2-indolone compound as VEGFR (vascular endothelial growth factor receptor) and PKa (prokaryotic kinase a) double-target inhibitor as well as preparation method and application of 2-indolone compound

Abstract

The invention relates to a 2-indolone compound as a VEGFR (vascular endothelial growth factor receptor) and PKa double-target inhibitor as well as a preparation method and application of the 2-indolone compound. Specifically, the invention relates to a 2-indolone compound as shown in a general formula (I), and a preparation method and application thereof, and particularly relates to a 2-indolone compound for treating or preventing microangiopathy related diseases mediated by high-level VEGF protein and PKa protein, such as diabetic macular edema (DME) and age-related macular degeneration (AMD). # imgabs0 #

Description

2-Indole ketone compounds as dual-target inhibitors of VEGFR and PKa, preparation methods thereof, and uses thereof Technical Field

[0001] The present invention belongs to the field of biomedicine and relates to 2-indolone compounds that are dual-target inhibitors of vascular endothelial growth factor receptor (VEGFR) and human plasma kallikrein (PKa), methods for their preparation, and uses thereof. Specifically, the present invention relates to a 2-indolone compound represented by general formula (I), methods for its preparation, and uses thereof, particularly for treating or preventing microangiopathy-related conditions mediated by elevated levels of VEGF and PKa proteins, such as diabetic macular edema (DME) and age-related macular degeneration (AMD). Background Art

[0002] Diabetic retinopathy (DR) is one of the most common microvascular complications of diabetes. It results from chronic, progressive diabetes-induced retinal microvascular leakage and obstruction, leading to a series of fundus pathologies such as microaneurysms, hard exudates, cotton-wool spots, neovascularization, vitreous proliferation, macular edema, and even retinal detachment. Chronic hyperglycemia causes glycosylation of proteins and lipids, altering their molecular structure and leading to reduced or ineffective function of enzymes and vital cellular components. Hyperglycemia also upregulates intracellular levels of diacylglycerol (DAG), a second messenger of lipid signaling. DAG acts as a physiological activator of protein kinase C (PKC). PKC activation leads to upregulation of vascular endothelial growth factor (VEGF). Increased VEGF expression becomes a driver of disease progression, accelerating the disease by stimulating leukocyte arrest, leading to the progression of retinal nonperfusion (RNP), which further increases VEGF expression, triggering a positive feedback loop of disease progression that alters vascular permeability, causing hypoxia, blood-retinal barrier (BRB) damage, and macular edema. According to statistics from the World Health Organization, approximately 285 million people worldwide are visually impaired, of which 4.8% suffer from diabetic retinopathy (DR). Diabetic macular edema (DME), a late-stage symptom of DR, is the leading cause of irreversible vision loss in diabetic patients. The number of DME patients in China increased from 6.1535 million in 2016 to 6.9201 million in 2020, with a compound annual growth rate of 3.0% from 2016 to 2020. The number is expected to grow at an average compound annual growth rate of 2.7% from 2020 onward, reaching 7.890 million by 2025.

[0003] The VEGF-VEGFR family is currently the most studied pro-angiogenic regulatory factor, inducing endothelial cell differentiation and angiogenesis. There are five main receptors that bind to VEGF ligands: VEGFR-1, VEGFR-2, VEGFR-3, and neuropilin-1 (NRP-1) and NRP-2. Three of these vascular endothelial growth factor receptors (VEGFR-1, VEGFR-2, and VEGFR-3) are receptor tyrosine kinases.

[0004] VEGFR-1 binds to VEGF-A, VEGF-B, and PlGF, and is exclusively involved in endothelial cell differentiation, not in early embryonic angiogenesis. VEGFR-2 has a high affinity for the low-molecular-weight VEGF-A isoform (110-165 amino acid residues) and VEGF-E, and a lower affinity for VEGF-C and VEGF-D. VEGFR-3 preferentially binds to VEGF-C and VEGF-D, activating downstream signaling pathways that regulate lymphatic vessel development and function. VEGFR-2 is a key regulator of VEGF signaling, which induces endothelial cell proliferation, increases vascular permeability, and promotes angiogenesis. Furthermore, VEGFR-2 has a greater affinity for VEGF than VEGFR-1. Studies have shown that VEGFR-2 is exclusively expressed in endothelial cells, and activation of VEGFR-2 can effectively stimulate angiogenesis. Therefore, VEGFR-2 is a primary target for the development of anti-angiogenic drugs. VEGFR-2 binds to various VEGF ligands (VEGF-A, C, D, and E) to form a homodimeric complex, altering its protein conformation, leading to autophosphorylation of tyrosine residues and signaling downstream. Key signaling pathways include Ras / Raf / MAPK and PI3K / AKT. Activation of these pathways leads to transcription factor activation, cell cycle disruption, cell migration promotion, cell proliferation maintenance, and inhibition of apoptosis. Furthermore, hypoxic conditions can activate HIF-1α, inducing the transcriptional expression and secretion of VEGF-A, which then binds to VEGFR-2 on surrounding vascular endothelial cells, stimulating angiogenesis. Therefore, VEGFR-2 signaling can be inhibited by antibodies binding to VEGF-A or VEGFR-2 proteins, or by small molecule inhibitors inhibiting VEGFR-2 receptor kinase activity, thereby reducing angiogenesis.

[0005] Under normal physiological conditions, angiogenesis is in a relatively dynamic equilibrium, tightly regulated by pro-angiogenic factors (VEGF) and anti-angiogenic factors (anti-VEGF). Under pathological conditions, the so-called angiogenic switch is activated, disrupting the balance between pro-angiogenic and anti-angiogenic factors. Pro-angiogenic factors take a dominant role, promoting endothelial cell proliferation, promoting new blood vessel formation, and increasing vascular permeability. Patients with diabetic macular edema (DME) and age-related macular degeneration (AMD) experience abnormally elevated VEGF levels in the eyes, leading to the formation of dysplastic and fragile new blood vessels in the choroid. These abnormal new vessels can rapidly invade the space between the retina and choroid, causing fundus hemorrhage and fluid leakage, further damaging photoreceptors and retinal pigment epithelial cells, and resulting in severe vision loss.

[0006] The pathogenesis of diabetic macular edema (DME) and age-related macular degeneration (AMD) is also associated with the kallikrein-kinin system (KKS) signaling pathway. Plasma prekallikrein (PPK), a pre-prokallikrein in the KKS signaling pathway, is an abundant serine protease zymogen that is converted to catalytically active plasma kallikrein (PKa) by factor XIIa (FXIIa) and participates in inflammatory responses. PKa cleaves kininogen to produce bradykinin. Bradykinin (BK) is a non-peptide that stimulates the BK receptor, which is abundantly expressed in vascular cells, glial cells, and neurons. BK is a strong inflammatory factor. Activation of BK receptors can cause it to bind to B1R and B2R, which can promote the increase of bradykinin levels by stimulating the release of mediators such as eicosanoids, endothelial-derived hyperpolarizing factor, NO, propylene glycol, tissue plasminogen activator (tPA) and glucose transporter-1 / 2 (glu1 / 2), further mediating a variety of signal cascade reactions. These mediators are associated with inflammation, angiogenesis, vasodilation, and increased vascular permeability, which can induce the formation of DME. Proteomic analysis of vitreous samples from patients with DME showed that PPK and PKa increased 11.0 times compared with the control vitreous of patients with macular holes (p<0.0001). Studies have shown that systemic continuous administration of PKa inhibitors can reduce the level of DME in diabetic mice. Abnormal retinal vascular permeability and retinal blood flow. Additional preclinical evidence suggests that plasma kallikrein (PKa) may cause VEGF-mediated retinal edema. Currently, the treatment of DME mainly relies on anti-VEGF therapy, but some patients (about 50%) do not respond to VEGF therapy. This means that 50% of patients do not receive effective treatment. According to literature reports, systemic administration of PKa inhibition has a significant effect on the treatment of VEGF-induced retinal thickening in mice. The present invention provides an effective 2-indole ketone compound as a dual-targeted small molecule inhibitor of VEGFR and PKa, which provides an effective therapeutic solution for diseases such as diabetic macular edema (DME) and age-related macular degeneration (AMD) caused by retinal microvascular disorders.

[0007] Summary of the Invention

[0008] The present invention provides a 2-indolinone compound that acts as a dual-target inhibitor of VEGFR and PKa, a method for preparing the same, and uses thereof. Specifically, the present invention provides a method for preparing a 2-indolinone compound that acts as a small molecule dual-target inhibitor of VEGFR2 and PKa. The compound chemically binds a VEGFR2 ligand to a PKa ligand to prepare a dual-target small molecule inhibitor of VEGFR2 and PKa. This dual-target inhibitor has the ability to target both the "active core" of VEGFR2 and the "active core" of PKa, resulting in compounds exhibiting "dual efficacy" against VEGFR2 and PKa. The compound is a dual inhibitor of both VEGFR2 and PKa proteins and can be used as a therapeutic agent for treating conditions characterized by abnormal VEGFR2 and PKa expression, such as cardiovascular and cerebrovascular diseases, and diabetic retinopathy (e.g., diabetic macular edema and age-related macular degeneration). By targeting both VEGFR2 and PKa, the compounds of the present invention are suitable for treating subjects with elevated expression of both VEGF and PKa proteins.

[0009] Specifically, the present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof:

[0010] in,

[0011] L1 is selected from

[0012] L2 is absent or selected from a linear -(CH2) 1-22 - group, wherein the straight chain -(CH2) 1-22 - one or more -(CH2)- in the group are optionally and independently replaced by one or more selected from -C(O)-, -O-, -(NR 2 )-, -S-, -(C=C)-, -(C≡C)-, a divalent cycloalkyl group, a divalent aryl group, a divalent heterocyclic group, and a divalent heteroaryl group, L2 is optionally substituted by 1, 2, or 3 R 1 replace;

[0013] Each R 1 independently selected from C1-C6 alkyl, halogenated C1-C6 alkyl, C1-C6 alkoxy, halogenated C1-C6 alkyl, halogen, cyano and hydroxy,

[0014] Each R 2 independently selected from hydrogen, C1-C6 alkyl and acyl, wherein the C1-C6 alkyl is optionally substituted by 1, 2 or 3 R 1 replace;

[0015] The group Ar1 is selected from -(CH2)-aryl, -(CH2)-heteroaryl and wherein ring a is a 5-6 membered heterocyclic ring, the aryl, heteroaryl and Optionally, one or more selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, cyano, C1-C6 alkoxy, halo C1-C6 alkoxy, amino, hydroxy, The group substitution;

[0016] The group Ar2 is selected from -C1-C6 alkyl, -C1-C6 haloalkyl, cycloalkyl, aryl and heteroaryl, wherein the group Ar2 is optionally substituted by one or more selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 cycloalkyl, C1-C6 cycloalkylalkyl, 5-6 membered heterocyclyl and substituted by a group, a 5-6 membered heterocyclic group or Optionally substituted with one or more halogens.

[0017] In a particular embodiment, the compound represented by formula (I) or a pharmaceutically acceptable salt thereof is a compound represented by formula (II) or a pharmaceutically acceptable salt thereof:

[0018] Wherein, X is selected from oxygen atom or NR 2 ;

[0019] L2, R 2 , group Ar1 and group Ar2 are as defined above in formula (I).

[0020] In a particular embodiment, the compound represented by formula (I) or a pharmaceutically acceptable salt thereof is a compound represented by formula (III) or a pharmaceutically acceptable salt thereof:

[0021] L2, the group Ar1 and the group Ar2 are as defined above for formula (I).

[0022] In a preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, Ar1 is selected from -(CH2)-phenyl, -(CH2)-pyridyl and Wherein ring a is a 5-6 membered heteroaryl, preferably imidazolyl or pyrazolyl, more preferably for

[0023] The phenyl, pyridyl and Optionally, one or more selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, cyano, C1-C6 alkoxy, halo C1-C6 alkoxy, amino, hydroxy, The group substitution;

[0024] R is selected from hydrogen and C1-C6 alkyl.

[0025] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof:

[0026] Ar2 is selected from C 6-10 Aryl is preferably phenyl, 5 to 10 membered heteroaryl, -C1-C6 alkyl, -C1-C6 haloalkyl, C3-C6 cycloalkyl, wherein the C 6-10 Aryl, 5 to 10 membered heteroaryl, optionally substituted by one or more selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 cycloalkyl, C1-C6 cycloalkylalkyl, 5-6 membered heterocyclyl and The 5-6 membered heterocyclic group or Optionally substituted with one or more halogens.

