CN113336774B

CN113336774B - Substituted chiral diaryl macrocyclic compounds as TRK inhibitors

Info

Publication number: CN113336774B
Application number: CN202110712429.3A
Authority: CN
Inventors: 范为正; 张永杰; 唐春雷; 刘彦; 邵俊兰; 丁若洋; 范懿庆
Original assignee: Nanjing Leizheng Pharmaceutical Technology Co ltd; Jiangnan University
Current assignee: Nanjing Leizheng Pharmaceutical Technology Co ltd; Jiangnan University
Priority date: 2021-06-25
Filing date: 2021-06-25
Publication date: 2023-05-23
Anticipated expiration: 2041-06-25
Also published as: CN113336774A

Abstract

The present invention relates to substituted chiral diaryl macrocyclic compounds as TRK inhibitors, belonging to the class of compounds. The chiral diaryl macrocyclic compound has a structure shown in a general formula (I), and has good and specific TRK family protein tyrosine kinase inhibition activity.

Description

Substituted chiral diaryl macrocyclic compounds as TRK inhibitors

Technical Field

The present invention is in the field of chemical medicine and relates to substituted chiral diaryl macrocyclic compounds as TRK inhibitors.

Background

Three members of the tropomyosin receptor kinase (tropomyosin receptor kinase, TRK) family TRKA (NTRK 1 encoded), TRKB (NTRK 2 encoded) and TRKC (NTRK 3 encoded) belong to the receptor tyrosine kinases (receptor tyrosine kinase, RTKs) and are transmembrane proteins. These kinases are expressed primarily in neuronal tissue and are activated by neurotrophic factors, playing an important role in regulating cell survival, proliferation, fate of neural precursors, growth and pattern formation of axons and dendrites; neurotrophins are specific ligands, namely the Nerve Growth Factor (NGF) ligand of TRKA, the brain-derived growth factor (BDGF) ligand of TRKB, and the NT-4/5 ligand, the NT3 ligand of TRKC, respectively. (Annual Review of Biochemistry, 2003.72:609-642).

The TRK receptor family plays an important role in the growth and development of neurons, and the rearranged state may drive malignancy. The chimeric TRK proteins produced by gene fusion either result in overexpression of the kinase domain or have constitutive activity of the kinase function. This rearrangement, whether by point mutation, chromosomal rearrangement and gene fusion, or gene deletion, results in spontaneous ligand independent dimerization and subsequent activation of signal transduction (Cancer Cell,2004.6 (4): 347-360). NTRK rearrangements and fusions were found in a variety of tumor types with different but overall lower proportions of cases. Among the various molecular mechanisms of TRK activation, NTRK gene chromosomal translocation is the most studied (Esmo Open,2016.1 (2); cancer Discovery,2015.5 (1): 25-34). Typically, the 3 'region of the NTRK gene is linked to the 5' region of the chaperone gene due to chromosomal rearrangements (Pharmacology & Therapeutics, 2017.173:58-66). NTRK gene fusion, point mutations and in-frame deletion mutations produce a ligand independent constitutively activated protein with oncogenic potential (Molecular and cellular biology,1990.10 (8): 4202-4210;Journal of Thoracic Oncology,2015.10 (12): 1670-1674). Because of the structural or functional deletions of the extracellular domains, these genetic abnormalities are difficult to regulate by antibody therapy or the like (Annual Review of Neuroscience, 2003.26:299-330), and small molecule inhibitors have become a promising approach (Proceedings of the National Academy of Sciences of the United States of America,1987.84 (19): 6707-6711; oncogene,1998.16 (6): 809-816).

Larotrib (Larotrectinib) was developed by Loxo Oncology in concert with Bayer AG for the treatment of adult and pediatric cancer patients with NTRK gene fusion; it is a highly selective, potent TRKA, TRKB and TRKC inhibitor (50% inhibition constant 5-11 nmol/L in vitro), almost inactive against the activity of other kinases and non-kinase targets, belonging to the first generation of TRK inhibitors (Journal of Medicinal Chemistry,2019.62 (4): 1731-1760; drugs,2019.79 (2): 201-206) that are ATP-competitive and bind to the ATP binding site. The compound can induce cell cycle arrest in cell G1 phase and apoptosis of KM-12 cells (Nature Medicine,2013.19 (11): 1469- +). First global approval was obtained for larotinib at day 26, 11, 2018, and phase 1 and phase 2 clinical trials were conducted in adult and pediatric patients in the united states; screening conditions for these patients included: solid tumors with NTRK gene fusions without acquired drug resistance mutations, metastasis or surgical excision may lead to serious morbidity, with no satisfactory replacement therapy or progression after treatment (Journal of Medicinal Chemistry,2019.62 (4): 1731-1760).

