (… WT Compared to the mutated DOT1L R231Q The protein has a more flexible and larger ligand-binding cavity, and the existing DOT1L WT Inhibitors against mutated DOT1L R231Q The proteins did not exhibit good inhibitory effects.

[0006] In addition, the existing DOT1L WT Most inhibitors belong to the adenosine class of compounds, which are organic compounds containing an adenosine or adenosine-like nucleus, such as 7-azapurine compounds and 2'-C-methyl substituted nucleosides. The technologies disclosed in invention patents WO2012075500, WO2012075381, CN109748943, and CN110092804 all belong to the adenosine class. Although adenosine compounds are specific, have low toxicity, typically exhibit bioactivity against multiple targets, can stimulate anti-tumor responses in the immune system, and possess excellent bioavailability, making them a highly promising compound, their structural characteristic—containing a metabolically unstable acetalamine fragment in their nucleus—leads to their easy recognition and rapid cleavage by various metabolic enzymes in the human body (Khirsariya P, (P, Maier L, et al., Journal of Medicinal Chemistry, 2022, 65: 5701-5723.). Therefore, adenosine compounds generally have the disadvantages of high clearance rates and poor pharmacokinetic properties, which greatly limits their application in the preparation of anti-tumor drugs. Therefore, there is a need to develop a non-adenosine inhibitor with good pharmacokinetic properties that can show good inhibitory activity against both wild-type and mutant DOT1L. Summary of the Invention

[0007] To address the problems existing in the background art, the present invention provides a non-adenosine DOT1L inhibitor with high metabolic stability, a 1,3-disubstituted piperidine derivative.

[0008] The purpose of this invention is to provide a 1,3-disubstituted piperidine derivative, the chemical structural formula of which is shown in general formula (I) or (II):

[0009]

[0010] In the formula:

[0011] m is independently 0-1;

[0012] n is independently 0-3;

[0013] X is

[0014] Het is a 5-9 membered heteroaryl group substituted with 0-4 independent R1 atoms, wherein the heteroaryl group contains 1-4 heteroatoms optionally selected from N, O and S;

[0015] R1 is a halogen, hydroxyl, amino, nitro, cyano, (C1-C6)alkyl, (C1-C6)alkoxy, (C1-C6)amide, (C1-C6)sulfonamide, halogen or hydroxyl or cyano or carboxyl substituted (C1-C6)alkyl or amino substituted with 1-2 (C1-C6)alkyl.

[0016] R2 is hydrogen, (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)ynyl, halogenated or amino-, cyano-, or carboxyl-substituted (C1-C6)alkyl, (CH2) p (C3-C6)cycloalkyl, (C1-C6)acyl, (C1-C6)sulfonyl, (CH2) p NR4R5, (CH2) p CONR4R5、(CH2) p COOR4 or (CH2) p OR4;

[0017] R4 and R5 may be the same or different, and are independently selected from hydrogen, (C1-C6) alkyl and halogen or amino or hydroxy or cyano-substituted (C1-C6) alkyl; or R4 and R5 together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or heteroaryl group, wherein the heterocyclic group or heteroaryl group contains 0-4 heteroatoms selected from N, O and S in addition to the nitrogen atom attached to R4 and R5;

[0018] R3 represents 1-4 substituents, each independently selected from hydrogen, halogen, hydroxyl, amino, nitro, cyano, (C1-C6)alkyl, (C1-C6)alkoxy, (C3-C6)cycloalkyl, (C1-C6)alkyl substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)alkoxy substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)acyl, (C1-C6)amide substituted with 0-3 halogens, hydroxyl groups, or amino groups, and (C1-C6)sulfonamide substituted with 0-3 halogens, hydroxyl groups, or amino groups.

[0019] p independently ranges from 0 to 6.

[0020] Preferably, in formula (I) or (II):

[0021] m is independently 0-1;

[0022] n is independently 0-3;

[0023] X is

[0024] Het is a 5-9 membered heteroaryl group substituted with 0-4 independent R1 atoms, wherein the heteroaryl group contains 1-4 heteroatoms optionally selected from N, O and S;

[0025] R1 is a halogen, hydroxyl, amino, nitro, cyano, (C1-C3)alkyl, (C1-C3)alkoxy, (C1-C3)amide, (C1-C3)sulfonamide, halogen or hydroxyl or cyano or carboxyl substituted (C1-C3)alkyl or amino substituted with 1-2 (C1-C3)alkyl groups.

[0026] R2 is hydrogen, (C1-C3)alkyl, (C2-C4)alkenyl, (C2-C4)alkynyl, halogenated or amino-, cyano-, or carboxyl-substituted (C1-C3)alkyl, (CH2) p (C3-C6)cycloalkyl, (C1-C3)acyl, (C1-C3)sulfonyl, (CH2) p NR4R5, (CH2) p CONR4R5、(CH2) p COOR4 or (CH2) p OR4;

[0027] R4 and R5 may be the same or different, and are independently selected from hydrogen, (C1-C3) alkyl and halogen or amino or hydroxy or cyano-substituted (C1-C3) alkyl; or R4 and R5 together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or heteroaryl group, wherein the heterocyclic group or heteroaryl group contains 0-4 heteroatoms selected from N, O and S in addition to the nitrogen atom attached to R4 and R5;

[0028] R3 represents 1-4 substituents, each independently selected from hydrogen, halogen, hydroxyl, amino, nitro, cyano, (C1-C6)alkyl, (C1-C6)alkoxy, (C3-C6)cycloalkyl, (C1-C6)alkyl substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)alkoxy substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)acyl, (C1-C6)amide substituted with 0-3 halogens, hydroxyl groups, or amino groups, and (C1-C6)sulfonamide substituted with 0-3 halogens, hydroxyl groups, or amino groups.

[0029] p independently ranges from 0 to 4.

[0030] More preferably, in the general formula (I) or (II):

[0031] m is independently 0-1;

[0032] n is independently 0-3;

[0033] X is

[0034] Het is a 5-9 membered heteroaryl group substituted with 0-4 independent R1 atoms, wherein the heteroaryl group contains 1-4 heteroatoms optionally selected from N, O and S;

[0035] R1 is a halogen, hydroxyl, amino, nitro, cyano, (C1-C3)alkyl, (C1-C3)alkoxy, (C1-C3)amide, (C1-C3)alkyl substituted with halogen or hydroxyl or cyano or carboxyl, or an amino substituted with 1-2 (C1-C3)alkyl groups.

[0036] R2 is hydrogen, (C1-C3)alkyl, halogen, or (C1-C3)alkyl substituted with amino, cyano, or carboxyl groups, or (CH2). p (C3-C6)cycloalkyl, (C1-C3)acyl, (C1-C3)sulfonyl, (CH2) p NR4R5, (CH2) p CONR4R5、(CH2) p COOR4 or (CH2) p OR4;

[0037] R4 and R5 may be the same or different, and are independently selected from hydrogen, (C1-C3) alkyl and halogen or amino or hydroxy or cyano-substituted (C1-C3) alkyl; or R4 and R5 together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or heteroaryl group, wherein the heterocyclic group or heteroaryl group contains 0-4 heteroatoms selected from N, O and S in addition to the nitrogen atom attached to R4 and R5;

[0038] R3 represents 1-4 substituents, each independently selected from hydrogen, halogen, hydroxyl, amino, nitro, cyano, (C1-C6)alkyl, (C1-C6)alkoxy, (C3-C6)cycloalkyl, (C1-C6)alkyl substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)alkoxy substituted with 0-3 halogens, hydroxyl groups, or amino groups, (C1-C6)acyl, (C1-C6)amide substituted with 0-3 halogens, hydroxyl groups, or amino groups, and (C1-C6)sulfonamide substituted with 0-3 halogens, hydroxyl groups, or amino groups.

[0039] p independently ranges from 0 to 4.