[0027] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, L2 is selected from C 1-2 alkylene;

[0028] Alternatively, L2 is selected from -(CH2) t -(C=C)-* and -(CH2) t -(C≡C)-*; wherein t is an integer from 0 to 6, preferably an integer from 1 to 4, more preferably 1 or 2; * is the position of connection to the pyrazole ring;

[0029] Alternatively, L2 is selected from Wherein, X1, X2, X3, X4 are each independently selected from a bond, -O-, -(NR 2 )-, -C(O)NH-, -NHC(O)-, -S-, -(C=C)-, -(C≡C)-, a 4-6 membered divalent heterocyclic group, a 3-6 membered divalent cycloalkyl group, a 5-10 membered divalent heteroaryl group, and a 6-10 membered divalent aryl group, wherein the 4-6 membered divalent heterocyclic group, the 3-6 membered divalent cycloalkyl group, the 5-10 membered divalent heteroaryl group, or the 6-10 membered divalent aryl group is optionally substituted by 1, 2, or 3 R 1 Replace; each R 2 independently selected from hydrogen, C1-C6 alkyl and acyl, wherein the C1-C6 alkyl is optionally substituted by 1, 2 or 3 R 1 Replace, each R 1 Independently selected from C1-C6 alkyl, halogenated C1-C6 alkyl, C1-C6 alkoxy, halogenated C1-C6 alkyl, halogen, cyano and hydroxyl; t1, t2, t3, t4, t5 are each independently an integer from 0 to 6; * is the position connected to the pyrazole ring.

[0030] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, the divalent heterocyclic group is selected from:

[0031] The divalent heteroaryl group is selected from:

[0032] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, -L2- is selected from:

[0033] Wherein: G is a 5-6 membered heteroaryl group, preferably More preferred

[0034] G1 and G2 are each independently selected from a 4-6 membered heterocyclic group, preferably

[0035] p is 1 or 2;

[0036] s is an integer from 0 to 6, preferably 0 or 1;

[0037] s1 is an integer from 1 to 10;

[0038] s2 is 1 or 2, preferably 1;

[0039] s3 and s4 are each independently selected from an integer of 1 to 6, preferably an integer of 1 to 4, more preferably 1 or 2;

[0040] v is an integer from 0 to 4, preferably 0 or 1;

[0041] * indicates the position of connection to the pyrazole ring.

[0042] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, Ar1 is selected from

[0043] Each R 3 Each is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, C1-C6 alkoxy, halogenated C1-C6 alkoxy, Preferred are halogen, C1-C6 alkoxy, Each R 4Each is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, C1-C6 alkoxy, halo-C1-C6 alkoxy, amino, hydroxy, preferably C1-C6 alkyl, halogen, C1-C6 alkoxy, amino;

[0044] R 5 Selected from C1-C6 alkyl;

[0045] m is 0, 1, 2 or 3, preferably 2 or 3, more preferably 3;

[0046] n is 0, 1, 2 or 3, preferably 2 or 3, more preferably 3.

[0047] In another preferred embodiment, in the compound represented by formula (I), formula (II) or formula (III) or a pharmaceutically acceptable salt thereof, Ar2 is selected from C1-C6 alkyl,

[0048] R 6 is hydrogen or halogen;

[0049] R 7 is hydrogen or C3-C6 cycloalkyl;

[0050] R 8 is a C3-C6 cycloalkyl group;

[0051] Each R 9 Independently selected from hydrogen, halogen, cyano, 5-6 membered heterocyclyl, wherein the 5-6 membered heterocyclyl is optionally substituted with one or more halogens;

[0052] q is 0, 1, or 2.

[0053] Examples of compounds represented by formula (I) include, but are not limited to:

[0054] or a pharmaceutically acceptable salt thereof.

[0055] The present invention also provides a pharmaceutical composition comprising a compound represented by formula (I), formula (II) or formula (III) as described above, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.

[0056] The present invention also relates to the use of the compound represented by formula (I), formula (II) or formula (III) as described above, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, in the preparation of a drug for treating or preventing vascular disease-related diseases.

[0057] The present invention also relates to the compounds represented by formula (I), formula (II) or formula (III) as described above, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions comprising the same, for use as medicines.

[0058] The present invention also relates to the compounds represented by formula (I), formula (II) or formula (III) as described above, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions comprising the same, which are used for treating or preventing vascular disease-related diseases.

[0059] The present invention also relates to a method for treating or preventing vascular disease-related diseases, which comprises administering to a subject in need thereof a therapeutically effective amount of a compound represented by formula (I), formula (II) or formula (III) as described above, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same.

[0060] In particular embodiments, the vasculopathy-related condition is selected from diabetic retinopathy (DR), diabetic macular edema (DME), retinal vein occlusion (RVO), central retinal vein occlusion (CRVO), macular degeneration, wet age-related macular degeneration (wet AMD), retinopathy of prematurity (ROP), neovascular glaucoma, retinitis pigmentosa (RP), proliferative and non-proliferative retinopathy, retinal angiomatous proliferation, macular telangiectasia, ischemic retinopathy, iris neovascularization, intraocular neovascularization, corneal neovascularization, retinal neovascularization, polypoidal choroidal vasculopathy (PCV), choroidal neovascularization and retinal degeneration. In particular, the vasculopathy-related condition is selected from diabetic retinopathy (DR), diabetic macular edema (DME), retinal vein occlusion (RVO), central retinal vein occlusion (CRVO), wet age-related macular degeneration (wet AMD). In particular, the vasculopathy-related condition is selected from hereditary angioedema (HAE) and brain edema after stroke. In particular, the vasculopathy-related disorder is selected from diabetic complications of retinal vascular permeability associated with diabetic retinopathy and diabetic macular edema.

[0061] Preferably, the vasculopathy-related disorder is selected from diabetic macular edema (DME), wet age-related macular degeneration (AMD), non-exudative choroidal neovascularization (CNV), hereditary angioedema (HAE) and brain edema after stroke.

[0062] Terminology

[0063] Unless otherwise stated, the terms used in the specification and claims have the following meanings.

[0064] The term "alkyl" refers to a saturated straight-chain or branched aliphatic hydrocarbon group having 1 to 20 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) carbon atoms (i.e., C 1-20 The alkyl group is preferably an alkyl group having 1 to 12 carbon atoms (i.e., C 1-12 alkyl), more preferably an alkyl group having 1 to 6 carbon atoms (i.e., C 1-6 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, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2 ,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2,2-diethylpentyl, n-decyl, 3,3-diethylhexyl, 2,2-diethylhexyl, and various branched-chain isomers thereof.

[0065] The term "alkylene" refers to a divalent alkyl group, wherein alkyl is as defined above, having from 1 to 20 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) carbon atoms (i.e., C 1-20 The alkylene group is preferably an alkylene group having 1 to 12 carbon atoms (i.e., C 1-12 alkylene), more preferably an alkylene group having 1 to 6 carbon atoms (i.e., C 1-6 Alkylene). Non-limiting examples include: -CH2-, -CH(CH3)-, -C(CH3)2-, -CH2CH2-, -CH(CH2CH3)-, -CH2CH(CH3)-, -CH2C(CH3)2-, -CH2CH2CH2-, -CH2CH2CH2CH2-, and the like.

[0066] The term "alkenyl" refers to an alkyl group containing at least one carbon-carbon double bond in the molecule, wherein alkyl is as defined above and has 2 to 12 (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12) carbon atoms (i.e., C 2-12 The alkenyl group is preferably an alkenyl group having 2 to 6 carbon atoms (i.e., C 2-6 Non-limiting examples include ethenyl, propenyl, isopropenyl, butenyl, and the like.

[0067] The term "alkynyl" refers to an alkyl group containing at least one carbon-carbon triple bond in the molecule, wherein alkyl is as defined above and has 2 to 12 (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12) carbon atoms (i.e., C 2-12 The alkynyl group is preferably an alkynyl group having 2 to 6 carbon atoms (i.e., C 2-6 Non-limiting examples include ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.

[0068] The term "alkoxy" refers to -O-(alkyl), wherein alkyl is as defined above. Non-limiting examples include methoxy, ethoxy, propoxy, butoxy, and the like.

[0069] The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic all-carbon ring (i.e., monocyclic cycloalkyl) or polycyclic ring system (i.e., polycyclic cycloalkyl) having from 3 to 20 (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) ring atoms (i.e., C 3-20 The cycloalkyl group is preferably a cycloalkyl group having 3 to 12 ring atoms (i.e., C 3-12 cycloalkyl), more preferably a cycloalkyl having 3 to 8 ring atoms (ie, C 3-8 cycloalkyl), cycloalkyl having 4 to 7 ring atoms (i.e., C 4-7 cycloalkyl) or a cycloalkyl having 3 to 6 ring atoms (i.e., C 3-6 cycloalkyl).

[0070] Non-limiting examples of the monocyclic cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl and cyclooctyl.

[0071] The polycyclic cycloalkyl group includes: spirocycloalkyl group, fused cycloalkyl group and bridged cycloalkyl group.

[0072] The term "divalent cycloalkyl" refers to a cycloalkyl group with one hydrogen atom removed, which has two sites of attachment.

[0073] The term "spiroalkyl" refers to a polycyclic ring system having a common carbon atom (called a spiro atom) between the rings, which may contain one or more double bonds within the ring, or one or more heteroatoms selected from nitrogen, oxygen and sulfur (the nitrogen may be optionally oxidized to form nitrogen oxides; the sulfur may be optionally oxoed to form sulfoxides or sulfones, but does not include -OO-, -OS- or -SS-), provided that it contains at least one all-carbon ring and the point of attachment is on the all-carbon ring, and has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) ring atoms (i.e., a 5- to 20-membered spiroalkyl). The spiroalkyl preferably has 6 to 14 ring atoms (i.e., a 6- to 14-membered spiroalkyl), and more preferably has 7 to 10 ring atoms (i.e., a 7- to 10-membered spiroalkyl). The spirocycloalkyl group includes a monospirocycloalkyl group and a polyspirocycloalkyl group (such as a bispirocycloalkyl group, etc.), preferably a monospirocycloalkyl group or a bispirocycloalkyl group, more preferably a 3-membered / 4-membered, 3-membered / 5-membered, 3-membered / 6-membered, 4-membered / 4-membered, 4-membered / 5-membered, 4-membered / 6-membered, 5-membered / 3-membered, 5-membered / 4-membered, 5-membered / 5-membered, 5-membered / 6-membered, 5-membered / 7-membered, 6-membered / 3-membered, 6-membered / 4-membered, 6-membered / 5-membered, 6-membered / 6-membered, 6-membered / 7-membered, 7-membered / 5-membered or 7-membered / 6-membered monospirocycloalkyl group. Non-limiting examples include:

[0074] The term "fused cycloalkyl" refers to a polycyclic ring system in which two adjacent carbon atoms are shared between the rings, which is a monocyclic cycloalkyl fused to one or more monocyclic cycloalkyls, or a monocyclic cycloalkyl fused to one or more heterocyclyls, aryls, or heteroaryls, wherein the point of attachment is on the monocyclic cycloalkyl, which may contain one or more double bonds within the ring, and has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) ring atoms (i.e., a 5- to 20-membered fused cycloalkyl). The fused cycloalkyl is preferably a fused cycloalkyl having 6 to 14 ring atoms (i.e., a 6- to 14-membered fused cycloalkyl), more preferably a fused cycloalkyl having 7 to 10 ring atoms (i.e., a 7- to 10-membered fused cycloalkyl). The fused cycloalkyl group includes bicyclic fused cycloalkyl groups and polycyclic fused cycloalkyl groups (such as tricyclic fused cycloalkyl groups, tetracyclic fused cycloalkyl groups, etc.), preferably bicyclic fused cycloalkyl groups or tricyclic fused cycloalkyl groups, more preferably 3-membered / 4-membered, 3-membered / 5-membered, 3-membered / 6-membered, 4-membered / 4-membered, 4-membered / 5-membered, 4-membered / 6-membered, 5-membered / 3-membered, 5-membered / 4-membered, 5-membered / 5-membered, 5-membered / 6-membered, 5-membered / 7-membered, 6-membered / 3-membered, 6-membered / 4-membered, 6-membered / 5-membered, 6-membered / 6-membered, 6-membered / 7-membered, 7-membered / 5-membered or 7-membered / 6-membered bicyclic fused cycloalkyl groups. Non-limiting examples include:

[0075] The term "bridged cycloalkyl" refers to a full carbon polycyclic ring system that shares two carbon atoms that are not directly connected between the rings, which may contain one or more double bonds within the ring and has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) carbon atoms (i.e., a 5 to 20-membered bridged cycloalkyl). The bridged cycloalkyl preferably has a bridged cycloalkyl of 6 to 14 carbon atoms (i.e., a 6 to 14-membered bridged cycloalkyl), more preferably a bridged cycloalkyl of 7 to 10 carbon atoms (i.e., a 7 to 10-membered bridged cycloalkyl). The bridged cycloalkyl includes bicyclic bridged cycloalkyl and polycyclic bridged cycloalkyl (e.g., tricyclic bridged cycloalkyl, tetracyclic bridged cycloalkyl, etc.), preferably bicyclic bridged cycloalkyl or tricyclic bridged cycloalkyl. Non-limiting examples include:

[0076] The term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic heterocycle (i.e., a monocyclic heterocyclyl) or a polycyclic heterocyclic ring system (i.e., a polycyclic heterocyclyl) containing at least one (e.g., 1, 2, 3, or 4) heteroatoms selected from nitrogen, oxygen, and sulfur (the nitrogen may be optionally oxidized, i.e., to form nitrogen oxides; the sulfur may be optionally oxoed, i.e., to form sulfoxides or sulfones, but excluding -OO-, -OS-, or -SS-) in the ring, and having 3 to 20 (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) ring atoms (i.e., a 3- to 20-membered heterocyclyl). The heterocyclic group is preferably a heterocyclic group having 3 to 12 ring atoms (i.e., a 3- to 12-membered heterocyclic group); further preferably a heterocyclic group having 3 to 8 ring atoms (i.e., a 3- to 8-membered heterocyclic group) or a heterocyclic group having 4 to 7 ring atoms (i.e., a 4- to 7-membered heterocyclic group); more preferably a heterocyclic group having 3 to 6 ring atoms (i.e., a 3- to 6-membered heterocyclic group) or a heterocyclic group having 5 or 6 ring atoms (i.e., a 5- or 6-membered heterocyclic group).