Entrictinib (Entrectrinib) is a potent multi-target ALK, ROS1 and TRKs inhibitor, with nanomolar activity on the corresponding target oncogenic driver cell line; was originally developed as an ALK inhibitor, and was subsequently found to also have ROS1 and TRK inhibitory activity (Molecular Cancer Therapeutics,2016.15 (4): 628-639;Journal of Medicinal Chemistry,2019.62 (17): 8364-8364). It exhibits good pharmacokinetic profiles in both rodents and non-rodents (Journal of Medicinal Chemistry,2019.62 (17): 8364-8364). Entrictinib was selected for development based on its allowable safety and tolerability, was used in phase I/II clinical trials to treat ALK-, ROS 1-and TRKs-dependent tumor patients, and showed a significant efficacy (British Journal of Cancer,2015.113 (12): 1730-1734; jnci-Journal of the National Cancer Institute,2016.108 (1)).

Secondary drug resistance mutations at the front of TRK solvents, such as the G595R and G667C mutations of TRKA, the G623R mutation of TRKC, have been shown to produce acquired drug resistance in patients and preclinical models by interfering with the binding of laratinib or emtrictinib to the ATP pocket (Cancer Discovery,2016.6 (1): 36-44;Cancer Discovery,2017.7 (9): 934-936;Cancer Discovery,2017.7 (9): 963-972). Similar secondary mutations in other kinase fusion paradigms, such as ALK fusion positive lung cancer and FGFR2 fusion positive cholangiocarcinoma, have been shown to lead to acquired resistance, but next generation kinase inhibitors can be developed to overcome these changes (Clinical Cancer Research,2011.17 (8): 2081-2086;Cancer Discovery,2017.7 (3): 252-263).

Selitetinib (seliteretinib) is a novel, selective TRK TKI aimed at overcoming acquired resistance mediated by kinase domain (solvent front and xDFG) mutations. The activity of these acquired mutations was demonstrated in enzyme and cell-based assays and in vivo tumor models. Two cases of TRK fusion positive cancer patients have developed acquired drug resistance mutations when using Larotigotine therapy, and researchers have used rapid dose determination under pharmacokinetic assessment guidance, first using treatment with selatinib on a human basis. In both cases, this approach resulted in a rapid tumor response and prolonged the overall duration of disease control achieved by TRK inhibition. The next generation TRK kinase inhibitor, celetinib, was identified as having potent and selective activity against all three TRK kinases, their fusions and acquired resistance mutations, both preclinical and in patients (Cancer Discovery,2017.7 (9): 963-972).

Lopatinib (reportectrinib) is a rationally designed low molecular weight macrocyclic kinase inhibitor, selective and potent for wild-type ROS1, TRKA-C, and ALK. More importantly, lopatinib exhibits inhibitory activity against a variety of solvent front mutated kinases in vivo and in vitro, and also potently inhibits other clinically relevant non-solvent front mutations. As clinical evidence, in one phase I/II clinical trial, lopatinib obtained a defined response in patients with ROS1 or NTRK3 fusion positive cancers that relapsed due to solvent front substitution mediated resistance to the prior generation TKIs (Cancer Discovery,2018.8 (10): 1227-1236).

In addition, inhibition of the neurotrophic factor/TRK pathway has been shown to be effective in treating inflammatory diseases in preclinical modes. For example, inhibition of the neurotrophic factor/Trk pathway is associated with preclinical patterns of the following diseases: inflammatory lung diseases including asthma (Pharmacology & Therapeutics,2008,117 (1): 52-76), interstitial cystitis (The Journal of Urology,2005,173 (3): 1016-1021), inflammatory bowel diseases including ulcerative colitis and Crohn's disease (Gut, 2000,46 (5): 670-678), and inflammatory skin diseases such as atopic dermatitis (Archives of Dermatological Research,2006,298 (1): 31-37), eczema and psoriasis (Journal of Investigative Dermatology,2004,122 (3): 812-819).