[0040] More preferably, in the general formula (I) or (II):

[0041] m is independently 0-1;

[0042] n is independently 0-2;

[0043] X is

[0044] Het is a 5-9 membered heteroaryl group substituted with 0-3 independent R1 atoms, wherein the heteroaryl group contains 1-4 heteroatoms optionally selected from N, O and S;

[0045] R1 is a halogen, hydroxyl, amino, nitro, cyano, (C1-C3)alkyl, or (C1-C3)alkoxy group;

[0046] R2 is a (C1-C3) alkyl, halogen, or amino, cyano, or carboxyl-substituted (C1-C3) alkyl, (CH2) p NR4R5, (CH2) p CONR4R5、(CH2) p COOR4 or (CH2) p OR4;

[0047] R4 and R5 may be the same or different, and are independently selected from hydrogen and (C1-C3) alkyl; or R4 and R5 together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or heteroaryl group, wherein the heterocyclic group or heteroaryl group contains 0-4 heteroatoms selected from N, O and S in addition to the nitrogen atom attached to R4 and R5.

[0048] R3 represents 1-4 substituents, each independently selected from hydrogen, halogen, (C1-C6)alkyl, (C1-C6)alkoxy, (C1-C6) amide with 0-3 halogen, hydroxyl or amino substituted groups, and (C1-C6) sulfonamide with 0-3 halogen, hydroxyl or amino substituted groups.

[0049] p independently ranges from 0 to 4.

[0050] More preferably, in the general formula (I) or (II):

[0051] m is independently 0-1;

[0052] n is independently 0-1;

[0053] X is

[0054] Het is a 5-9 membered heteroaryl group substituted with 0-2 R1 atoms, wherein the heteroaryl group contains 1-4 heteroatoms optionally selected from N, O and S;

[0055] R1 is a halogen, hydroxyl, amino, cyano, or (C1-C3) alkyl group;

[0056] R2 is a (C1-C3) alkyl group or (CH2) p NR4R5 or (CH2) p OR4;

[0057] R4 and R5 may be the same or different, and are independently selected from hydrogen and (C1-C3) alkyl; or R4 and R5 together with the nitrogen atom to which they are attached form a 6-membered heterocyclic group, wherein the heterocyclic group is morpholine or thiomorpholine;

[0058] R3 represents 1-4 substituents, each independently selected from hydrogen, halogen, (C1-C6)alkyl, (C1-C6)alkoxy and (C1-C6) amide with 0-3 halogen or hydroxyl or amino substituted groups.

[0059] p independently ranges from 0 to 4.

[0060] More preferably, in the general formula (I) or (II):

[0061] m is independently 0-1;

[0062] n is independently 0-1;

[0063] X is

[0064] Het is a 5-9 membered heteroaryl group substituted with 0-2 independent R1s, wherein the heteroaryl group is optionally 1H-pyrazolo[3,4-d]pyrimidine, imidazo[1,2-a]pyrazine, purine, pyridine, pyrazine or imidazole;

[0065] R1 is a halogen, hydroxyl, amino, cyano, or methyl group;

[0066] R2 is

[0067] R3 represents 1-3 substituents, each independently selected from hydrogen, halogen, methyl, tert-butyl, methoxy, acetamido, and pivaloylamino.

[0068] The 1,3-disubstituted piperidine derivative is specifically selected from the following compounds:

[0069] N-{2-{{4-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0070] N-{2-{{4-{(3-morpholinopropyl){[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0071] N-{2-{{4-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0072] N-{2-{{4-{{[1-(6-chloro-1H-pyrazolo[3,4-d]pyrimidin-4-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0073] N-{2-{{4-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0074] N-{2-{{4-{(3-morpholinopropyl){[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide;

[0075] N-{2-{{4-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazo-5-yl}neopentamide;

[0076] 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0077] 1-[4-(tert-butyl)phenyl]-3-{3-{methyl{[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea;

[0078] 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea;

[0079] 1-{3-{{[1-(1H-imidazol-1-carbonyl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0080] 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea;

[0081] 1-[4-(tert-butyl)phenyl]-3-{3-{methyl{[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea;

[0082] 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0083] 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea;

[0084] 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea;

[0085] 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea;

[0086] 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0087] 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea;

[0088] 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0089] 1-{3-{{[1-(8-bromo-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0090] 1-{3-{{[1-(2-amino-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0091] 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea;

[0092] 1-{3-{{[1-(2-amino-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea;

[0093] 1-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}-3-(3,5-dimethoxyphenyl)urea;

[0094] 1-(4-bromophenyl)-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}urea;

[0095] 1-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}-3-(3,4,5-trimethylphenyl)urea;

[0096] N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}ureo}phenyl}acetamide;

[0097] 1-{2-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}ethyl}-3-(3,5-dimethoxyphenyl)urea;

[0098] N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-methoxypropyl)amino}propyl}ureoyl}phenyl}acetamide.

[0099] The 1,3-disubstituted piperidine derivative of the invention forms a pharmaceutically acceptable salt with an acid, wherein the acid is: hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, naphthalenedisulfonic acid, acetic acid, propionic acid, lactic acid, trifluoroacetic acid, maleic acid, citric acid, fumaric acid, oxalic acid, tartaric acid, or benzoic acid.

[0100] This invention provides a method for preparing the 1,3-disubstituted piperidine derivative, wherein the preparation method is selected from one of method A and method B, and the preparation steps and route are shown below;

[0101] Method A:

[0102] Step 1: Intermediate M1 undergoes a reductive amination reaction with primary amines of different substitutions in the presence of a reducing agent to generate intermediate M2;

[0103] Step 2: Intermediate M3 and intermediate M4 of different lengths undergo a condensation reaction in the presence of carbonyl diimidazole (CDI) to generate intermediate M5;

[0104] Step 3: In the presence of an oxidizing agent, intermediate M5 oxidizes its terminal hydroxyl group to an aldehyde group to generate intermediate M6; or intermediate M5 reacts with an iodine reagent to generate intermediate M7.

[0105] Step 4: Intermediate M6 and intermediate M2 undergo a reductive amination reaction in the presence of a reducing agent to generate intermediate M8; or intermediate M6 and intermediate M2 undergo an N-alkylation reaction under alkaline conditions to generate intermediate M8;

[0106] Step 5: Intermediate M8 is deprotected by tert-butyloxycarbonyl (Boc) under acidic conditions to generate intermediate M9;

[0107] Step 6: Intermediate M9 undergoes a condensation reaction with heteroaryl carboxylic acids with different substitutions in the presence of a condensing agent to obtain compound I; or intermediate M9 undergoes an N-alkylation reaction with heteroaryl halides with different substitutions under alkaline conditions to obtain compound I.

[0108]

[0109] Method B:

[0110] Step 1: Intermediate M10 reacts with primary amines of different substitutions in the presence of a reducing agent to generate intermediate M11;

[0111] Step 2: Intermediate M11 undergoes a reductive amination reaction with intermediate M1 in the presence of a reducing agent to generate intermediate M12;

[0112] Step 3: Intermediate M12 is deprotected by tert-butyloxycarbonyl (Boc) under acidic conditions to generate intermediate M13;

[0113] Step 4: Intermediate M13 undergoes a condensation reaction with heteroaryl carboxylic acids with different substitutions in the presence of a condensing agent to obtain compound II; or intermediate M13 undergoes an N-alkylation reaction with heteroaryl halides with different substitutions under alkaline conditions to obtain compound II.

[0114]

[0115] On the other hand, the present invention provides the use of the aforementioned 1,3-disubstituted piperidine derivative or a pharmaceutically acceptable salt thereof in the preparation of a tumor-treating drug.

[0116] The present invention provides a pharmaceutical composition comprising a 1,3-disubstituted piperidine derivative thereof or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0117] The present invention also provides the use of the above-described pharmaceutical composition in the preparation of a drug for treating tumors.

[0118] Beneficial effects of the present invention

[0119] This invention provides a DOT1L inhibitor with a 1,3-disubstituted piperidine core, filling a gap in DOT1L inhibitor research. It possesses the advantages of novel structure, stability, and ease of storage. Furthermore, this invention provides a method for preparing the 1,3-disubstituted piperidine derivative, which is simple, easy to purify, and uses readily available raw materials. The reaction process is also environmentally friendly and suitable for large-scale industrial production, demonstrating good feasibility. Compared to existing DOT1L inhibitors, this 1,3-disubstituted piperidine derivative is a non-adenosine inhibitor, avoiding the use of adenosine in its molecular structure, fundamentally solving the pharmacokinetic problems caused by the metabolic instability of adenosine. It exhibits superior DOT1L inhibitory activity in both wild-type and mutant forms. Detailed Implementation

[0120] The present invention will be further described below with reference to specific implementation examples, but the present invention is not limited to these embodiments.