[0077] Non-limiting examples of the monocyclic heterocyclic group include pyrrolidinyl, tetrahydropyranyl, dihydropyridinyl, tetrahydropyridinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl and homopiperazinyl.

[0078] The polycyclic heterocyclic group includes a spiro heterocyclic group, a fused heterocyclic group and a bridged heterocyclic group.

[0079] The term "divalent heterocyclic group" refers to a heterocyclic group with one hydrogen atom removed, which has two sites of attachment.

[0080] The term "spiroheterocyclyl" refers to a polycyclic heterocyclic ring system in which the rings share one atom (called a spiro atom), which may contain one or more double bonds in the ring and at least one (e.g., 1, 2, 3 or 4) heteroatom selected from nitrogen, oxygen and sulfur (the nitrogen may be optionally oxidized, i.e., to form a nitrogen oxide; the sulfur may be optionally oxidized, i.e., to form a sulfoxide or sulfone, but excluding -OO-, -OS- or -SS-), provided that it contains at least one monocyclic heterocyclic group and the point of attachment is on the monocyclic heterocyclic group, which has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) ring atoms (i.e., a 5- to 20-membered spiroheterocyclyl). The spiro heterocyclic radical preferably has a spiro heterocyclic radical (i.e., a 6 to 14 yuan spiro heterocyclic radical) of 6 to 14 ring atoms, more preferably a spiro heterocyclic radical (i.e., a 7 to 10 yuan spiro heterocyclic radical) with 7 to 10 ring atoms. The spiro heterocyclic radical includes monospiro heterocyclic radical and polyspiro heterocyclic radical (such as dispiro heterocyclic radical etc.), preferably monospiro heterocyclic radical or dispiro heterocyclic radical, more preferably 3 yuan / 4 yuan, 3 yuan / 5 yuan, 3 yuan / 6 yuan, 4 yuan / 4 yuan, 4 yuan / 5 yuan, 4 yuan / 6 yuan, 5 yuan / 3 yuan, 5 yuan / 4 yuan, 5 yuan / 5 yuan, 5 yuan / 6 yuan, 5 yuan / 7 yuan, 6 yuan / 3 yuan, 6 yuan / 4 yuan, 6 yuan / 5 yuan, 6 yuan / 6 yuan, 6 yuan / 7 yuan, 7 yuan / 5 yuan or 7 yuan / 6 yuan monospiro heterocyclic radical. Non-limiting examples include:

[0081] wait.

[0082] The term "fused heterocyclyl" refers to a polycyclic heterocyclic ring system that shares two adjacent atoms between the rings, which may contain one or more double bonds within the ring and at least one (e.g., 1, 2, 3, or 4) heteroatom selected from nitrogen, oxygen, and sulfur (the nitrogen may be optionally oxidized, i.e., to form a nitrogen oxide; the sulfur may be optionally oxidized, i.e., to form a sulfoxide or sulfone, but excluding -OO-, -OS-, or -SS-), which is a monocyclic heterocyclyl fused to one or more monocyclic heterocyclyls, or a monocyclic heterocyclyl fused to one or more cycloalkyl, aryl, or heteroaryl groups, wherein the point of attachment is on the monocyclic heterocyclyl, and has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) ring atoms (i.e., a 5- to 20-membered fused heterocyclyl). The fused heterocyclic radical preferably has a fused heterocyclic radical of 6 to 14 ring atoms (i.e., a 6 to 14-membered fused heterocyclic radical), more preferably a fused heterocyclic radical of 7 to 10 ring atoms (i.e., a 7 to 10-membered fused heterocyclic radical). The fused heterocyclic radical includes bicyclic and polycyclic fused heterocyclic radicals (such as tricyclic fused heterocyclic radicals, tetracyclic fused heterocyclic radicals, etc.), preferably bicyclic fused heterocyclic radicals or tricyclic fused heterocyclic radicals, more preferably 3 yuan / 4 yuan, 3 yuan / 5 yuan, 3 yuan / 6 yuan, 4 yuan / 4 yuan, 4 yuan / 5 yuan, 4 yuan / 6 yuan, 5 yuan / 3 yuan, 5 yuan / 4 yuan, 5 yuan / 5 yuan, 5 yuan / 6 yuan, 5 yuan / 7 yuan, 6 yuan / 3 yuan, 6 yuan / 4 yuan, 6 yuan / 5 yuan, 6 yuan / 6 yuan, 6 yuan / 7 yuan, 7 yuan / 5 yuan or 7 yuan / 6 yuan bicyclic fused heterocyclic radicals. Non-limiting examples include:

[0083] wait.

[0084] The term "bridged heterocyclic group" refers to a polycyclic heterocyclic ring system that shares two atoms that are not directly connected between the rings, which may contain one or more double bonds within the ring and at least one (e.g., 1, 2, 3, or 4) heteroatom selected from nitrogen, oxygen, and sulfur (the nitrogen may be optionally oxidized, i.e., to form a nitrogen oxide; the sulfur may be optionally oxidized, i.e., to form a sulfoxide or sulfone, but excluding -OO-, -OS-, or -SS-), and has 5 to 20 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20) ring atoms (i.e., a 5- to 20-membered bridged heterocyclic group). The bridged heterocyclic group is preferably a bridged heterocyclic group having 6 to 14 ring atoms (i.e., a 6- to 14-membered bridged heterocyclic group), and more preferably a bridged heterocyclic group having 7 to 10 ring atoms (i.e., a 7- to 10-membered bridged heterocyclic group). According to the number of constituent rings, heterocyclic groups can be divided into bicyclic bridged heterocyclic groups and polycyclic bridged heterocyclic groups (such as tricyclic bridged heterocyclic groups, tetracyclic bridged heterocyclic groups, etc.), preferably bicyclic bridged heterocyclic groups or tricyclic bridged heterocyclic groups. Non-limiting examples include:

[0085] wait.

[0086] The term "aryl" refers to a monocyclic all-carbon aromatic ring (i.e., monocyclic aromatic group) or a polycyclic aromatic ring system (i.e., polycyclic aromatic group) having a conjugated π electron system, which has 6 to 14 (e.g., 6, 7, 8, 9, 10, 11, 12, 13, or 14) ring atoms (i.e., 6- to 14-membered aromatic group). The aryl group is preferably an aromatic group having 6 to 10 ring atoms (i.e., 6- to 10-membered aromatic group), more preferably an aromatic group having 8 to 10 ring atoms (i.e., 8- to 10-membered polycyclic aromatic group). The monocyclic aromatic group is, for example, phenyl. Non-limiting examples of the polycyclic aromatic group include: naphthyl, anthracenyl, phenanthrenyl, etc.

[0087] The term "divalent aryl" refers to an aryl group with one hydrogen atom removed, which has two sites of attachment.

[0088] The term "heteroaryl" refers to a monocyclic heteroaromatic ring (i.e., a monocyclic heteroaryl) or a polycyclic heteroaromatic ring system (i.e., a polycyclic heteroaryl) having a conjugated π electron system, which contains at least one (e.g., 1, 2, 3, or 4) heteroatoms selected from nitrogen, oxygen, and sulfur (the nitrogen may be optionally oxidized, i.e., to form a nitrogen oxide; the sulfur may be optionally oxidized, i.e., to form a sulfoxide or sulfone, but excluding -OO-, -OS-, or -SS-), and has 5 to 14 (e.g., 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14) ring atoms (i.e., a 5- to 14-membered heteroaryl). The heteroaryl is preferably a heteroaryl having 5 to 10 ring atoms (i.e., a 5- to 10-membered heteroaryl), more preferably a heteroaryl having 5 or 6 ring atoms (i.e., a 5- or 6-membered monocyclic heteroaryl), or preferably a heteroaryl having 8 to 10 ring atoms (i.e., an 8- to 10-membered polycyclic heteroaryl).

[0089] The monocyclic heteroaryl groups include, but are not limited to, furyl, thienyl, thiazolyl, isothiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, furazanyl, pyrrolyl, N-alkylpyrrolyl, pyridyl, pyrimidinyl, pyridonyl, N-alkylpyridone (e.g. etc.), pyrazinyl, pyridazinyl, etc.

[0090] Non-limiting examples of the polycyclic heteroaryl group include indolyl, indazolyl, quinolyl, isoquinolyl, quinoxalinyl, phthalazinyl, benzimidazolyl, benzothiophenyl, benzofuranyl, quinazolinyl, carbazolyl, pyrrolotriazinyl, and the like.

[0091] The term "divalent heteroaryl" refers to a heteroaryl group with one hydrogen atom removed, which has two sites of attachment.

[0092] The term "haloalkyl" refers to an alkyl group substituted with one or more halogens, wherein alkyl is as defined above.

[0093] The term "haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein alkoxy is as defined above.

[0094] The term "hydroxyalkyl" refers to an alkyl group substituted with one or more hydroxy groups, wherein alkyl is as defined above.

[0095] The term "halogen" refers to fluorine, chlorine, bromine or iodine.

[0096] The term "hydroxy" refers to -OH.

[0097] The term "mercapto" refers to -SH.

[0098] The term "amino" refers to -NH2.

[0099] The term "cyano" refers to -CN.

[0100] The term "nitro" refers to -NO2.

[0101] The term "oxo" or "oxo" refers to "=0".

[0102] The term "carbonyl" refers to C=O.

[0103] The term "carboxy" refers to -C(O)OH.

[0104] The compounds of the present invention may exist in specific stereoisomeric forms. The term "stereoisomer" refers to isomers with identical structures but different arrangements of atoms in space. These include cis and trans (or Z and E) isomers, (-)- and (+)-isomers, (R)- and (S)-enantiomers, diastereomers, (D)- and (L)-isomers, tautomers, atropisomers, conformers, and mixtures thereof (e.g., racemates, mixtures of diastereomers). Substituents in the compounds of the present invention may have additional asymmetric atoms. All of these stereoisomers and their mixtures are included within the scope of the present invention. Optically active (-)- and (+)-isomers, (R)- and (S)-enantiomers, and (D)- and (L)-isomers can be prepared by chiral synthesis, chiral reagents, or other conventional techniques. An isomer of a compound of the present invention can be prepared by asymmetric synthesis or chiral auxiliary, or, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), by forming a diastereomeric salt with an appropriate optically active acid or base, followed by diastereomeric resolution by conventional methods known in the art to obtain the pure isomer. Furthermore, separation of enantiomers and diastereoisomers is typically accomplished by chromatography.

[0105] In the chemical structure of the compound of the present invention, the bond Indicates that the configuration is not specified, that is, if chiral isomers exist in the chemical structure, the bond Can be or or include both and Two configurations.

[0106] "Optional" or "optionally" means that the event or circumstances described subsequently may but need not occur, and includes both situations in which the event or circumstances occur and do not occur. For example, "alkyl optionally substituted with halogen or cyano" includes both situations in which the alkyl is substituted with halogen or cyano and situations in which the alkyl is not substituted with halogen and cyano.