The neurotrophic factor/TRK pathway, and in particular the BDNF/TRKB pathway, is also implicated in the etiology of neurodegenerative diseases, including multiple sclerosis, parkinson's disease and Alzheimer's disease (Frontiers in Neuroendocrinology,2006,27 (4): 404-414). Modulation of the neurotrophic factor/Trk pathway may be useful in the treatment of these and related diseases.

Studies have shown that the TRKA receptor is critical for the disease process of trypanosoma cruzi (Chagasdisease) in infection by parasitic infection in human hosts (Cell Host & Microbe,2007,1 (4): 251-261). Thus, inhibition of TRKA may be useful in the treatment of chagas' disease and related protozoal infections.

Meanwhile, the TRK inhibitor can also be used for treating diseases related to imbalance of bone remodeling regulation, such as osteoporosis, rheumatoid arthritis and bone metastasis. Bone metastasis is a frequent complication of cancer that can occur in up to 70% of patients with advanced breast or prostate cancer and in about 15 to 30% of patients with lung, colon, stomach, bladder, uterine, rectal, thyroid or kidney cancer. Osteolytic metastasis can cause severe pain, pathological fractures, life-threatening hypercalcemia, spinal cord compression and other nerve compression syndromes. Therefore, drugs that induce apoptosis in proliferative osteoblasts would be highly advantageous. The expression of TRKA receptors and TRKC receptors has been observed in the osteogenic region of the fracture mouse model (Bone, 2000,26 (6): 625-633). In addition, NGF distribution was observed in almost all osteoblasts. pan-TRK inhibitors inhibit tyrosine signaling activated by neurotrophins binding to all 3 TRK receptors in human hFOB osteoblasts. These data support the theory of using TRK inhibitors to treat bone remodeling diseases (e.g., bone metastasis in cancer patients).

The lopatinib is used as a multi-target inhibitor, and has strong inhibition effect on wild ALK and ROS1 except TRK. The physiological function of ALK has not been fully elucidated as reported in the literature, but some evidence has demonstrated the regulatory role of ALK in normal development and function of the central and peripheral nervous systems (Oncogene 1997, 14:439-449). There is also evidence that ROS1 may be involved in epithelial-to-mesenchymal transition of mammalian kidneys, lungs, heart, intestines and testes (Biochim Biophys Acta,2009;1795:37-52; EMBOJ,1991,10:3693-3702;Development 1992,115:11-20). The existing strong TRK inhibitor has poor selectivity, can not effectively realize specific inhibition of TRK, and can also generate obvious inhibition effect on some non-kinase targets, thereby bringing about obvious side effects.

Disclosure of Invention

In order to solve the above problems in the prior art, the present invention provides a compound having the general formula (I) (including stereoisomers and tautomers thereof) or pharmaceutically acceptable salts thereof, a pharmaceutical composition and uses thereof. The compound of the invention has better specificity, good pharmaceutical performance and metabolic stability.

The present invention first provides a compound having the general formula (I) (including stereoisomers and tautomers thereof) or a pharmaceutically acceptable salt thereof,

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Wherein:

M ¹ m and M ² Independently selected from CH, CD or N;

X ¹ x is X ² Independently selected from S or O;

each R is ¹ 、R ² 、R ³ R is R ⁴ Independently selected from H, deuterium, methoxy or (1-3C) alkyl; wherein each hydrogen atom in the (1-3C) alkyl group is optionally independently substituted with: deuterium, halogen, methoxy or CN;

R ⁵ selected from H, deuterium or (1-3C) alkyl; wherein each hydrogen atom in the (1-3C) alkyl group is optionally independently substituted with: deuterium, halogen or CN;

a is-R ⁶ CH-, or-R ⁶ CHR ^6’ CH-, or-R ⁶ CHR ^6’ CHR ^6” CH-、-R ⁶ CHR ^6’ CHR ^6” CHR ^6”’ CH-；R ⁶ 、R ^6’ 、R ^6” R is R ^6”’ Each independently selected from H, deuterium, methoxy, substituted or unsubstituted (1-2C) alkyl; by substituted is meant that each hydrogen atom in the (1-2C) alkyl group is optionally independently substituted with: deuterium, halogen, methoxy or CN;

R ⁷ selected from halogen, CN, methoxy, substituted or unsubstituted (1-2C) alkyl; by substituted is meant that each hydrogen atom in the (1-2C) alkyl is optionally independently substituted with: deuterium, halogen, methoxy or CN.

In one embodiment of the invention, R ⁵ Selected from H or deuterium.

In one embodiment of the invention, R ⁷ Selected from halogen (flurochlorinated bromine iodine) or CN.