[0121] In the following examples, the proton NMR spectra of the compounds were measured using a Bruker ARX-400 / 600, and the mass spectra were measured using an Agilent 1100 LC / MSD; all reagents used were analytical grade or chemically pure.

[0122] Example 1: N-{2-{{4-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (1)

[0123]

[0124] The route is shown below:

[0125]

[0126] Step A1: Synthesis of intermediate A-I

[0127] At room temperature, 10.0 g (0.07 mol) of 4-nitro-o-phenylenediamine, 12.3 g (0.13 mol) of chloroacetic acid, and 50 mL of water were added to 133.0 mL of concentrated hydrochloric acid. The mixture was then heated to reflux and stirred for 4 h. The reaction solution was cooled to room temperature, the pH was adjusted to 9 with ammonia, and the mixture was filtered to give an orange-red solid. The crude product was recrystallized from methanol to give 8.9 g of a pale yellow solid, with a yield of 64.5%. Analytical data: ESI-MS [M+H] + (m / z):212.09.

[0128] Step A2: Synthesis of intermediate A-II

[0129] Intermediate A-I (9.0 g, 0.04 mol) was dissolved in tetrahydrofuran (100.0 mL) at room temperature, followed by the addition of 4-piperidinone hydrochloride hydrate (7.8 g, 0.08 mol) and anhydrous potassium carbonate (10.8 g, 0.08 mol). The reaction mixture was reacted at 50 °C for 6 h. Most of the tetrahydrofuran in the reaction solution was distilled off, and the residue was poured into water (100.0 mL). After stirring for 30 min, the mixture was filtered to give 8.0 g of a pale yellow solid, with a yield of 74.9%. Analytical data: ESI-MS [M+H] + (m / z):275.39.

[0130] Step A3: Synthesis of intermediate A-III

[0131] At room temperature, intermediate A-II (3.0 g, 0.011 mol) and reduced iron powder (3.1 g, 0.055 mol) were added to methanol (30.0 mL), and the mixture was heated to 65 °C and reacted for 30 min. Then, a solution of ammonium chloride (3.0 g, 0.057 mol) in water (15.0 mL) was added, and the reaction was continued with stirring for 1.5 h. The mixture was filtered while hot, washed with hot methanol, and the filtrate was evaporated to dryness. The residue was poured into water (30 mL), stirred, and the pH was adjusted to 9 with saturated sodium bicarbonate solution. The aqueous phase was extracted with a mixed solvent (DCM:MeOH = 8:1, 30.0 mL × 3). The combined organic phases were evaporated to dryness to give 1.8 g of a yellow solid, with a yield of 65.2%. Analytical data: ESI-MS [M+H] + (m / z):245.12.

[0132] Step A4: Synthesis of intermediate A-Ⅳ

[0133] At room temperature, intermediate A-III (3.8 g, 0.004 mol) and triethylamine (4.7 g, 0.012 mol) were added to isopropanol (38.0 mL), and pentanoyl chloride (3.7 g, 0.008 mol) was slowly added dropwise at 0 °C. After the addition was complete, the temperature was raised to 35 °C, and the reaction was stirred overnight. The reaction mixture was evaporated to dryness, and the residue was poured into water (40.0 mL) and extracted with ethyl acetate (30.0 mL × 3). The combined organic phases were evaporated to dryness to give 3.6 g of a yellow solid, yield 72.0%. Analytical data: ESI-MS [M+H] + (m / z):329.19.

[0134] Step A5: Synthesis of intermediate A-V

[0135] Intermediate A-Ⅳ (0.5 g, 1.52 mmol) and N-(3-aminopropyl)morpholine (0.35 g, 2.44 mmol) were dissolved in dichloromethane (5 mL). One drop of glacial acetic acid was added, and the mixture was stirred at room temperature for 0.5 h. Sodium triacetoxyborohydride (1.3 g, 6.09 mmol) was added, and the mixture was stirred at room temperature for 2 h. The reaction solution was quenched with a saturated sodium bicarbonate aqueous solution (15 mL), then extracted with dichloromethane (10 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to give 0.44 g of a yellow, foamy solid, with a yield of 63.3%. Analytical data: ESI-MS [M+H] + (m / z):457.32.

[0136] Step A6: Synthesis of intermediate A-VI

[0137] Methyl N-Boc-3-piperidincarboxylate (4 g, 15.5 mmol) was dissolved in tetrahydrofuran (60 mL). Lithium aluminum tetrahydrogenate (1.18 g, 31.1 mmol) was added in portions under ice bath conditions. After the addition was complete, the mixture was stirred at 0 °C for 1 h. The reaction solution was quenched with water (1.2 mL) and 15% sodium hydroxide aqueous solution (1.2 mL). After stirring at room temperature for 30 min, the mixture was filtered. The filtrate was evaporated to dryness to give 3.6 g of a pale yellow oil, with a yield of 100%. Analytical data: ESI-MS [M+H] + (m / z):216.15.

[0138] Step A7: Synthesis of intermediate A-VII

[0139] Intermediate A-VI (3.5 g, 16.2 mmol) was dissolved in dichloromethane (50 mL), and sodium bicarbonate (4 g, 48.7 mmol) and Desmartin oxidant (10.3 g, 24.3 mmol) were added. The mixture was stirred at room temperature for 1 h. The reaction solution was quenched with saturated sodium thiosulfate aqueous solution (50 mL), and the aqueous phase was extracted with dichloromethane (15 mL × 3). The combined organic phases were washed with saturated saline solution (30 mL), dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to give 2.4 g of a yellow oil, yield 69.1%. Analytical data: ESI-MS [M+H] + (m / z):214.14.

[0140] Step A8: Synthesis of intermediate A-VIII

[0141] Intermediates A-VII (94 mg, 0.44 mmol) and A-V (100 mg, 0.22 mmol) were dissolved in dichloromethane (1.5 mL), and 1 drop of glacial acetic acid was added. The mixture was stirred at room temperature for 0.5 h, followed by the addition of sodium triacetoxyborohydride (102 mg, 0.48 mmol) and stirring at room temperature for 16 h. The reaction mixture was quenched with saturated sodium bicarbonate aqueous solution (5 mL), extracted with dichloromethane (3 mL × 5), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography to give 130 mg of a colorless oil, with a yield of 80%. Analytical data: ESI-MS [M+H] + (m / z):654.46.

[0142] Step A9: Synthesis of intermediate A-IX

[0143] Trifluoroacetic acid (0.42 g, 2.8 mmol) was added to a solution of intermediate A-VIII (110 mg, 0.18 mmol) in dichloromethane (1.5 mL), and the mixture was stirred at room temperature for 2 h. After the reaction was complete, the reaction solution was evaporated to dryness to give 150 mg of a yellow oil, with a yield of 100%. Analytical data: ESI-MS [M+H] + (m / z):554.41.

[0144] Step A10: Synthesis of Example 1

[0145] Intermediate A-IX (100 mg, 0.18 mmol) and N,N-diisopropylethylamine (117 mg, 0.9 mmol) were dissolved in isopropanol (2 mL), and 6-chloropurine (84 mg, 0.54 mmol) was added. The mixture was stirred at 80 °C for 48 h. The reaction solution was evaporated to dryness, and the residue was diluted with dichloromethane (5 mL), washed with saturated ammonium chloride aqueous solution (2 mL × 3), and dried over anhydrous sodium sulfate. The organic phase was evaporated to dryness to obtain the crude product, which was purified by column chromatography to give 53 mg of a pale yellow solid, with a yield of 43.8%. Analytical data: mp: 201.3–205.6 °C; ESI-MS [M+H] + (m / z): 672.40; 1 H NMR(400MHz,DMSO-d6)δ12.95(s,1H),12.15(s,1H),9.14(s,1H),8.16(s,1H ),8.07(s,1H),7.88(s,1H),7.56-7.16(m,2H),3.65(s,2H),3.46(s,6H),3.1 8(s,2H),2.97-2.84(m,2H),2.79(s,1H),2.44(q,J=7.0Hz,3H),2.36-2.17( m,7H),2.11-1.94(m,2H),1.90-1.78(m,1H),1.75-1.41(m,8H),1.24(s,9H); 13 C NMR(101MHz,DMSO-d6)δ176.61,153.70,152.28,151.82,138.12,119.16,66.57,57.68,56 .31,54.35,53.84,53.76,48.46,39.49,35.69,29.70,28.23,27.81,27.02,25.61,25.34.