[0107] "Substitution" or "substituted" means that one or more hydrogen atoms, preferably 1-6, more preferably 1-3, and even more preferably 1-2 hydrogen atoms in a group are replaced independently of one another by a corresponding number of substituents. Those skilled in the art can determine (by experiment or theory) whether substitution is possible or not without undue effort. For example, an amino group or a hydroxyl group having free hydrogen may be unstable when combined with a carbon atom having an unsaturated bond (such as an alkene).

[0108] A "pharmaceutical composition" refers to a mixture containing one or more compounds described herein, or pharmaceutically acceptable salts thereof, and other chemical components, as well as other components such as pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitating absorption of the active ingredient and thereby exerting its biological activity.

[0109] "Pharmaceutically acceptable salts" refer to salts of the compounds of the present invention, which may be selected from inorganic or organic salts. Such salts are safe and effective for use in mammals and possess the desired biological activity. They can be prepared during the final isolation and purification of the compound, or separately by reacting a suitable group with a suitable base or acid. Bases commonly used to form pharmaceutically acceptable salts include inorganic bases, such as sodium hydroxide and potassium hydroxide, and organic bases, such as ammonia. Acids commonly used to form pharmaceutically acceptable salts include inorganic acids and organic acids.

[0110] As used herein, the term "treat" includes treatment of a disease or condition in a mammal, particularly a human, and includes: (a) inhibiting the infection, disease or condition, i.e., arresting or delaying the development of the infection, disease or condition; (b) relieving the infection, disease or condition, i.e., causing regression of the disease or condition, and / or (c) curing the infection, disease or condition.

[0111] As used herein, the term "prophylaxis" includes prophylactic therapy in mammals, particularly humans, to reduce the likelihood of developing an infection, disease, or condition. Patients may be selected for prophylactic therapy based on their increased risk of infection, disease, or condition compared to the general population. "Prophylaxis" may include treatment of subjects who have not yet developed an infection or clinical condition, and prevention of a second occurrence of the same or similar infection or clinical condition.

[0112] With respect to a drug or pharmacologically active agent, the term "therapeutically effective amount" refers to an amount of the drug or agent sufficient to achieve, or at least partially achieve, the desired effect. The determination of a therapeutically effective amount varies from person to person, depending on the age and general condition of the recipient, as well as the specific active substance. The appropriate therapeutically effective amount in each individual case can be determined by those skilled in the art through routine experimentation.

[0113] The term "pharmaceutically acceptable" as used herein refers to compounds, materials, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with patient tissues without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio, and effective for the intended use.

[0114] As used herein, the singular form "a," "an," and "the" include plural references and vice versa unless the context clearly dictates otherwise.

[0115] When the term "about" is applied to a parameter such as pH, concentration, temperature, etc., it indicates that the parameter can vary by ±10%, and sometimes more preferably within ±5%. As will be understood by those skilled in the art, when a parameter is not critical, numbers are generally given for illustration purposes only and are not limiting.

[0116] It should be noted that, unless otherwise explicitly stated, the term "independently selected" used in this invention should be broadly interpreted, meaning that each substituent is selected independently of the others. Therefore, each substituent may be the same as or different from the others. More specifically, the term "independently selected" can mean that the specific options represented by the same symbol in different groups do not affect each other, or that the specific options represented by the same symbol in the same group do not affect each other. DETAILED DESCRIPTION

[0117] The following examples are only used to illustrate the present invention, but are not used to limit the present invention. Any modifications, changes, variations, etc. made within the scope of protection of the present invention are all within the scope of protection of the present invention.

[0118] The compounds provided herein can be prepared using the specific synthetic schemes described below from readily available starting materials, which can be synthesized using methods known in the art or commercially available. Experimental methods in the following examples, where specific conditions are not specified, can be determined by conventional methods and conditions by those skilled in the art through routine optimization procedures.

[0119] The structures of the compounds were determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). -6 The unit of ppm is given. NMR measurements were performed using a Bruker dps400 nuclear magnetic spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), and deuterated methanol (CD3OD), and tetramethylsilane (TMS) was used as the internal standard.

[0120] LC-MS measurements were performed using an 1100 Series LC / MSD Trap (ESI) mass spectrometer (manufacturer: Agilent).

[0121] GC-MS was performed using a GCMS-QP2010 SE.

[0122] Preparative liquid chromatography was performed using an LC3000 high performance liquid chromatograph and an LC6000 high performance liquid chromatograph (manufacturer: Chuangxin Tongheng). The column chromatography was performed using a Daisogel C18 10 μm 60A (20 mm×250 mm).

[0123] High performance liquid chromatography (HPLC) was performed using a Shimadzu LC-20AD high pressure liquid chromatograph (Agilent TC-C18 250×4.6 mm 5 μm column chromatography) and a Shimadzu LC-2010AHT high pressure liquid chromatograph (Phenomenex C18 250×4.6 mm 5 μm column chromatography).

[0124] The thin layer chromatography silica gel plate used was Qingdao Ocean Chemical GF254 silica gel plate. The silica gel plate used in thin layer chromatography (TLC) had a specification of 0.15 mm to 0.2 mm, and the specification used for thin layer chromatography separation and purification products was 0.4 mm to 0.5 mm.

[0125] Column chromatography generally uses Qingdao marine silica gel 100-200 mesh and 200-300 mesh silica gel as the carrier.

[0126] The known starting materials of the present invention can be synthesized by methods known in the art, or can be purchased from online shopping malls, Beijing Coupling, Sigma, Bailingwei, Yishiming, Shanghai Shuya, Yinuokai, Nanjing Yaoshi, Anaiji Chemical and other companies.

[0127] Unless otherwise specified in the examples, all reactions can be carried out under an argon atmosphere or a nitrogen atmosphere.

[0128] Argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a capacity of about 1 L.

[0129] A CEM Discover SP microwave reactor was used for the microwave reaction.

[0130] Unless otherwise specified in the examples, the solution refers to an aqueous solution.

[0131] Unless otherwise specified in the examples, the reaction temperature is room temperature, particularly 20°C to 30°C.

[0132] The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent systems used in the reactions were: A: dichloromethane and methanol system, B: n-hexane and ethyl acetate system, C: petroleum ether and ethyl acetate system, and D: acetone. The volume ratio of the solvents was adjusted according to the polarity of the compounds.

[0133] The eluent system for column chromatography and the developing solvent system for thin-layer chromatography used to purify the compound include: A: dichloromethane and methanol system, B: petroleum ether, ethyl acetate and dichloromethane system, C: petroleum ether and ethyl acetate system. The volume ratio of the solvent is adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid can also be added for adjustment.

[0134] Example

[0135] Preparation Example 1: Preparation of tert-butyl 2-(2-hydroxyethoxy)ethoxyacetate (Intermediate 1)

[0136] Step 1: Under a nitrogen atmosphere, compound Int1-1 (10.0 g, 94.34 mmol) was added to THF (50 mL). The temperature was cooled to 0°C, and sodium hydride (8.3 g, 0.21 mol, 60% purity) was added portionwise. The mixture was stirred at 0°C for 1 h. Benzyl bromide (16.1 g, 94.34 mmol) dissolved in THF (50 mL) was slowly added dropwise. The temperature was raised to 70°C and stirred for 16 h. The mixture was cooled to room temperature, and saturated aqueous ammonium chloride (200 mL) was added. The THF was removed and the mixture was concentrated under reduced pressure. The mixture was then extracted with ethyl acetate (100 mL x 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound Int1-2 (10.0 g, yield: 56%) as a yellow oily liquid.

[0137] MS (ESI) M / Z: 219.10 [M+Na] + .

[0138] 1 H NMR (400MHz, CHCl3-d) δ7.38–7.26(m,5H),4.58(s,2H),3.76–3.68(m,4H),3.66–3.60(m,4H).

[0139] Step 2: Under nitrogen atmosphere, compound Int1-2 (5.0 g, 25.51 mmol) was added to THF (25 mL), cooled to 0°C, and sodium hydride (8.3 g, 30.61 mmol) was added portionwise. The mixture was stirred at 0°C for 1 h. 2-Butyl 2-bromoacetate (1.2 g, 94.34 mmol) dissolved in THF (50 mL) was slowly added dropwise, and the mixture was stirred at room temperature for 16 h. Saturated aqueous ammonium chloride (100 mL) was added, and the THF was removed by concentration under reduced pressure. The mixture was extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound Int1-3 (3.0 g, yield: 38%) as a yellow oily liquid.

[0140] MS (ESI) M / Z: 311.2 [M+H] + .

[0141] 1 H NMR (400MHz, CHCl3-d) δ7.37–7.25(m,5H),4.57(s,2H),4.03(s,2H),3.81–3.58(m,8H),1.47(s,9H).

[0142] Step 3: Under a nitrogen atmosphere, compound Int1-3 (3.0 g, 9.68 mmol), acetic acid (580 mg, 9.68 mmol), and Pd / C (10%, 300 mg) were added sequentially to ethanol. The mixture was replaced with hydrogen three times and allowed to react at room temperature for 16 h. The mixture was filtered, the filtrate was collected, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain Intermediate 1 (2.0 g, yield: 94%) as a yellow oil.

[0143] MS (ESI) M / Z: 243.10 [M+Na] + .

[0144] 1H NMR (400MHz, CDCl3-d) δ4.01(s,2H),3.75–3.68(m,6H),3.64–3.60(m,2H),1.47(s,9H).

[0145] Preparation Example 2: Preparation of ethyl 3-(bromomethyl)-1-(4-((2-oxopyridin-1(2H)yl)methyl)benzyl)-1H-pyrazole-4-carboxylate (Intermediate 2)

[0146] Step 1: At room temperature, compound Int2-1 (10.0 g, 42.02 mmol) and DMF-DMA (5.0 g, 42.02 mmol) were added sequentially to DMF (100 mL). The mixture was stirred at 110°C for 16 h. The mixture was cooled to room temperature and concentrated under reduced pressure to afford crude compound Int2-2 (13.0 g) as a yellow oil. This was used in the next step without further purification.

[0147] MS (ESI) M / Z: 292.1 [M+H] + .

[0148] Step 2: Compound Int2-2 (13.0 g, 42.02 mmol), HOAc (2.5 g, 42.02 mmol), and N-tert-butyl hydrazine (80%, 2.6 g, 42.02 mmol) were added sequentially to EtOH (100 mL) at room temperature. The temperature was raised to 80°C and stirred for 16 h. The mixture was cooled to room temperature and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound Int2-3 (8.9 g, yield: 82%) as a yellow oil.

[0149] MS (ESI) M / Z: 261.1 [M+H] + .

[0150] 1 H NMR (400MHz, DMSO-d6) δ13.42(d,J=70.4Hz,1H),8.06(d,J=153.2Hz,1H),7.44–7. 20(m,5H),4.74(s,2H),4.56(s,2H),4.19(q,J=7.2Hz,2H),1.24(t,J=7.2Hz,3H).

[0151] Step 3: Under a nitrogen atmosphere, compound Int2-3 (8.0 g, 30.77 mmol), acetic acid (1.8 g, 30.77 mmol), and Pd / C (10%, 1.0 g) were added sequentially to ethanol. The mixture was replaced with hydrogen three times and allowed to react at room temperature for 16 h. The filtrate was filtered, collected, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (EA:PE = 3:1) to obtain compound Int2-4 (4.8 g, yield: 92%) as a yellow solid.

[0152] MS (ESI) M / Z: 171.1 [M+H] + .

[0153] 1 H NMR (400MHz, CHCl3-d) δ7.94 (s, 1H), 4.89 (s, 2H), 4.29 (q, J = 7.2Hz, 2H), 2.04 (s, 3H), 1.34 (t, J = 7.2Hz, 3H).

[0154] Step 4: Compound Int2-4 (4.8 g, 28.24 mmol) and potassium carbonate (19.5 g, 141.20 mmol) were added sequentially to DMF (100 mL) at room temperature and stirred at room temperature for 5 minutes. Compound Int2-5 (9.3 g, 33.89 mmol) was added and stirred at room temperature for 16 hours. Water (500 mL) was added and the mixture was extracted with dichloromethane (200 mL × 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride solution (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (EA:PE = 0-100%) to obtain compound Int2-6 (7.6 g, yield: 74%) as a white solid.

[0155] MS (ESI) M / Z: 368.1 [M+H] + .