[0146] Example 2: N-{2-{{4-{(3-morpholinopropyl){[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (2)

[0147]

[0148] Step A11: Synthesis of Example 2

[0149] At room temperature, pyrazine-2-carboxylic acid (34 mg, 0.27 mmol), N,N-diisopropylethylamine (47 mg, 0.36 mmol), and 2-(7-benzotriazole oxide)-N,N,N',N'-tetramethylurea hexafluorophosphate (103 mg, 0.27 mmol) were added to dichloromethane (2 mL), and the mixture was stirred at room temperature for 1 h. Then, intermediate A-IX (100 mg, 0.18 mmol) was added, and the reaction was continued for 23 h. After the reaction was complete, water (10 mL) was added to the reaction solution, and the mixture was extracted with dichloromethane (10.0 mL × 3). The combined organic phases were evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography to give 70 mg of a pale yellow solid, with a yield of 58.8%. Analytical data: mp: 186.9-189.7℃; ESI-MS [M+H] + (m / z): 660.42; 1 H NMR (400MHz, DMSO-d6) δ12.14(s,1H),9.14(s,1H),8.84-8.77(m,1H),8.72(dd,J=4.6,2.6H z,1H),8.64(ddd,J=14.6,2.6,1.5Hz,1H),7.89(s,1H),7.49-7.22(m,2H),4.46(dd,J=39.2 ,12.0Hz,1H),3.84-3.47(m,8H),3.02(t,J=12.0Hz,1H),2.95-2.61(m,3H),2.49-2.42(m,1 H), 2.38-2.08 (m, 7H), 2.02 (ddd, J=19.1, 9.0, 3.7Hz, 3H), 1.82-1.37 (m, 7H), 1.24 (s, 14H).

[0150] Example 3: N-{2-{{4-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (3)

[0151]

[0152] Following the method in step A10 of Example 1, using intermediate A-IX as a starting material, an N-alkylation reaction was carried out with 2,6-dichloropurine to prepare Example 3, with a yield of 42.5%. Analytical data: mp: 203.7-207.1℃; ESI-MS [M+H] + (m / z): 706.40; 1HNMR (400MHz, DMSO-d6) δ13.08(s,1H),12.13(s,1H),9.13(s,1H),8.08(d,J=8.3Hz,1H),7.89(s,1H),7.34(d,J=42.2Hz,2H),5.59(s,1H),4.85( s,1H),3.65(s,2H),3.57-3.38(m,6H),2.88(d,J=10.7Hz,4H),2.48-2.1 0(m,9H),2.00(dd,J=18.4,7.9Hz,2H),1.87-1.35(m,9H),1.24(s,11H). 13 C NMR(101MHz,DMSO-d6)δ176.59,153.76,152.99,138.76,129.73,118.09,66.64,57.58,56 .37,56.26,54.33,53.91,53.80,48.29,39.49,29.45,27.82,26.68,25.82,22.56,14.41.

[0153] Example 4: N-{2-{{4-{{[1-(6-chloro-1H-pyrazolo[3,4-d]pyrimidin-4-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (4)

[0154]

[0155] Following the method in step A10 of Example 1, using intermediate A-IX as a starting material, Example 4 was prepared by N-alkylation reaction with 4,6-dichloro-1H-pyrazolo[3,4-d]pyrimidine, with a yield of 66.7%. Analytical data: mp: 208.4-210.6℃; ESI-MS [M+H] + (m / z): 706.40; 1H NMR(400MHz,DMSO-d6)δ13.66(s,1H),12.12(s,1H),9.11(s,1H),8.18(s,1H),7.89(s ,1H),7.33(d,J=42.6Hz,2H),4.76(d,J=86.0Hz,1H),4.39(s,1H),3.65(s,2H),3.46( t,J=4.7Hz,4H),3.07(s,1H),2.88(d,J=10.7Hz,2H),2.43(tq,J=13.0,6.2Hz,3H),2. 26(d,J=24.8Hz,8H),2.04(dt,J=23.3,11.9Hz,2H),1.80-1.38(m,10H),1.23(s,11H). 13 C NMR(101MHz,DMSO-d6)δ176.05,156.82,156.61,156.07,133.75,98.10,66.11,57 .29,55.92,55.79,53.78,53.29,53.12,38.98,28.49,27.31,25.90,25.40,24.52.

[0156] Example 5: N-{2-{{4-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (5)

[0157]

[0158] Following the method in step A10 of Example 1, using intermediate A-IX as a starting material, an N-alkylation reaction was carried out with 2-chloro-4-cyanopyridine to prepare Example 5, with a yield of 55.9%. Analytical data: mp: 199.9-203.4℃; ESI-MS [M+H] + (m / z): 656.42; 1H NMR (400MHz, DMSO-d6) δ9.18(s,1H),8.23(d,J=5.0Hz,1H),7.93(d,J=1.9Hz,1H),7.42(d ,J=8.6Hz,1H),7.32(dd,J=8.7,1.9Hz,1H),7.23(s,1H),6.85(dd,J=5.0,1.2Hz,1H),4.24 (dd,J=54.1,12.9Hz,2H),3.84(s,2H),3.63(dq,J=10.4,6.5,5.1Hz,6H),3.19-2.88(m,5 H), 2.65 (d, J = 21.1Hz, 7H), 2.35 (d, J = 62.0Hz, 3H), 1.85-1.57 (m, 7H), 1.35-1.24 (m, 14H).

[0159] Example 6: N-{2-{{4-{(3-morpholinopropyl){[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide (6)

[0160]

[0161] Following the method in step A11 of Example 2, using intermediate A-IX as a starting material, an N-acylation reaction was carried out with 2-oxo-1,2-dihydropyridine-3-carboxylic acid to prepare Example 6, with a yield of 82.4%. Analytical data: mp: 198.5-202.3℃; ESI-MS [M+H] + (m / z): 675.43; 1 H NMR (400MHz, DMSO-d6) δ11.84(s,2H),9.14(s,1H),7.90(d,J=1.9Hz,1H),7.40(dt,J=9.3,3 .7Hz,3H),7.29(dd,J=8.5,1.9Hz,1H),6.20(dt,J=22.8,6.6Hz,1H),4.35(d,J=13.5Hz,1H) ,3.82-3.68(m,2H),3.59(dd,J=9.5,4.5Hz,6H),2.88(d,J=32.3Hz,3H),2.62(dd,J=24.4,1 2.4Hz, 2H), 2.29 (d, J = 53.7Hz, 7H), 2.05 (d, J = 14.1Hz, 3H), 1.75-1.45 (m, 7H), 1.24 (s, 14H).

[0162] Example 7: N-{2-{{4-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazo-5-yl}neopentamide (7)

[0163]

[0164] Following the method in step A10 of Example 1, using intermediate A-IX as a starting material, an N-alkylation reaction was carried out with 3-bromo-8-chloro-imidazolium[1,2-a]pyrazine to prepare Example 7, with a yield of 44.8%. Analytical data: mp: 204.2-208.1℃; ESI-MS [M+H] + (m / z): 749.35; 1 H NMR (400MHz, DMSO-d6) δ12.15(s,1H),9.14(s,1H),7.90(s,1H),7.68(s,1H),7.61(d,J=4.5Hz,1H ),7.45(d,J=4.5Hz,1H),7.42-7.19(m,2H),5.43-5.04(m,2H),3.66(s,2H),3.47(t,J=4.9Hz,4H), 3.14(t,J=11.6Hz,1H),2.99-2.67(m,3H),2.48-2.39(m,2H),2.33-2.15(m,7H),2.02(q,J=11.4, 10.6Hz, 2H), 1.87-1.76 (m, 1H), 1.75-1.54 (m, 4H), 1.46 (p, J = 8.9, 8.4Hz, 4H), 1.29-1.12 (m, 13H).

[0165] Example 8: 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (8)

[0166]

[0167] The route is shown below:

[0168]

[0169] Step B1: Synthesis of intermediate B-I

[0170] At room temperature, carbonyl diimidazole (7.06 g, 43.55 mmol) was dissolved in tetrahydrofuran (75 mL), and 4-tert-butylaniline (5 g, 33.5 mmol) was added and the mixture was stirred for 6 h. Then, 3-aminopropanol (3.02 g, 40.2 mmol) was added, and the reaction was continued for 1.5 h. The reaction mixture was evaporated to dryness, and the residue was dissolved in ethyl acetate (300 mL). The organic phase was then washed with 1M hydrochloric acid aqueous solution (100 mL × 3), saturated sodium bicarbonate aqueous solution (100 mL), and saturated brine (100 mL), respectively. After drying with anhydrous sodium sulfate and filtration, the organic phase was evaporated to dryness to give 8 g of a yellow oily substance, with a yield of 95.3%. Analytical data: ESI-MS [M+H] + (m / z):251.17.

[0171] Step B2: Synthesis of intermediate B-II

[0172] Intermediate B-I (2 g, 8 mmol) was dissolved in dry dichloromethane (80 mL). Under nitrogen protection, the mixture was cooled to -10 °C, and Dysmartin oxidant (5.09 g, 12 mmol) was slowly added in portions. After the addition was complete, the mixture was heated to 0 °C and reacted for 1.5 h. The reaction solution was quenched with saturated sodium thiosulfate solution (20 mL) and saturated sodium bicarbonate solution (20 mL). After stirring at room temperature for 15 min, the mixture was filtered, and the filtrate was separated. The separated aqueous phase was extracted with dichloromethane (20 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to obtain the crude product. The crude product was recrystallized from ethyl acetate to give 0.9 g of a white solid, with a yield of 45%. Analytical data: ESI-MS [M+H] + (m / z):249.15.

[0173] Step B3: Synthesis of intermediate B-III

[0174] Intermediate A-VII (2 g, 9.38 mmol) and 40% methylamine aqueous solution (1.1 g, 14.1 mmol) were added to methanol (20 mL), followed by the addition of 1 drop of glacial acetic acid and reaction at room temperature for 1 h. Sodium cyanoborohydride (1.2 g, 18.8 mmol) was then added, and the reaction continued for 16 h. The reaction solution was quenched with saturated sodium bicarbonate aqueous solution (40 mL), followed by extraction with dichloromethane (20 mL × 3). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and evaporated to dryness to give 0.86 g of a yellow oil, yield 40.2%. Analytical data: ESI-MS [M+H] + (m / z):229.18.

[0175] Step B4: Synthesis of intermediate B-Ⅳ

[0176] Intermediate B-II (0.78 g, 3.15 mmol) and intermediate B-III (0.6 g, 2.63 mmol) were dissolved in dichloromethane (6 mL), and 1 drop of glacial acetic acid was added. The mixture was stirred at room temperature for 1 h, followed by the addition of sodium triacetoxyborohydride (0.72 g, 3.42 mmol), and the reaction was continued for 16 h. The reaction solution was quenched with saturated sodium bicarbonate aqueous solution (10 mL), and then extracted with dichloromethane (10 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate and evaporated to dryness to give 0.8 g of a colorless oil, yield 59%. Analytical data: ESI-MS [M+H] + (m / z):461.40.

[0177] Step B5: Synthesis of intermediate B-V

[0178] Trifluoroacetic acid (3.09 g, 27 mmol) was added to a solution of intermediate B-Ⅳ (0.8 g, 1.74 mmol) in dichloromethane (1.5 mL), and the mixture was stirred at room temperature for 2 h. After the reaction was complete, the reaction solution was evaporated to dryness to give 0.76 g of a white solid, with a yield of 100%. Analytical data: ESI-MS [M+H] + (m / z):361.30.

[0179] Step B6: Synthesis of Example 8

[0180] Intermediate B-V (80 mg, 0.18 mmol) and N,N-diisopropylethylamine (117 mg, 0.9 mmol) were dissolved in isopropanol (2 mL), and 6-chloropurine (102 mg, 0.54 mmol) was added. The mixture was stirred at 80 °C for 48 h. The reaction solution was evaporated to dryness, and the residue was diluted with dichloromethane (5 mL), washed with saturated ammonium chloride aqueous solution (2 mL × 3), and dried over anhydrous sodium sulfate. The organic phase was evaporated to dryness to obtain the crude product, which was purified by column chromatography to give 43 mg of a pale yellow solid, with a yield of 49.9%. Analytical data: mp: 183.6–185.8 °C; ESI-MS [M+H] + (m / z): 479.30; 1 H NMR (400MHz, DMSO-d6) δ12.98(s,1H),8.54(s,1H),8.21(s,1H),8.14(s,1H),7.33-7.16(m,4H),6.32(t,J=5.7Hz,1H),5.15(s,2 H),3.22-2.88(m,4H),2.67(s,3H),2.50-2.31(m,3H),1.96-1.81(m,2H),1.73(d,J=12.7Hz,3H),1.57-1.27(m,3H),1.23(s,9H).

[0181] Example 9: 1-[4-(tert-butyl)phenyl]-3-{3-{methyl{[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea (9)

[0182]

[0183] Following the method in step A11 of Example 2, using intermediate B-V as a starting material, N-acylation reaction was carried out with pyrazine-2-carboxylic acid to prepare Example 9, with a yield of 59.5%. Analytical data: mp: 185.1-189.1℃; ESI-MS [M+H] + (m / z): 467.31; 1 HNMR (400MHz, DMSO-d6) δ8.82(t,J=1.8Hz,1H),8.72(dd,J=9.8,2.6Hz,1H),8.67(q,J=2.1Hz,1H),8.46(s,1H),7.34-7.15(m,4H),6.20(d,J=38. 6Hz,1H),4.37(dd,J=49.0,12.9Hz,1H),3.74-3.54(m,1H),3.21-2.57(m ,5H),2.25(d,J=69.2Hz,5H),1.93-1.42(m,6H),1.24(d,J=1.9Hz,11H).

[0184] Example 10: 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea (10)

[0185]

[0186] Following the method in step B6 of Example 8, intermediate B-V was used as a starting material and reacted with 2,6-dichloropurine via an N-alkylation reaction to produce Example 10, with a yield of 30.4%. Analytical data: mp: 187.4-190.7℃; ESI-MS [M+H] + (m / z): 513.30; 1 HNMR(400MHz,DMSO-d6)δ13.21(s,1H),8.54(s,1H),8.16(s,1H),7.34-7.17(m,4H),6.31(s,1H),5.47(s,1H),4.66(s,1H), 3.15(q,J=7.4,6.9Hz,4H),2.47(s,3H),2.02-1.81(m,2H),1.81-1.61(m,3H),1.42(dd,J=65.1,11.9Hz,2H),1.23(s,10H).

[0187] Example 11: 1-{3-{{[1-(1H-imidazol-1-carbonyl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (11)

[0188]

[0189] Step B7: Synthesis of Example 11

[0190] Carbonyl diimidazole (117 mg, 0.72 mmol) was dissolved in tetrahydrofuran (2 mL), and intermediate B-V (80 mg, 0.18 mmol) was slowly added in portions at room temperature, with the mixture stirred for 24 h. The reaction solution was evaporated to dryness to obtain the crude product, which was purified by column chromatography to give 28 mg of a white solid, with a yield of 34.1%. Analytical data: mp: 179.6-182.6℃; ESI-MS [M+H] + (m / z):455.30.

[0191] Example 12: 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea (12)

[0192]

[0193] Following the method in step B6 of Example 8, using intermediate B-V as a starting material, an N-alkylation reaction was carried out with 2-chloro-4-cyanopyridine to prepare Example 12, with a yield of 33.7%. Analytical data: mp: 188.9-191.4℃; ESI-MS [M+H] + (m / z): 463.20; 1 H NMR (400MHz, DMSO-d6) δ8.55 (s, 1H), 8.26 (d, J = 5.1Hz, 1H), 7.39-7.14 (m, 4H),6.91-6.81(m,1H),6.32(s,1H),4.48-4.29(m,1H),4.16(d,J=13.1Hz, 1H),3.13(q,J=6.5Hz,2H),2.94(ddd,J=13.9,11.2,2.9Hz,1H),2.83-2.5 1(m,4H),2.42(s,3H),2.04-1.56(m,5H),1.53-1.40(m,1H),1.24(s,11H).