[0156] 1 H NMR (400MHz, DMSO-d6) δ8.37(s,1H),7.79–7.73(m,1H),7.47–7.33(m,1H),7.25(s,4H),6.43–6.36(m,1H),6.22(td,J=6.4,1. 2Hz, 1H), 5.28 (s, 2H), 5.06 (s, 2H), 4.84 (t, J = 6.0Hz, 1H), 4.56 (d, J = 6.0Hz, 2H), 4.18 (q, J = 7.2Hz, 2H), 1.25 (t, J = 7.2Hz, 3H).

[0157] Step 5: Compound Int2-6 (1.0 g, 2.72 mmol) and phosphorus tribromide (1.1 g, 4.09 mmol) were added sequentially to acetonitrile (10 mL) at room temperature and stirred at 70°C for 1 h. The mixture was cooled to room temperature and poured into an ice-cold saturated aqueous sodium bicarbonate solution (100 mL). Extraction was performed with dichloromethane (50 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to afford Intermediate 2 (1.1 g, yield: 92%) as a white solid.

[0158] MS (ESI) M / Z: 429.1 [M+H] + .

[0159] Preparation Example 3: Preparation of (Z)-N-(2-aminoethyl)-5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamide (Intermediate 3)

[0160] Step 1: Under nitrogen atmosphere, compound Int3-1 (780 mg, 8.0 mmol) was added to H2O (18 mL), cooled to 0°C, and potassium hydroxide (500 mg, 16.0 mmol) was added portionwise. The temperature was raised to 100°C and stirred for 4 h. HCl (10 mL) was added at 0°C, and the mixture was rinsed with DCM (30 mL × 3). The mixture was filtered under reduced pressure to obtain compound Int3-2 (640 mg, crude) as a yellow oily solid.

[0161] MS (ESI) M / Z: 167.1 [M+H] + .

[0162] Step 2: Compound Int3-2 (1.8 g, 11.9 mmol) was added to ethanol (18 mL) at room temperature. Compound Int3-3 (1.92 g, 11.9 mmol) and piperidine (2.4 mL, 24 mmol) were added to the mixture in sequence. The mixture was stirred at room temperature for 0.5 h, then heated to 90°C and stirred for 16 h. HCl (10 mL) was added at 0°C and filtered under reduced pressure to obtain a solid. The solid was slurried with a mixture of DCM (30 mL) and MeOH (3 mL). The residue was filtered and washed with EtOH (30 mL) to obtain Int3-4 (2.4 g, 63%) as a yellow solid.

[0163] MS (ESI) M / Z: 301.1 [M+H] + .

[0164] 1H NMR (400MHz, DMSO-D6) δ 13.84 (s, 1H), 12.14 (s, 1H), 10.92 (s, 1H), 7.85 – 7.63 (m, 2H), 6.98 – 6.76 (m, 2H), 2.47 (d, J = 2.2Hz, 6H).

[0165] Step 3: Compound Int3-4 (100 mg, 0.33 mmol), compound Int3-5 (64 mg, 0.40 mmol), HOBT (67.5 mg, 0.50 mmol), EDCI (95.05 mg, 0.50 mmol), and triethylamine (67.30 mg, 0.66 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 12 h. Water (10 mL) was added, and the mixture was extracted with DCM (10 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (dichloromethane:methanol = 0-10%) to obtain compound Int3-6 (39 mg, yield: 26.7%) as a yellow solid.

[0166] MS (ESI) M / Z: 443.6 [M+H] + .

[0167] Step 4: Compound Int3-6 (29 mg, 0.07 mmol) was added to a hydrochloric acid / dioxane solution (3 mL) at room temperature, cooled to 0°C, and stirred at room temperature for 1 hour. The pH of the reaction solution was adjusted to alkaline with saturated aqueous sodium bicarbonate solution, extracted with DCM (10 mL × 3), and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain intermediate 3 (30 mg, hydrochloride) as a white solid.

[0168] MS (ESI) M / Z: 343.14 [M+1] + .

[0169] Preparation Example 4: Preparation of (S,Z)-5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-N-(pyrrolidin-3-yl)-1H-pyrrole-3-carboxamide (Intermediate 4)

[0170] Step 1: Compound Int3-4 (500 mg, 1.67 mmol), compound Int4-1 (372 mg, 2.00 mmol), HOBT (338 mg, 2.50 mmol), EDCI (477.5 mg, 2.50 mmol), and triethylamine (336.00 mg, 3.34 mmol) were added sequentially to DMF (5 mL) at room temperature and stirred at room temperature for 12 h. Water (60 mL) was added, and the mixture was extracted with DCM (60 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (PE:EA = 0-100%) to obtain compound Int4-2 (491 mg, yield: 62.9%) as a yellow solid.

[0171] MS (ESI) M / Z: 469.6 [M+H] + .

[0172] Step 2: Compound Int4-2 (80 mg, 0.17 mmol) was added to a hydrochloric acid / dioxane solution (10 mL) at room temperature, cooled to 0°C, and stirred at room temperature for 1 hour. The pH of the reaction solution was adjusted to alkaline with saturated aqueous sodium bicarbonate solution, and extracted with DCM (10 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford Intermediate 4 (80 mg, hydrochloride salt) as a white solid.

[0173] MS (ESI) M / Z: 369.14 [M+1] + .

[0174] Preparation Example 5: Preparation of (S,Z)-3-((2-(4-(3-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamide)pyrrolidine-1-carbonyl)piperazin-1-yl)ethoxy)methyl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxylic acid (Intermediate 5)

[0175] Step 1: Under nitrogen atmosphere, compound Int5-1 (386 mg, 1.68 mmol) was added to THF (5 mL), cooled to 0°C, and sodium hydride (67 mg, 1.68 mmol) was added. The mixture was stirred at 0°C for 1 h, and intermediate 2 (600 mg, 1.40 mmol) dissolved in DMF (5 mL) was slowly added dropwise. The mixture was stirred at room temperature for 16 h. Saturated aqueous ammonium chloride (50 mL) was added, and the THF was removed by concentration under reduced pressure. The mixture was extracted with ethyl acetate (50 mL × 3). The organic phases were combined, washed with water (100 mL) and saturated aqueous sodium chloride (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound Int5-2 (115 mg, yield: 15%) as a yellow oily liquid.

[0176] MS (ESI) M / Z: 580.3 [M+H] + .

[0177] Step 2: Compound Int5-2 (115 mg, 0.20 mmol) was added to a solution of hydrogen chloride / dioxane (4.0 M, 5 mL), reacted at room temperature for 1 hour, and concentrated under reduced pressure to obtain compound Int5-3 (95 mg, yield: 100%) as a white solid.

[0178] MS (ESI) M / Z: 480.3 [M+H] + .

[0179] Step 3: Under nitrogen atmosphere, compound Int5-3 (95 mg, 0.2 mmol) was dissolved in DCM (2 mL), cooled to 0°C, and a solution of triphosgene (0.1 mmol) dissolved in DCM (2 mL) was added dropwise. The mixture was stirred at 0°C for 30 minutes, and intermediate 4 (73 mg, 0.2 mmol) dissolved in DMF (2 mL) was added dropwise. The mixture was naturally returned to room temperature and reacted for 1 hour. Water (20 mL) was added and extracted with DCM (20×3). The organic phases were combined, washed with water (30 mL) and saturated sodium chloride solution (30 mL), dried over anhydrous sulfuric acid, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound Int5-4 (100 mg, yield: 57%) as a yellow solid.

[0180] MS (ESI) M / Z: 874.4 [M+H] + .

[0181] Step 4: Compound Int5-4 (100 mg, 0.11 mmol) was added to ethanol (3 mL) at room temperature. A solution of sodium hydroxide (23 mg, 0.57 mmol) dissolved in water (0.6 mL) was added and stirred at room temperature for 24 hours. The pH of the reaction solution was adjusted to acidic with 1N aqueous hydrogen chloride solution and extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to afford Intermediate 5 (60 mg, yield: 62%) as a yellow solid.

[0182] MS (ESI) M / Z: 846.4 [M+H] + .

[0183] Preparation Example 6: Preparation of tert-butyl 2-(prop-2-en-1-oxy)acetate (Intermediate 6)

[0184] Under nitrogen atmosphere, tert-butyl 2-hydroxyacetate (6.7 g, 50.85 mmol) was added to THF (25 mL). The temperature was cooled to 0°C, and sodium hydride (2.0 g, 50.85 mmol) was added portionwise. The mixture was stirred at 0°C for 1 h. 3-Bromo-1-propyne (5.0 g, 42.37 mmol) dissolved in THF (25 mL) was slowly added dropwise. The mixture was stirred at room temperature for 16 h. The temperature was cooled to 0°C, and saturated aqueous ammonium chloride (100 mL) was added. The mixture was concentrated under reduced pressure to remove THF and extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain intermediate 6 (2.3 g, yield: 32%) as a yellow oily liquid.

[0185] MS (ESI) M / Z: 171.1 [M+H] + .

[0186] Preparation Example 7: Preparation of tert-butyl 4-(prop-2-en-1-oxy)piperidine-1-carboxylate (Intermediate 7)

[0187] Under a nitrogen atmosphere, compound intermediate 7-1 (6 g, 30 mmol) was added to THF (40 mL), cooled to 0°C, and sodium hydride (1.21 g, 30 mmol) was added portionwise. The mixture was stirred at 0°C for 1 h. 3-Bromo-1-propyne (3.77 g, 32 mmol) dissolved in THF (25 mL) was slowly added dropwise, and the mixture was stirred at room temperature for 16 h. The mixture was cooled to 0°C, and saturated aqueous ammonium chloride (100 mL) was added. The THF was removed by concentration under reduced pressure, and the mixture was extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound intermediate 7 (4.3 g, yield: 60%) as a yellow oily liquid.

[0188] MS (ESI) M / Z: 240.2 [M+H] + .

[0189] Preparation Example 8: Preparation of tert-butyl 3-(prop-2-en-1-oxy)azetidine-1-carboxylate (Intermediate 8)

[0190] Under a nitrogen atmosphere, compound intermediate 8-1 (6.92 g, 40 mmol) was added to THF (60 mL). The temperature was lowered to 0°C, and sodium hydride (1.62 g, 40 mmol) was added portionwise. The mixture was stirred at 0°C for 1 h. 3-Bromo-1-propyne (4.77 g, 40 mmol) dissolved in THF (25 mL) was slowly added dropwise, and the mixture was stirred at room temperature for 16 h. The temperature was lowered to 0°C, and saturated aqueous ammonium chloride (100 mL) was added. The THF was removed by concentration under reduced pressure, and the mixture was extracted with ethyl acetate (100 mL x 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound intermediate 8 (5.5 g, yield: 65%) as a yellow oily liquid.

[0191] MS (ESI) M / Z: 212.1 [M+H] + .

[0192] Preparation Example 9: Preparation of tert-butyl (R)-3-(prop-2-yn-1-yloxy)pyrrolidine-1-carboxylate (Intermediate 9)

[0193] Under nitrogen atmosphere, compound intermediate 9-1 (5.61 g, 30 mmol) was added to THF (50 mL), cooled to 0°C, and sodium hydride (1.22 g, 30 mmol) was added portionwise. The mixture was stirred at 0°C for 1 h. 3-Bromo-1-propyne (4.17 g, 35 mmol) dissolved in THF (25 mL) was slowly added dropwise, and the mixture was stirred at room temperature for 16 h. The mixture was cooled to 0°C, and saturated aqueous ammonium chloride (100 mL) was added. The THF was removed by concentration under reduced pressure, and the mixture was extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with water (200 mL) and saturated aqueous sodium chloride (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (ethyl acetate:petroleum ether = 0-100%) to obtain compound intermediate 9 (3.92 g, yield: 58%) as a yellow oily liquid.

[0194] MS (ESI) M / Z: 226.1 [M+H] + .

[0195] Preparation Example 10: Preparation of tert-butyl (R)-3-(prop-2-yn-1-yloxy)pyrrolidine-1-carboxylate (Intermediate 10)

[0196] Intermediate 10 was prepared by the same synthetic method as Intermediate 9.

[0197] MS (ESI) M / Z: 226.1 [M+H] + .

[0198] Preparation Example 11: Preparation of ethyl 3-bromo-1-(4-(2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxylate (Intermediate 11)

[0199] Step 1: To a solution of pyridin-2(H)-one (25.0 g, 262 mmol, 1.00 eq) in MeCN (500 mL) was added 1,4-bis(bromomethyl)benzene (104.08 g, 394.33 mmol, 1.50 eq) and K2CO3 (72.7 g, 525 mmol, 2.00 eq). The reactants were stirred at 80 ° C for 12 hours. The reaction mixture was filtered and the filter cake was washed with DCM (200 mL). It was then diluted with H2O (500 mL) and extracted with DCM 1500 mL (500 mL×3). The combined organic layers were washed with brine (500 mL×3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO 2 , PE / EtOAc=1 / 0 to 1 / 1, plate 2, PE:EtOAc=1:1, Rf=0.45) to obtain intermediate 11-1 (8.00 g, 28.8 mmol, 10.9% yield) as a white solid.