[0194] Example 13: 1-[4-(tert-butyl)phenyl]-3-{3-{methyl{[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea (13)

[0195]

[0196] Following the method in step A11 of Example 2, using intermediate B-V as a starting material, an N-acylation reaction was carried out with 2-oxo-1,2-dihydropyridine-3-carboxylic acid to prepare Example 13, with a yield of 26.5%. Analytical data: mp: 187.7-189.8℃; ESI-MS [M+H] + (m / z): 482.30; 1 H NMR (400MHz, DMSO-d6) δ11.90 (s, 1H), 8.35 (d, J = 10.8Hz, 1H), 7.50-7.39 (m, 2H), 7.32 -7.25(m,2H),7.21(t,J=8.7Hz,2H),6.23(td,J=6.6,4.3Hz,1H),6.13(dt,J=39.3,5. 8Hz,1H),4.37-4.17(m,1H),3.19-2.91(m,3H),2.90-2.52(m,2H),2.47-2.28(m,2H), 2.13(d,J=50.5Hz,4H),1.79-1.53(m,4H),1.49-1.36(m,2H),1.24(d,J=1.9Hz,11H).

[0197] Example 14: 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (14)

[0198]

[0199] Following the method in step B6 of Example 8, using intermediate B-V as a starting material, Example 14 was prepared by N-alkylation reaction with 3-bromo-8-chloroimidazole[1,2-a]pyrazine, with a yield of 59.9%. Analytical data: mp: 193.1-195.2℃; ESI-MS [M+H] + (m / z): 556.20; 1H NMR (400MHz, DMSO-d6) δ8.34(s,1H),7.71(s,1H),7.63(d,J=4.5Hz,1H),7.49(d,J=4. 5Hz,1H),7.28(d,J=8.8Hz,2H),7.21(d,J=8.8Hz,2H),6.12(t,J=5.7Hz,1H),5.22(t, J=17.1Hz,2H),3.21-3.06(m,3H),2.84(t,J=11.6Hz,1H),2.49-2.25(m,3H),2.20(s, 4H), 1.89-1.76 (m, 2H), 1.72 (dt, J = 13.3, 3.5Hz, 1H), 1.66-1.45 (m, 3H), 1.23 (s, 10H).

[0200] Example 15: 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea (15)

[0201]

[0202] The route is shown below:

[0203]

[0204] Step B8: Synthesis of intermediate B-VI

[0205] Following the method in step B3 of Example 8, intermediate B-VI was prepared by reductive amination of intermediate A-VII with isopropylamine, with a yield of 67.6%. Analytical data: ESI-MS [M+H] + (m / z):257.22.

[0206] Step B9: Synthesis of intermediate B-VII

[0207] Following the method in step B4 of Example 8, intermediate B-VI was prepared by reductive amination with intermediate B-II to obtain intermediate B-VII, with a yield of 77.6%. Analytical data: ESI-MS [M+H] + (m / z):489.37.

[0208] Step B10: Synthesis of intermediate B-VIII

[0209] Following the method in step B5 of Example 8, intermediate B-VIII was prepared from intermediate B-VII via a deprotection reaction, with a yield of 100%. Analytical data: ESI-MS [M+H] + (m / z):389.40.

[0210] Step B11: Synthesis of Example 15

[0211] Following the method in step B6 of Example 8, intermediate B-VIII was used as a starting material and reacted with 2-chloro-4-cyanopyridine via an N-alkylation reaction to prepare Example 15, with a yield of 59.9%. Analytical data: mp: 195.9-199.3℃; ESI-MS [M+H] + (m / z): 491.30; 1 H NMR (400MHz, DMSO-d6) δ8.32 (s, 1H), 8.25 (d, J = 5.0Hz, 1H), 7.30-7.24 (m, 2H), 7.24-7.16 (m, 3H),6.83(d,J=5.0Hz,1H),6.07(s,1H),4.32(d,J=13.0Hz,1H),4.21-4.11(m,1H),3.11(qd, J=6.6,3.0Hz,2H),3.02-2.78(m,2H),2.61(t,J=11.6Hz,1H),2.34(d,J=52.4Hz,3H),1.88-1 .78(m,1H),1.72-1.49(m,4H),1.45-1.37(m,1H),1.24(s,11H),0.96(dd,J=11.8,6.4Hz,6H).

[0212] Example 16: 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea (16)

[0213]

[0214] Following the method in step A11 of Example 2, using intermediate B-VIII as a starting material, an N-acylation reaction was carried out with 2-oxo-1,2-dihydropyridine-3-carboxylic acid to prepare Example 16, with a yield of 34.9%. Analytical data: mp: 194.2-197.9℃; ESI-MS [M+H] + (m / z): 510.20; 1H NMR (400MHz, DMSO-d6) δ11.89 (s, 1H), 8.29 (d, J = 12.4Hz, 1H), 7.50-7.38 (m, 2H), 7.35-7.26 ( m,2H),7.21(dd,J=8.9,7.0Hz,2H),6.21(dt,J=8.6,6.6Hz,1H),6.13-5.91(m,1H),4.39(d,J =12.7Hz,1H),3.21-2.83(m,3H),2.82-2.52(m,2H),2.44-1.94(m,4H),1.83-1.47(m,4H),1. 24(d,J=1.5Hz,12H),1.17-1.06(m,1H),0.93(dd,J=19.3,6.4Hz,3H),0.79(d,J=6.5Hz,3H).

[0215] Example 17: 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea (17)

[0216]

[0217] Following the method in step A11 of Example 2, using intermediate B-VIII as a starting material, an N-acylation reaction was carried out with pyrazine-2-carboxylic acid to prepare Example 17, with a yield of 75.3%. Analytical data: mp: 196.7-198.8℃; ESI-MS [M+H] + (m / z):495.40.

[0218] Example 18: 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (18)

[0219]

[0220] Following the method in step B6 of Example 8, intermediate B-VIII was used as a starting material and reacted with 6-chloropurine via an N-alkylation reaction to produce Example 18, with a yield of 46.1%. Analytical data: mp: 198.5-202.1℃; ESI-MS [M+H] + (m / z): 507.40; 1HNMR(400MHz,DMSO-d6)δ12.96(s,1H),8.28(s,1H),8.19(s,1H),8.09(s,1H),7.27 (d,J=8.8Hz,2H),7.20(d,J=8.8Hz,2H),6.04(t,J=5.6Hz,1H),3.13(qd,J=6.8,2.3 Hz,3H),2.86(d,J=59.8Hz,2H),2.32(d,J=52.5Hz,4H),1.86(dd,J=13.5,3.7Hz,1H ),1.78-1.62(m,2H),1.61-1.44(m,3H),1.23(s,10H),0.95(dd,J=19.5,6.5Hz,6H).

[0221] Example 19: 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea (19)

[0222]

[0223] Following the method in step B6 of Example 8, intermediate B-VIII was used as a starting material and reacted with 2,6-dichloropurine via an N-alkylation reaction to produce Example 19, with a yield of 33.9%. Analytical data: mp: 199.8-203.7℃; ESI-MS [M+H] + (m / z): 541.30; 1 HNMR(400MHz,DMSO-d6)δ13.13(s,1H),8.32(s,1H),8.12(s,1H),7.26(d, J=8.9Hz,2H),7.20(d,J=8.8Hz,2H),6.05(s,1H),5.62(s,1H),4.86(s,1H) ,3.21-2.90(m,4H),2.35(d,J=43.2Hz,4H),1.84(d,J=12.8Hz,1H),1.80-1 .64(m,2H),1.62-1.42(m,3H),1.23(s,11H),0.97(dd,J=27.4,6.5Hz,6H).

[0224] Example 20: 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (20)

[0225]

[0226] Following the method in step B6 of Example 8, using intermediate B-VIII as a starting material, Example 20 was prepared by N-alkylation reaction with 3-bromo-8-chloroimidazole[1,2-a]pyrazine, with a yield of 46.6%. Analytical data: mp: 197.4-200.6℃; ESI-MS [M+H] + (m / z): 584.30; 1 H NMR (400MHz, DMSO-d6) δ8.30(s,1H),7.70(s,1H),7.62(d,J=4.5Hz,1H),7.48(d,J=4.5Hz,1 H),7.27(d,J=8.8Hz,2H),7.20(d,J=8.8Hz,2H),6.05(t,J=5.5Hz,1H),5.38-5.16(m,2H),3 .19-3.07(m,3H),2.95(s,1H),2.79(t,J=11.3Hz,1H),2.34(d,J=55.9Hz,4H),1.84(d,J=12 .3Hz,1H),1.79-1.66(m,2H),1.62-1.46(m,3H),1.23(s,10H),0.95(dd,J=18.9,6.5Hz,6H).

[0227] Example 21: 1-{3-{{[1-(8-bromo-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (21)

[0228]

[0229] Following the method in step B6 of Example 8, using intermediate B-VIII as a starting material, Example 21 was prepared by N-alkylation reaction with 8-bromo-6-chloro-9H-purine, with a yield of 28.6%. Analytical data: mp: 201.0-202.7℃; ESI-MS [M+H] + (m / z):585.20.

[0230] Example 22: 1-{3-{{[1-(2-amino-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (22)

[0231]

[0232] Following the method in step B6 of Example 8, intermediate B-VIII was used as a starting material and reacted with 2-amino-6-chloropurine via an N-alkylation reaction to prepare Example 22, with a yield of 31.4%. Analytical data: mp: 200.6-202.1℃; ESI-MS [M+H]+ (m / z):522.40.

[0233] Example 23: 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea (23)

[0234]

[0235] Following the method in step B6 of Example 8, using intermediate B-VIII as a starting material, Example 23 was prepared by N-alkylation reaction with 2,6-dichloro-9-methyl-9H-purine, with a yield of 45.1%. Analytical data: mp: 202.8-204.4℃; ESI-MS [M+H] + (m / z):555.30.

[0236] Example 24: 1-{3-{{[1-(2-amino-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea (24)

[0237]

[0238] Following the method in step B6 of Example 8, using intermediate B-VIII as a starting material, Example 24 was prepared by N-alkylation reaction with 6-chloro-9-methyl-9H-purine-2-amine, with a yield of 42.0%. Analytical data: mp: 203.1-205.1℃; ESI-MS [M+H] + (m / z):536.40.

[0239] Example 25: 1-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}-3-(3,5-dimethoxyphenyl)urea (25)

[0240]

[0241] The route is shown below:

[0242]

[0243] Step C1: Synthesis of intermediate C-I

[0244] Following the method in step B1 of Example 8, intermediate C-I was prepared by condensation reaction of 3,5-dimethoxyaniline with 3-aminopropanol, with a yield of 82.7%. Analytical data: ESI-MS [M+H] + (m / z):255.20.

[0245] Step C2: Synthesis of intermediate C-II

[0246] Triphenylphosphine (0.6 g, 2.29 mmol) and imidazole (0.15 g, 2.29 mmol) were dissolved in dichloromethane (4 mL). Elemental iodine (0.6 g, 2.29 mmol) was added at 0 °C, and the mixture was stirred for 5 min. Then, a dichloromethane solution (4 mL) of intermediate C-I (0.39 g, 1.52 mmol) was added, and the mixture was stirred at room temperature for 3 h. The reaction solution was diluted with dichloromethane (20 mL) and washed with saturated ammonium chloride solution (10 mL), saturated sodium bicarbonate solution (10 mL), and saturated brine solution (10 mL), respectively. The solution was then dried over anhydrous sodium sulfate, filtered, and evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography to obtain 0.4 g of a yellow solid, with a yield of 72%. Analytical data: ESI-MS [M+H] + (m / z):365.10.

[0247] Step C3: Synthesis of intermediate C-Ⅲ

[0248] Following the method in step B3 of Example 8, intermediate C-III was prepared by reductive amination of intermediate A-VII with N-(3-aminopropyl)morpholine, with a yield of 59.1%. Analytical data: ESI-MS [M+H] + (m / z):342.30.

[0249] Step C4: Synthesis of intermediate C-Ⅳ

[0250] Potassium carbonate (0.46 g, 3.30 mmol), intermediate C-II (0.4 g, 1.10 mmol), and intermediate C-III (0.45 g, 1.32 mmol) were added to acetonitrile (5 mL) and reacted at 60 °C for 24 h. Inorganic salts were removed from the reaction solution by filtration, and the filtrate was evaporated to dryness to obtain the crude product. The crude product was purified by column chromatography to give 0.37 g of a pale yellow solid, with a yield of 58%. Analytical data: ESI-MS [M+H] + (m / z):378.40.

[0251] Step C5: Synthesis of intermediate C-V

[0252] Following the method in step B5 of Example 8, intermediate C-V was obtained by deprotection of intermediate C-Ⅳ under trifluoroacetic acid conditions, with a yield of 100%. Analytical data: ESI-MS [M+H] + (m / z):478.30.

[0253] Step C6: Synthesis of Example 25

[0254] Following the method in step B6 of Example 8, intermediate C-V was used as a starting material and reacted with 2,6-dichloropurine via an N-alkylation reaction to prepare Example 25, with a yield of 25.7%. Analytical data: mp: 226.1-227.8℃; ESI-MS [M+H] + (m / z): 630.32; 1 HNMR (400MHz, DMSO-d6) δ13.17(s,1H),8.63(s,1H),8.13(s,1H),6.61(d,J=2.3Hz,2H),6.28(s,1H),6.04(t,J=2.3Hz,1H),5.56(s,1H),4.76(s,1 H),3.67(s,6H),3.54(t,J=4.7Hz,6H),3.23-2.95(m,4H),2.60(dd,J=27 .4,11.6Hz,4H),2.44-2.22(m,3H),1.92-1.54(m,8H),1.50-1.16(m,4H).

[0255] Example 26: 1-(4-bromophenyl)-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}urea (26)

[0256]

[0257] Following steps C1-C6 of Example 25, 4-bromoaniline was used as the starting material, and Example 26 was prepared through a 6-step reaction. Analytical data: mp: 229.2-231.4℃; ESI-MS [M+H] + (m / z):648.21.

[0258] Example 27: 1-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}-3-(3,4,5-trimethylphenyl)urea (27)

[0259]

[0260] Following steps C1-C6 of Example 25, 3,4,5-trimethylaniline was used as the starting material, and Example 27 was prepared through a 6-step reaction. Analytical data: mp: 233.6-234.8℃; ESI-MS [M+H] + (m / z):612.35.

[0261] Example 28: N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}ureo}phenyl}acetamide (28)

[0262]

[0263] Following steps C1-C6 of Example 25, 4-aminoacetanilide was used as a starting material, and Example 28 was prepared through a 6-step reaction. Analytical data: mp: 234.7-236.2℃; ESI-MS [M+H] + (m / z):627.30.

[0264] Example 29: 1-{2-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}ethyl}-3-(3,5-dimethoxyphenyl)urea (29)

[0265]

[0266] Following steps C1-C6 of Example 25, 3,5-dimethoxyaniline and 2-hydroxyethylamine were used as starting materials, and Example 29 was prepared through a 6-step reaction. Analytical data: mp: 230.2-231.9℃; ESI-MS [M+H] + (m / z):616.30.

[0267] Example 30: N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-methoxypropyl)amino}propyl}ureido}phenyl}acetamide (30)

[0268]

[0269] Following steps C1-C6 of Example 25, 3-methoxypropylamine was used as a starting material, and Example 30 was prepared through a 6-step reaction. Analytical data: mp: 228.5-229.7℃; ESI-MS [M+H] + (m / z):572.20.

[0270] Example 31: Antitumor activity experiment on the compounds prepared in the above examples.

[0271] I. In vitro antitumor cell activity

[0272] The compounds from Examples 1-30 of this invention were tested in vitro to inhibit the growth of human lung cancer cell line H460-DOT1L. WT H460-DOT1L R231Q The impact on cell survival.

[0273] 1. Cell Culture

[0274] Using RPMI 1640 medium as the basal culture medium, a cell culture medium containing 10% fetal bovine serum was prepared. Cells were cultured in a 37°C incubator containing 5% CO2. The culture medium was changed daily. When the cells reached 80%-90% confluence, the original culture medium was discarded, the cells were washed once with PBS, digested with 0.25% trypsin, and passaged according to experimental requirements.

[0275] 2. Drug preparation

[0276] The required volume of DMSO was calculated based on the compound's molecular weight and the mass weighed to prepare a stock solution of the same concentration (100 mM), which was then stored at 4°C.