[0200] LCMS (ESI): m / z 278[M+1] + .

[0201] 1 H NMR (400MHz, CDCl3) δ7.70-7.69 (d, J=1.6Hz, 1H), 7.68-7.53 (m, 1H), 7.40 -7.38(d,J=8Hz,2H),7.28-7.26(d,J=8Hz,2H),6.59-6.56(d,J=8.8Hz,1H),6.41-6.38(m,1H),5.19(s,2H),4.53(s,2H).

[0202] Step 2: Intermediate 11-1 (200 mg, 0.72 mmol), ethyl 3-bromo-1H-pyrazole-4-carboxylate (157 mg, 0.72 mmol), and potassium carbonate (197 mg, 1.44 mmol) were added sequentially to DMF (5 mL) and reacted at room temperature overnight. The mixture was filtered, the filter cake was washed with ethyl acetate, and the filtrate was concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (petroleum ether / ethyl acetate = 0-100%) to obtain Intermediate 11 (250 mg, yield: 83.7%) as a white solid.

[0203] MS (ESI) M / Z: 416.0 [M+H] + .

[0204] Example 1: Preparation of (S,Z)-N-((6-amino-2,4-dimethylpyridin-3-yl)methyl)-3-((2-(2-(2-(3-(5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)pyrrolidin-1-yl)-2-oxoethoxy)ethoxyethoxymethyl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamido (Compound 46)

[0205] Step 1: Under nitrogen atmosphere, intermediate 2 (466 mg, 2.33 mmol) was added to THF (5 mL), cooled to 0°C, and sodium hydride (93 mg, 2.33 mmol) was added. The mixture was stirred at 0°C for 1 h, and intermediate 1 (500 mg, 1.16 mmol) dissolved in DMF (5 mL) was slowly added dropwise. The mixture was stirred at room temperature for 16 h. Saturated aqueous ammonium chloride (50 mL) was added, and the THF was removed by concentration under reduced pressure. The mixture was extracted with ethyl acetate (50 mL × 3). The organic phases were combined, washed with water (100 mL) and saturated aqueous sodium chloride (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 46-1 (150 mg, yield: 23%) as a yellow oily liquid.

[0206] MS (ESI) M / Z: 569.3 [M+H] + .

[0207] Step 2: Compound 46-1 (150 mg, 0.26 mmol) was added to a solution of hydrogen chloride / dioxane (4.0 M, 5 mL), reacted at room temperature for 1 hour, and concentrated under reduced pressure to obtain compound 46-2 (150 mg, yield: 100%) as a white solid.

[0208] MS (ESI) M / Z: 514.2 [M+H] + .

[0209] Step 3: Compound 46-2 (150 mg, 0.29 mmol), HATU (133 mg, 0.35 mmol), and triethylamine (59 mg, 0.58 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Intermediate 4 (128 mg, 0.35 mmol) was then added. The mixture was stirred at room temperature for 0.5 h, and water (10 mL) was added. The mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 46-3 (85 mg, yield: 33%) as a yellow solid.

[0210] MS (ESI) M / Z: 864.4 [M+H] + .

[0211] Step 4: Compound 46-3 (85 mg, 0.10 mmol) was added to ethanol (3 mL) at room temperature, and a solution of sodium hydroxide (20 mg, 0.40 mmol) dissolved in water (0.6 mL) was added. The mixture was stirred at room temperature for 24 hours. The pH of the reaction solution was adjusted to acidic with 1N aqueous hydrogen chloride solution, and extracted with DCM (10 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 46-4 (58 mg, yield: 69%) as a yellow solid.

[0212] MS (ESI) M / Z: 836.3 [M+H] + .

[0213] Step 5: Compound 46-4 (58 mg, 0.07 mmol), HATU (31 mg, 0.08 mmol), and triethylamine (21 mg, 0.21 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 46-5 (16 mg, 0.08 mmol) was then added. The mixture was stirred at room temperature for 0.5 h, and water (10 mL) was added. The mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 46 (47 mg, yield: 70%) as a yellow solid.

[0214] MS (ESI) M / Z: 969.4 [M+H] + .

[0215] Example 2: Preparation of (S,Z)-N-(4-aminoiminoylbenzyl)-3-((2-(2-(3-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)pyrrolidin-1-yl)-2-oxoethoxy)ethoxy)ethoxy)methyl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 47)

[0216] Compound 46-4 (24 mg, 0.03 mmol), HATU (15 mg, 0.04 mmol), and triethylamine (15 mg, 0.15 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 47-1 (6 mg, 0.04 mmol) was then added. The mixture was stirred at room temperature for 0.5 h, and water (10 mL) was added. The mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 47 (21 mg, yield: 92%) as a yellow solid.

[0217] MS (ESI) M / Z: 877.41 [M+H] + .

[0218] 1 H NMR(400MHz,DMSO-d6)δ13.68(s,1H),10.91(s,1H),9.27(s,2H),8.93(s,1 H),8.56–8.47m,1H),8.33(s,1H),7.87–7.60(m,4H),7.50(d,J=8.4Hz,2H), 7.43–7.31(m,1H),7.26(s,5H),7.23–7.14(m,1H),6.67(s,1H),6.40–6.37(m,1H),6.24–6.19(m,1H),5.33–5.3 0(m,2H),5.06(s,2H),4.65(s,2H),4.50–4.42(m,4H),3.98(s,2H),3.28–3.24(m,4H),2.42(s,3H),2.41(s,3H).

[0219] Example 3: Preparation of (Z)-N-(4-aminoiminoylbenzyl)-3-(1-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrol-3-yl)-5-methyl-1-oxo-8,11-dioxa-2,5-diazadodec-12-yl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 49)

[0220] Step 1: Under a nitrogen atmosphere, intermediate 2 (148 mg, 1.40 mmol) was added to THF (5 mL), cooled to 0°C, and sodium hydride (56 mg, 1.40 mmol) was added. The mixture was stirred at 0°C for 1 h. Diethylene glycol (300 mg, 0.70 mmol) dissolved in DMF (5 mL) was slowly added dropwise, and the mixture was stirred at room temperature for 16 h. Saturated aqueous ammonium chloride (50 mL) was added, and the THF was removed by concentration under reduced pressure. The mixture was then extracted with DCM (50 mL x 3). The organic phases were combined, washed with water (100 mL) and saturated aqueous sodium chloride (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 49-1 (70 mg, yield: 22%) as a colorless oil.

[0221] MS (ESI) M / Z: 456.2 [M+H] + .

[0222] Step 2: Compound 49-1 (70 mg, 0.15 mmol) and Dess-Martin reagent (98 mg, 0.23 mmol) were added sequentially to DCM (3 mL) and stirred at room temperature for 16 hours. The mixture was filtered, the filtrate was collected, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 49-2 (35 mg, yield: 50%) as a colorless oil.

[0223] MS (ESI) M / Z: 454.2 [M+H] + .

[0224] Step 3: Under nitrogen atmosphere, compound 49-2 (35 mg, 0.08 mmol), compound 49-3 (27 mg, 0.08 mmol, prepared by the same method as Preparation Example 3) and acetic acid (5 mg, 0.08 mmol) were added sequentially to a mixed solution of DCM (2 mL) and methanol (0.2 mL). The temperature was cooled to 0°C, and STAB (34 mg, 0.16 mmol) was added. The mixture was naturally returned to room temperature and stirred for 3 hours. Water (10 mL) was added and extracted with DCM (10 mL × 3). The organic phases were combined, washed with water (20 mL) and saturated sodium chloride solution (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 49-4 (35 mg, yield: 57%) as a yellow solid.

[0225] MS (ESI) M / Z: 794.4 [M+H] + .

[0226] Step 4: Compound 49-4 (35 mg, 0.04 mmol) was added to ethanol (3 mL) at room temperature, followed by a solution of sodium hydroxide (9 mg, 0.22 mmol) dissolved in water (0.6 mL). The mixture was stirred at room temperature for 24 hours. The pH of the reaction mixture was adjusted to acidic with 1N aqueous hydrogen chloride solution, and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to afford compound 49-5 (30 mg, yield: 94%) as a yellow solid.

[0227] MS (ESI) M / Z: 766.3 [M+H] + .

[0228] Step 5: Compound 49-5 (30 mg, 0.04 mmol), HATU (18 mg, 0.05 mmol), and triethylamine (12 mg, 0.12 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. 4-Aminomethylbenzylamine dihydrochloride (compound 47-1) (9 mg, 0.05 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The combined organic phases were dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 49 (20 mg, yield: 55%) as a yellow solid.

[0229] MS (ESI) M / Z: 897.4 [M+H] + .

[0230] Example 4: Preparation of (S,Z)-N-(6-amino-2,4-dimethylpyridin-3-yl)-3-((2-(4-(3-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)pyrrolidin-1-carboxyl)piperazin-1-yl)ethoxy)methyl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 50)

[0231] Intermediate 5 (30 mg, 0.03 mmol), HATU (16 mg, 0.04 mmol), and triethylamine (10 mg, 0.10 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 1-5 (8 mg, 0.04 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The combined organic phases were dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 50 (21 mg, yield: 62%) as a yellow solid.

[0232] MS (ESI) M / Z: 979.5 [M+H] + .

[0233] 1 H NMR(400MHz, CH3OH-d4)δ8.11(s,1H),7.66(dd,J=6.8,2.0Hz,1H),7.55(d,J=2.4Hz,1H),7.52–7.46(m,1H),7.43 –7.39(m,1H),7.30–7.22(m,4H),6.88–6.84(m,2H),6.54–6.50(m,1H),6.47(s,1H),6.38–6.32m,1H),5.36–5.32 (m,1H),5.28(s,2H),5.14(s,2H),4.65(s,2H),4.44(s,3H),3.71–3.57(m,4H),3.51–3.41(m,2H),3.29–3.22(m, 4H),2.59–2.48(m,6H),2.47(s,3H),2.43(s,3H),2.40(s,3H),2.34(s,3H),2.25–2.17(m,2H),2.08–1.98(m,3H).

[0234] Example 5: Preparation of (Z)-N-((6-amino-2,4-dimethylpyridin-3-yl)methyl)-3-(3-(2-((2-(5-((5-fluoro-2-indolin-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)ethyl)amino)-2-oxoethoxy)prop-1-yn-1-yl)-1-(4-((2-pyridinone-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamido (Compound 48)

[0235] Step 1: Under a nitrogen atmosphere, intermediate 11 (500 mg, 1.20 mmol), intermediate 6 (408 mg, 2.40 mmol), cuprous iodide (23 mg, 0.12 mmol), and tetrakistriphenylphosphine palladium (138 mg, 0.12 mmol) were added sequentially to a mixed solution of triethylamine (2.5 mL) and DMF (2.5 mL). The mixture was heated to 100°C and stirred for 3 hours. The mixture was cooled to room temperature, and dichloromethane (25 mL) was added. The mixture was filtered, the filtrate was collected, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (dichloromethane:methanol = 0-100%) to obtain compound 48-2 (500 mg, purity 50%) as a yellow oily liquid, which was used directly in the next step.

[0236] MS (ESI) M / Z: 506.2 [M+H] + .

[0237] Step 2: Compound 48-2 (600 mg, 0.59 mmol) was added to a solution of hydrogen chloride / dioxane (4.0 M, 5 mL). The reaction was allowed to react at room temperature for 1 hour, and the mixture was concentrated under reduced pressure to obtain a residue. The residue was suspended in dichloromethane (50 mL). The pH was adjusted to approximately 8 with saturated aqueous sodium bicarbonate solution, and the mixture was allowed to stand. The layers were separated, and the aqueous phase was extracted with DCM (50 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (dichloromethane:methanol = 0-100%) to obtain compound 48-3 (97 mg, yield: 37%) as a white solid.

[0238] MS (ESI) M / Z: 450.1 [M+H] + .

[0239] Step 3: Compound 48-3 (43 mg, 0.10 mmol), HATU (44 mg, 0.11 mmol), and triethylamine (20 mg, 0.20 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound Int3-5 (18 mg, 0.11 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 48-4 (53 mg, yield: 88%) as a yellow solid.