[0277] 3. Experimental Procedure

[0278] (1) Cell seeding: Take cells in good growth condition during the logarithmic growth phase, digest the cells with 0.25% trypsin, then disperse them into a single cell suspension by pipetting with culture medium. After counting, seed them into a 96-well plate at an appropriate density (200 cells / well), 100 μL / well, and incubate in a saturated humidity, 37℃, 5% CO2 incubator.

[0279] (2) Drug treatment: After 24 hours of cell culture, six different concentration gradients of drug were added according to experimental needs, 10 μL per well, with three replicates per group and a blank group per plate. In this experiment, the test drug was diluted to the corresponding concentration gradients of 100 μM, 10 μM, 1 μM, 0.1 μM, and 0.01 μM and added for 72 hours. After drug addition, the cells were cultured in a 5% CO2, 37℃ constant temperature cell culture incubator for the corresponding time, and the cell state was observed under an inverted microscope.

[0280] (3) Color development and colorimetric analysis: After the drug reaches the action time point, add 10 μL of 2.5 mg / mL MTT solution to each well and continue incubation for 1-4 h; terminate the culture, discard the supernatant, add 100 μL of dimethyl sulfoxide solution to each well, shake at an appropriate speed for 5 min on a shaker to dissolve the purple crystals, observe the color change, place the 96-well plate in a microplate reader, detect the absorbance (OD) value of each well at a wavelength of 490 nm and compare it with the blank group.

[0281] (4) Data analysis: Calculate the cell proliferation rate and inhibition rate of each group.

[0282] Cell viability = (average OD value of test group / OD value of blank group) × 100%

[0283] Inhibition rate = (1 - average OD value of test group / OD value of blank group) × 100%

[0284] Calculate the half-maximal inhibitory concentration (IC50) of the drug. 50 Based on the drug concentration and the growth inhibition rate of the drug on cells, the IC50 was calculated using the regression-Probit method with SPSS 20.0 statistical analysis software. 50 value.

[0285] Compound inhibits human lung cancer cell line H460 WT and H460 R231Q The cell viability results are shown in the table below, where "A" indicates IC50. 50 Value < 10 μM; "B" indicates 10 μM < IC 50 Value <100μM; "C" indicates IC 50 Value > 100 μM.

[0286]

[0287]

[0288] II. In vitro enzyme inhibitory activity

[0289] 2μLDOT1L WT The enzyme solution, 2 μL of substrate (20 μM SAM, 0.15 ng / mL Nuclesomes), and 1 μL of either the test compound or a positive control compound were mixed. The control group included all substrates except the test compound. The total DOT1L test reaction volume was 5 μL. After dilution with Tris buffer (20 mM Tris-HCl, 0.01% Trixton-100, 0.1% BSA, 0.5 mM DTT, pH 8.0), the mixture was incubated at room temperature for 3 hours. The reaction was terminated by adding 2.5 μL of high-salt buffer (50 mM Tris-HCl, 1 M NaCl, 0.1% Tween-20, 0.3% poly-L-lysine, pH 7.4). The fluorescence intensity was measured using an assay kit at an excitation wavelength of 680 nM and an emission wavelength of 615 nM using Envision. All doses were set in triplicate. Nonlinear fitting was performed using GraphPadPrism 5 to calculate the IC50. 50 value.

[0290] Some compounds in this invention inhibit DOT1L WT The enzyme activity results are shown in the table below. In this table, "A" indicates IC50. 50 Value < 100 nM; "B" indicates 100 nM < IC 50 Value < 1 μM; "C" indicates 1 μM < IC 50 Value < 10 μM; "D" indicates 10 μM < IC50 Value <100μM.

[0291] Example <![CDATA[DOT1L WT IC 50 (μM / L)]]> SAH (positive control) 3.79 Example 7 C Example 10 B Example 12 B Example 14 A Example 15 B Example 18 B Example 19 A Example 20 A

Claims

1. A 1,3-disubstituted piperidine derivative, characterized in that, Selected from the following compounds: (1) 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (2) 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea; (3) 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}urea; (4) 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(methyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (5) 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea; (6) 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(2-oxo-1,2-dihydropyridine-3-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea; (7) 1-[4-(tert-butyl)phenyl]-3-{3-{isopropyl{[1-(pyrazin-2-carbonyl)piperidin-3-yl]methyl}amino}propyl}urea; (8) 1-{3-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (9) 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea; (10) 1-{3-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (11) 1-{3-{{[1-(8-bromo-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (12) 1-{3-{{[1-(2-amino-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (13) 1-[4-(tert-butyl)phenyl]-3-{3-{{[1-(2-chloro-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}urea; (14) 1-{3-{{[1-(2-amino-9-methyl-9H-purin-6-yl)piperidin-3-yl]methyl}(isopropyl)amino}propyl}-3-[4-(tert-butyl)phenyl]urea; (15) 1-(4-bromophenyl)-3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}urea; (16) 1-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}-3-(3,4,5-trimethylphenyl)urea; (17) N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}propyl}ureo}phenyl}acetamide; (18) N-{4-{3-{3-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-methoxypropyl)amino}propyl}ureido}phenyl}acetamide; (19) N-{2-{{4-{{[1-(9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide; (20) N-{2-{{4-{{[1-(2-chloro-9H-purin-6-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide; (21) N-{2-{{4-{{[1-(6-chloro-1H-pyrazolo[3,4-d]pyrimidin-4-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide; (22) N-{2-{{4-{{[1-(4-cyanopyridin-2-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazol-5-yl}neopentamide; (23) N-{2-{{4-{{[1-(3-bromoimidazo[1,2-a]pyrazin-8-yl)piperidin-3-yl]methyl}(3-morpholinopropyl)amino}piperidin-1-yl}methyl}-1H-benzo[d]imidazo-5-yl}neopentamide.

2. The process for the preparation of a 1,3-disubstituted piperidine derivative according to claim 1, characterized in that, Selected from Method A and Method B: Method A: Step 1: Intermediate M1 undergoes a reductive amination reaction with primary amines of different substitutions in the presence of a reducing agent to generate intermediate M2; Step 2: Intermediate M3 and intermediate M4 of different lengths undergo a condensation reaction in the presence of carbonyl diimidazole to generate intermediate M5; Step 3: In the presence of an oxidizing agent, intermediate M5 oxidizes its terminal hydroxyl group to an aldehyde group to generate intermediate M6; or intermediate M5 reacts with an iodine reagent to generate intermediate M7. Step 4: Intermediate M6 and intermediate M2 undergo a reductive amination reaction in the presence of a reducing agent to generate intermediate M8; or intermediate M6 and intermediate M2 undergo an N-alkylation reaction under alkaline conditions to generate intermediate M8; Step 5: Intermediate M8 is deprotected by tert-butyloxycarbonyl group under acidic conditions to generate intermediate M9; Step 6: (a) Intermediate M13 undergoes a condensation reaction with different substituted heteroaryl carboxylic acids in the presence of a condensing agent to obtain compounds (6) and (7); (b) Intermediate M9 undergoes an N-alkylation reaction with different substituted heteroaryl halides under alkaline conditions to obtain compounds (1)~(5) and (8)~(18). wherein R1, R2, R3and Het are the corresponding groups at the corresponding positions of compounds (6), (7); X is ; n = 1, m = 1; wherein R1, R2, R3and Het are the corresponding groups at the corresponding positions of compounds (1) to (5), (8) to (18); X is ; n = 1, m = 0; Method B: Step 1: Intermediate M10 reacts with primary amines of different substitutions in the presence of a reducing agent to generate intermediate M11; Step 2: Intermediate M11 undergoes a reductive amination reaction with intermediate M1 in the presence of a reducing agent to generate intermediate M12; Step 3: Intermediate M12 is deprotected by tert-butyloxycarbonyl group under acidic conditions to generate intermediate M13; Step 4: Intermediate M13 undergoes N-alkylation reaction with heteroaryl halides with different substitutions under alkaline conditions to obtain compounds (19)~(23). Where R1, R2, R3 and Het are the corresponding groups at the corresponding positions in compounds (19) to (23); X is , m=0.

3. A pharmaceutical composition comprising a 1,3-disubstituted piperidine derivative of claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

4. The use of a 1,3-disubstituted piperidine derivative of claim 1 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 3, in the preparation of a medicament for treating lung cancer.

Images