[0240] MS (ESI) M / Z: 592.3 [M+H] + .

[0241] Step 4: Compound 48-4 (53 mg, 0.09 mmol) was added to ethanol (3 mL) at room temperature, followed by a solution of sodium hydroxide (18 mg, 0.45 mmol) dissolved in water (0.6 mL). The mixture was stirred at room temperature for 48 hours. The pH of the reaction mixture was adjusted to acidic with 1N aqueous hydrogen chloride solution, and extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 48-5 (47 mg, yield: 94%) as a yellow solid.

[0242] MS (ESI) M / Z: 563.2 [M+H] + .

[0243] Step 5: Compound 48-5 (47 mg, 0.08 mmol), HATU (38 mg, 0.10 mmol), and triethylamine (24 mg, 0.24 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 1-5 (19 mg, 0.10 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 48-6 (50 mg, yield: 89%) as a yellow solid.

[0244] MS (ESI) M / Z: 697.3 [M+H] + .

[0245] Step 6: Compound 48-6 (50 mg, 0.07 mmol) was added to a solution of hydrogen chloride / dioxane (4.0 M, 5 mL), reacted at room temperature for 1 hour, and concentrated under reduced pressure to obtain compound 48-7 (42 mg, yield: 100%) as a white solid.

[0246] MS (ESI) M / Z: 597.3 [M+H] + .

[0247] Step 7: Compound 48-7 (42 mg, 0.07 mmol), HATU (32 mg, 0.08 mmol), and triethylamine (21 mg, 0.24 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound Int3-4 (24 mg, 0.08 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 48 (20 mg, yield: 33%) as a yellow solid.

[0248] MS (ESI) M / Z: 879.4 [M+H] + .

[0249] 1 H NMR(400MHz,DMSO-d6)δ13.67(s,1H),10.89(s,1H),8.27(s,1H),7.96(t, J=5.2Hz,1H),7.87(s,1H),7.78–7.73(m,2H),7.71(s,1H),7.68–7.61(m,1H),7.42-7 .37(m,1H),7.27–7.20(m,5H),6.96–6.88(m,1H),6.86–6.80(m,1H),6.38(d,J=10.4H z,2H),6.24–6.16(m,1H),5.28(s,2H),5.05(s,2H),4.42(s,2H),4.24(d,J=4.8Hz,2H ),3.99(s,2H),3.32–3.27(m,4H),2.43(s,3H),2.41(s,3H),2.39(s,3H),2.26(s,3H).

[0250] The compounds listed in the table below were prepared using procedures similar to those described in Examples 1, 2, 3, 4 and 5 using appropriate starting materials.

[0251] Table 1.

[0252] Example 6: Preparation of (Z)-N-(4-aminoiminoylbenzyl)-3-(3-(2-(2-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)ethyl)amino)-2-oxoethoxy)prop-1-yn-1-yl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 60)

[0253] Step 1: Intermediate 3 (97 mg, 0.22 mmol), HATU (99 mg, 0.26 mmol), and triethylamine (67 mg, 0.67 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 48-2 (85 mg, 0.22 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 60-1 (110 mg, yield: 65%) as a yellow solid.

[0254] MS (ESI) M / Z: 774.3 [M+H] + .

[0255] Step 2: Compound 60-1 (105 mg, 0.14 mmol) was added to ethanol (3 mL) at room temperature, followed by a solution of sodium hydroxide (27 mg, 0.68 mmol) dissolved in water (0.6 mL). The mixture was stirred at room temperature for 48 hours. The pH of the reaction mixture was adjusted to acidic with 1N aqueous hydrogen chloride solution, and extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 60-2 (24 mg, yield: 23%) as a yellow solid.

[0256] MS (ESI) M / Z: 746.3 [M+H] + .

[0257] Step 3: Compound 60-2 (24 mg, 0.03 mmol), HATU (15 mg, 0.04 mmol), and triethylamine (9 mg, 0.09 mmol) were added sequentially to DMF (1 mL) at room temperature and stirred at room temperature for 0.5 h. Compound 2-1 (9 mg, 0.04 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 60 (21 mg, yield: 75%) as a yellow solid.

[0258] MS (ESI) M / Z: 877.3 [M+H] + .

[0259] 1 H NMR(400MHz,DMSO-d6)δ13.68(s,1H),10.91(s,1H),9.27(s,2H),8.93(s,1H),8.55–8.50(m,1H),8 .33(s,1H),7.87–7.60(m,4H),7.50(d,J=8.4Hz,2H),7.43–7.31(m,1H),7.26(s,5H),7.23–7.14(m ,1H),6.86–6.81(m,1H),6.67(s,1H),6.40–6.37(m,1H),6.24-6.19(m,1H),5.32–5.31(m,2H),5.0 6(s,2H),4.65(s,2H),4.50–4.42(m,4H),3.98(s,2H),3.27–3.24(m,4H),2.42(s,3H),2.41(s,3H).

[0260] Example 7: Preparation of (S,Z)-N-(4-aminoiminoylbenzyl)-3-((2-(4-(3-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)pyrrolidin-1-carboxyl)piperazin-1-yl)ethoxy)methyl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 50)

[0261] Intermediate 5 (30 mg, 0.03 mmol), HATU (16 mg, 0.04 mmol), and triethylamine (10 mg, 0.10 mmol) were added sequentially to DMF (2 mL) at room temperature and stirred at room temperature for 0.5 h. 4-Aminomethylbenzylamine dihydrochloride (Compound 47-1) (9 mg, 0.04 mmol) was added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The combined organic phases were dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford Compound 50 (19 mg, yield: 56%) as a yellow solid.

[0262] MS (ESI) M / Z: 977.4 [M+H] + .

[0263] 1 H NMR(400MHz,CH3OH-d4)δ8.15(s,1H),7.79–7.75(m,2H),7.69–7.65(m,1H),7.59–7.54(m,3H),7.52–7.47(m,1H),7 .43–7.39(m,1H),7.31–7.25(m,4H),6.89–6.84(m,2H),6.55–6.50(m,1H),6.38–6.33(m,1H),5.36–5.33(m,1H),5. 31(s,2H),5.15(m,2H),4.70(s,2H),4.63(s,2H),4.47–4.40(m,1H),3.68–3.56(m,4H),3.50–3.37(m,2H),3.18–3. 11(m,4H),2.55(t,J=4.8Hz,2H),2.43(s,3H),2.40(s,3H),2.38–2.36(m,2H),2.22–2.16(m,2H),2.05–2.00(m,2H).

[0264] Example 8: Preparation of (S,Z)-N-(4-aminoiminoylbenzyl)-3-(3-((1-(3-(5-((5-fluoro-2-oxoindole-3-ylidene)methyl)-2,4-dimethyl-1H-pyrrole-3-carboxamido)pyrrolidin-1-carboxyl)piperidin-4-yl)oxy)prop-1-yn-1-yl)-1-(4-((2-oxopyridin-1(2H)-yl)methyl)benzyl)-1H-pyrazole-4-carboxamide (Compound 61)

[0265] Step 1: Under a nitrogen atmosphere, intermediate 11 (500 mg, 1.20 mmol), intermediate 7 (574 mg, 2.40 mmol), cuprous iodide (23 mg, 0.12 mmol), and tetrakistriphenylphosphine palladium (138 mg, 0.12 mmol) were added sequentially to a mixed solution of triethylamine (2.5 mL) and DMF (2.5 mL). The mixture was heated to 100° C. and stirred for 3 hours. The mixture was cooled to room temperature, and dichloromethane (25 mL) was added. The mixture was filtered, the filtrate was collected, and concentrated under reduced pressure to obtain a residue. The residue was separated by column chromatography (dichloromethane:methanol = 0-100%) to obtain compound 61-1 (470 mg, 68%) as a yellow oily liquid, which was used directly in the next step.

[0266] MS (ESI) M / Z: 575.3 [M+H] + .

[0267] Step 2: Compound 61-1 (470 mg, 0.82 mmol) was added to a solution of hydrogen chloride / dioxane (4.0 M, 5 mL) and allowed to react at room temperature for 1 hour. The mixture was concentrated under reduced pressure to obtain a residue, which was suspended in dichloromethane (50 mL) and adjusted to a pH of approximately 8 with saturated aqueous sodium bicarbonate. The mixture was allowed to stand for 2 minutes. The layers were separated and the aqueous phase was extracted with DCM (50 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (dichloromethane:methanol = 0-100%) to obtain compound 61-2 (237 mg, yield: 65%) as a white solid.

[0268] MS (ESI) M / Z: 447.2 [M+H] + .

[0269] Step 3: Under nitrogen atmosphere, compound 61-2 (178 mg, 0.4 mmol) was dissolved in DCM (2 mL), cooled to 0°C, and a solution of triphosgene (0.1 mmol) dissolved in DCM (4 mL) was added dropwise. The mixture was stirred at 0°C for 30 minutes, and intermediate 4 (147 mg, 0.2 mmol) dissolved in DMF (2 mL) was added dropwise. The mixture was allowed to return to room temperature and react for 1 hour. Water (20 mL) was added, and the mixture was extracted with DCM (20 × 3). The organic phases were combined, washed with water (30 mL) and saturated sodium chloride solution (30 mL), dried over anhydrous sulfuric acid, filtered, and concentrated under reduced pressure to obtain a residue. The residue was separated and purified by column chromatography (methanol:dichloromethane = 0-100%) to obtain compound 61-3 (191 mg, yield: 55%) as a yellow solid.

[0270] MS (ESI) M / Z: 869.4 [M+H] + .

[0271] Step 4: Compound 61-3 (190 mg, 0.22 mmol) was added to ethanol (3 mL) at room temperature, followed by a solution of sodium hydroxide (46 mg, 1.14 mmol) dissolved in water (1 mL). The mixture was stirred at room temperature for 24 hours. The pH of the reaction mixture was adjusted to acidic with 1N aqueous hydrogen chloride solution, and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to afford compound 61-4 (111 mg, yield: 60%) as a yellow solid.

[0272] MS (ESI) M / Z: 839.3 [M+H] - .

[0273] Step 5: Compound 61-4 (50 mg, 0.06 mmol), HATU (32 mg, 0.08 mmol), and triethylamine (20 mg, 0.20 mmol) were added sequentially to DMF (4 mL) at room temperature and stirred at room temperature for 0.5 h. 4-Aminomethylbenzylformamidine dihydrochloride (compound 47-1) (18 mg, 0.08 mmol) was then added and stirred at room temperature for 0.5 h. Water (10 mL) was added and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a residue. The residue was isolated and purified by column chromatography (methanol:dichloromethane = 0-100%) to afford compound 61 (31 mg, yield: 53%) as a yellow solid.

[0274] MS (ESI) M / Z: 972.4 [M+H] + .

[0275] The compounds listed in the table below were prepared using procedures similar to those described in Examples 6, 7, and 8 using appropriate starting materials.

[0276] Table 2.

[0277] Test Example 1: In vitro inhibition test of the compound of the present invention on plasma kallikrein (PKa)

[0278] The ability of compounds of the invention to inhibit plasma kallikrein (PKK) was determined in the presence of 1% (v / v) DMSO using the following biochemical assay in assay buffer (100 mM Tris, 150 mM NaCl, adjusted to pH 7.8 with HCl, containing 0.1% (w / v) BSA and 0.05% (v / v) Tween 20): Compounds were tested for inhibition of PKa enzymatic activity using the FI method. PKK (R&D, Cat#2497-SE) and Pro-Phe-Arg-AMC (Nanjing Peptide Industry, Cat#NJP23946) solutions were prepared in reaction buffer (50 mM Tris-HCl, pH 7.5, 250 mM NaCl). The final concentrations of the enzyme protein (R&D, Cat#2497-SE) and Pro-Phe-Arg-AMC in the reaction mixture were 10 nM and 60 μM, respectively. The positive reference substance Avoralstat (MCE, Cat#HY-16735) had a starting concentration of 100 nM, 3-fold dilution, and 10+0 dose. 0.05 μL of the compound in 100% DMSO was delivered to a 384-well plate by acoustic liquid delivery technology (Echo665; nanoliter range). The compound was tested in duplicate wells. Transfer 5 μL of 2× enzyme solution to a 384 reaction plate and centrifuge at 1000 rpm for 1 minute, incubate at 25°C for 15 minutes; transfer 5 μL of 2× Pro-Phe-Arg-AMC solution to a 384 reaction plate, centrifuge at 1000 rpm for 1 minute, and incubate at 25°C for 3 hours. Finally, fluorescence was measured using BMG and an Envision reader with an excitation wavelength set to 355 nm and an emission wavelength set to 460 nm, and the FI signal (ex380 / em460) was read. IC was obtained using GraphPad Prism software. 50 Values ​​and nonlinear regression curve fitting.

[0279] These data were fitted to produce the best-fit IC 50 The results of these measurements are shown in Table 2 below, with the numbers corresponding to the compound numbers.

[0280] Test Example 2: Evaluation of the inhibition of PKa by the compounds of the present invention in kaolin-activated human PPP

[0281] Platelet-poor plasma (PPP) obtained from human whole blood and anticoagulated with sodium citrate was incubated with various concentrations of the test compound and 25, 75, 250, or 750 μg / mL kaolin in assay buffer for 20 minutes at 37°C to obtain a concentration response of the test compound for each kaolin dose used. The positive drug, Avoralstat (MCE, Cat#HY-16735), was diluted to the reference concentration and diluted three-fold using the dilution as the first point.

[0282] 50 nL of compound was then transferred to a 384-well assay plate, with each well containing two replicates. The 384-well assay plate was centrifuged at 1000 rpm. 2.5 μL of human plasma working solution was added to each assay well. The 384-well assay plate was centrifuged at 1000 RPM. 2.5 μL of kaolin working solution was added to each assay well. The 384-well assay plate was centrifuged at 1000 RPM and incubated at 25°C for 20 minutes. 5 μL of Pro-Phe-Arg-AMC working solution was added to each assay well. The 384-well assay plate was centrifuged at 1000 RPM and incubated at 25°C for 60 minutes. Fluorescence signals were read by BMG.

[0283] The inhibition percentage of the compound was calculated according to the following formula:

[0284] Compound (inhibition %) = 100 × (average of negative control well values ​​- compound well values) / (average of negative control well values ​​- average of positive control well values)

[0285] These data were fitted to produce the best-fit IC 50 The results of these measurements are shown in Table 2 below, with the numbers corresponding to the compound numbers.

[0286] Test Example 3: In vitro VEGFR2 and PDGFRβ inhibitory activity test of compounds

[0287] The test compound was vortexed and thoroughly mixed, and then diluted with DMSO (Cat. No.: D4540, Manufacturer: Sigma) to 100× the detection concentration. Transfer 50 nL of compound to a 384-well assay plate (Cat. No. 784075, Greiner) using a 655SYSTEM. Prepare a 2x kinase solution using 1x kinase reaction buffer (1x Buffer, 5 mM MgCl2, 1 mM DTT, 1 mM MnCl2). Transfer 2.5 μL of VEGFR2 (0.07 nM, Carna, Cat. No. 08-191) solution to the 384-well plate. Centrifuge at 1000 rpm for 60 seconds using a microplate centrifuge (Plate pro 3200, Monad). Incubate in a biochemical incubator (LRH-250F, BluePard) at 25°C for 10 minutes. Prepare a 2× mixture of substrate (TK: 1 μM) and ATP (4 μM, manufacturer: Promega, catalog number: V915B) using kinase reaction buffer. Add 2.5 μL of the substrate and ATP mixture to the reaction plate to start the reaction. Centrifuge at 1000 rpm for 1 minute. Incubate at 25°C for 50 minutes. Add 5 μL of kinase detection reagent to each well of the reaction plate, centrifuge at 1000 rpm for 1 minute, and incubate at 25°C for 60 minutes. Use a microplate reader (manufacturer: BMG, model: PHERAstar FSX) to read the fluorescence signals at 620 nm (Cryptate) and 665 nm (XL665). Each reaction was tested in duplicate, and IC was analyzed using GraphPad Prism 8.0 software. 50 .

[0288] The test compound was vortexed and thoroughly mixed, and then diluted with DMSO (Cat. No.: D4540, Manufacturer: Sigma) to 100× the detection concentration. 655 SYSTEM) transfer 50 nL of compound to a 384 assay plate (Cat. No. 784075, Greiner). Prepare a 2x kinase solution using 1x kinase reaction buffer (1x Buffer, 5 mM MgCl2, 1 mM DTT, 1 mM MnCl2, 12.5 nM SEB). Transfer 2.5 μL of PDGFRβ (0.23 nM, Carna, Cat. No. 08-158) solution to the 384 reaction plate. Centrifuge at 1000 rpm for 60 seconds using a microplate centrifuge (Model: Plate pro 3200, Monad). Incubate in a biochemical incubator (Model: LRH-250F, Bluepard) at 25°C for 10 minutes. Prepare a 2× mixture of substrate (TK: 1 μM) and ATP (1 μM, manufacturer: Promega, catalog number: V915B) using kinase reaction buffer. Add 2.5 μL of the substrate and ATP mixture to the reaction plate to start the reaction. Centrifuge at 1000 rpm for 1 minute. Incubate at 25°C for 50 minutes. Add 5 μL of kinase detection reagent to each well of the reaction plate, centrifuge at 1000 rpm for 1 minute, and incubate at 25°C for 60 minutes. Use a microplate reader (manufacturer: BMG, model: PHERAstar FSX) to read the fluorescence signals at 620 nm (Cryptate) and 665 nm (XL665). Each reaction was tested in duplicate, and IC was analyzed using GraphPad Prism 8.0 software. 50 .

[0289] These data were fitted to produce the best-fit IC 50 The results of these measurements are shown in Table 3 below, with the numbers corresponding to the compound numbers.

[0290] Table 3.

[0291] The above results indicate that the compounds of the present invention have a strong inhibitory effect on plasma kallikrein (PKa), VEGFR2 and PDGFRβ.

[0292] It will be appreciated by those skilled in the art that the present disclosure is not limited to the foregoing illustrative embodiments but can be embodied in other specific forms without departing from its essential attributes. It is therefore intended that the present disclosure be considered in all respects as illustrative and not restrictive, and that reference to the appended claims is made to the embodiments other than the foregoing embodiments, and all variations that come within the meaning and range of equivalents of the claims are intended to be embraced herein.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof: in, L1 is selected from L2 is absent or selected from a linear -(CH2) 1-22 -group, in which the straight chain -(CH2) 1-22 - one or more -(CH2)- in the group are optionally and independently replaced by one or more selected from -C(O)-, -O-, -(NR 2 )-, -S-, -(C=C)-, -(C≡C)-, a divalent cycloalkyl group, a divalent aryl group, a divalent heterocyclic group, and a divalent heteroaryl group, and L2 is optionally substituted by 1, 2 or 3 R 1 replace; Each R 1 independently selected from C1-C6 alkyl, halogenated C1-C6 alkyl, C1-C6 alkoxy, halogenated C1-C6 alkyl, halogen, cyano and hydroxyl, Each R 2 independently selected from hydrogen, C1-C6 alkyl and acyl, wherein the C1-C6 alkyl is optionally substituted by 1, 2 or 3 R 1 replace; The group Ar1 is selected from -(CH2)-aryl, -(CH2)-heteroaryl and Wherein ring a is a 5-6 membered heterocyclic ring, the aryl, heteroaryl and Optionally, one or more selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, cyano, C1-C6 alkoxy, halo C1-C6 alkoxy, amino, hydroxyl, The group substitution; The group Ar2 is selected from -C1-C6 alkyl, -C1-C6 haloalkyl, cycloalkyl, aryl and heteroaryl, wherein the group Ar2 is optionally substituted by one or more selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 cycloalkyl, C1-C6 cycloalkylalkyl, 5-6 membered heterocyclyl and substituted by a group, a 5-6 membered heterocyclic group or Optionally substituted with one or more halogens.

2. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, which is a compound of formula (II) or a pharmaceutically acceptable salt thereof: in, X is selected from oxygen atoms, NR 2 ; L2, R 2 , group Ar1 and group Ar2 are as defined in claim 1.

3. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1, which is a compound of formula (III) or a pharmaceutically acceptable salt thereof: in, L2, the group Ar1 and the group Ar2 are as defined in claim 1.

4. A compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, wherein: Ar1 is selected from -(CH2)-phenyl, -(CH2)-pyridyl and Wherein ring a is a 5-6 membered heteroaryl group, preferably imidazolyl or pyrazolyl, more preferably for The phenyl, pyridyl and Optionally, one or more selected from C1-C6 alkyl, C1-C6 haloalkyl, halogen, cyano, C1-C6 alkoxy, halo C1-C6 alkoxy, amino, hydroxyl, The group substitution; R is selected from hydrogen and C1-C6 alkyl.

5. A compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, wherein: Ar2 is selected from C 6-10 Aryl is preferably phenyl, 5 to 10 membered heteroaryl, -C1-C6 alkyl, -C1-C6 haloalkyl, C3-C6 cycloalkyl, wherein the C 6-10 Aryl, 5 to 10 membered heteroaryl, optionally substituted by one or more selected from halogen, cyano, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 cycloalkyl, C1-C6 cycloalkylalkyl, 5-6 membered heterocyclyl and The 5-6 membered heterocyclic group or Optionally substituted with one or more halogens.

6. A compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof, wherein L2 is selected from C 1-2 Alkylene; Alternatively, L2 is selected from -(CH2) t -(C=C)-* and -(CH2) t -(C≡C)-*; wherein t is an integer from 0 to 6, preferably an integer from 1 to 4, more preferably 1 or 2; * is the position of connection to the pyrazole ring; Alternatively, L2 is selected from in, X1, X2, X3, and X4 are each independently selected from a bond, -O-, -(NR 2 )-, -C(O)NH-, -NHC(O)-, -S-, -(C=C)-, -(C≡C)-, a 4-6 membered divalent heterocyclic group, a 3-6 membered divalent cycloalkyl group, a 5-10 membered divalent heteroaryl group, and a 6-10 membered divalent aryl group, wherein the 4-6 membered divalent heterocyclic group, the 3-6 membered divalent cycloalkyl group, the 5-10 membered divalent heteroaryl group, or the 6-10 membered divalent aryl group is optionally substituted by 1, 2, or 3 R 1 Replace; each R 2 independently selected from hydrogen, C1-C6 alkyl and acyl, wherein the C1-C6 alkyl is optionally substituted by 1, 2 or 3 R 1 Replace, each R 1 Independently selected from C1-C6 alkyl, halogenated C1-C6 alkyl, C1-C6 alkoxy, halogenated C1-C6 alkyl, halogen, cyano and hydroxyl; t1, t2, t3, t4, t5 are each independently an integer from 0 to 6; * is the position connected to the pyrazole ring.

7. A compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 6, wherein: The divalent heterocyclic group is selected from: The divalent heteroaryl group is selected from:

8. A compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7, wherein: -L2-Selected from: Wherein: G is a 5-6 membered heteroaryl group, preferably More preferred G1 and G2 are each independently selected from a 4-6 membered heterocyclic group, preferably p is 1 or 2; s is an integer from 0 to 6, preferably 0 or 1; s1 is an integer from 1 to 10; s2 is 1 or 2, preferably 1; s3 and s4 are each independently selected from an integer of 1 to 6, preferably an integer of 1 to 4, more preferably 1 or 2; v is an integer from 0 to 4, preferably 0 or 1; * is the position of connection to the pyrazole ring.

9. A compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 8, selected from: or a pharmaceutically acceptable salt thereof.

10. A pharmaceutical composition comprising a compound of formula (I) according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.

11. Use of the compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 9 or the pharmaceutical composition according to claim 10 in the preparation of a medicament for treating or preventing vascular disease-related diseases.

12. The use according to claim 11, wherein the vascular disease-related disorder is selected from diabetic retinopathy (DR), diabetic macular edema (DME), retinal vein occlusion (RVO), central retinal vein occlusion (CRVO), macular degeneration, wet age-related macular degeneration (wet AMD), retinopathy of prematurity (ROP), neovascular glaucoma, retinitis pigmentosa (RP), proliferative and non-proliferative retinopathy, retinal angiomatous proliferation, macular telangiectasia, ischemic retinopathy, iris neovascularization, intraocular neovascularization, corneal neovascularization, retinal neovascularization, polypoidal choroidal vasculopathy (PCV), choroidal neovascularization. The invention relates to a retinal vascular disease and a retinal degeneration, in particular selected from diabetic retinopathy (DR), diabetic macular edema (DME), retinal vein occlusion (RVO), central retinal vein occlusion (CRVO), wet age-related macular degeneration (wet AMD), in particular selected from hereditary angioedema (HAE) and brain edema after stroke, in particular selected from diabetic complications of retinal vascular permeability associated with diabetic retinopathy and diabetic macular edema, preferably selected from diabetic macular edema (DME), wet age-related macular degeneration (AMD), non-exudative choroidal neovascularization (CNV), hereditary angioedema (HAE) and brain edema after stroke.

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