At the bookIn one embodiment of the invention, A is-R ⁶ CHR ^6’ CH-, or-R ⁶ CHR ^6’ CHR ^6” CH-。

In one embodiment of the invention, R ¹ 、R ² 、R ³ R is R ⁴ Independently selected from H, deuterium, or methyl;

Preferably:

in certain embodiments of the invention, M ¹ M and M ² CH.

In certain embodiments of the invention, X ¹ X is X ² Is O.

In certain embodiments of the invention, R ¹ 、R ² 、R ³ R is R ⁴ Selected from H or methyl. Specific examples, R ¹ 、R ² 、R ³ R is R ⁴ At least one of which is methyl, the inventors have found that such groups result in better kinase inhibitory activity due to more favourable hydrophobic interactions.

In certain embodiments of the invention, R ⁵ Selected from H.

In certain embodiments of the invention, A is-R ⁶ CHR ^6’ CH-. At this time R ⁶ R is R ^6’ Each independently selected from H or methyl.

In certain embodiments of the invention, A is-R ⁶ CHR ^6’ CHR ^6” CH-. At this time R ⁶ 、R ^6’ R is R ^6” All are H.

In certain embodiments of the invention, R ⁷ Selected from fluorine.

In certain embodiments of the invention, the compounds have the following structure and steric configuration:

in certain embodiments of the invention, to obtain better TRK inhibition activity, in certain embodiments of the invention, it is preferred that the compounds have the following structure and steric configuration:

in one embodiment of the present invention, the pharmaceutically acceptable salt is an inorganic salt or an organic salt, and the inorganic salt includes hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, nitrate, phosphate, and acid phosphate; the organic salt is selected from acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, sulfonate, benzenesulfonate, salicylate.

The invention also provides a pharmaceutical composition comprising the compound or pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier, excipient or diluent.

The invention also provides the use of the above compound or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of cancer, pain, inflammation, a neurodegenerative disease, trypanosoma cruzi infection or osteolytic disease in a mammal.

In one embodiment of the invention, the cancer comprises melanoma, non-small cell lung cancer, thyroid cancer, acute myelogenous leukemia, glioblastoma, astrocytoma and medulloblastoma, colon cancer, neuroblastoma, ovarian cancer, breast cancer, prostate cancer, pancreatic cancer, multiple myeloma, and large cell neuroendocrine tumor; wherein the pain is related to cancer, surgery, bone fracture, bone pain caused by tumor metastasis, osteoarthritis, psoriatic arthritis, rheumatoid arthritis, interstitial cystitis, chronic pancreatitis, visceral pain, inflammatory pain, migraine, chronic lumbar back pain, bladder pain syndrome, and neuropathic pain; wherein the inflammation comprises asthma, interstitial cystitis, ulcerative colitis, crohn's disease, atopic dermatitis, eczema, and psoriasis.

Furthermore, the present invention provides the use of a compound of the present invention or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of cancer, pain, inflammation, neurodegenerative disease, trypanosoma cruzi infection or osteolytic disease in a mammal. Preferably, wherein the cancer comprises melanoma, non-small cell lung cancer, thyroid cancer, acute myelogenous leukemia, glioblastoma, astrocytoma and medulloblastoma, colon cancer, neuroblastoma, ovarian cancer, breast cancer, prostate cancer, pancreatic cancer, multiple myeloma, and large cell neuroendocrine tumor; wherein the pain is related to cancer, surgery, bone fracture, bone pain caused by tumor metastasis, osteoarthritis, psoriatic arthritis, rheumatoid arthritis, interstitial cystitis, chronic pancreatitis, visceral pain, inflammatory pain, migraine, chronic lumbar back pain, bladder pain syndrome, and neuropathic pain; wherein the inflammation comprises asthma, interstitial cystitis, ulcerative colitis, crohn's disease, atopic dermatitis, eczema, and psoriasis.

The beneficial effects are that:

the structural compound shown in the general formula (I) provided by the invention has better selectivity to TRK and more novel structure compared with the lopatinib (reportectrinib) under the condition of ensuring good TRK inhibitory activity and metabolic stability.

Detailed Description

The technical scheme of the present invention will be described in detail with reference to examples.

In the context of the present invention "(1-2C) alkyl" refers to a saturated branched or branched monovalent hydrocarbon radical of 1 to 2 carbon atoms, respectively. Examples include, but are not limited to, methyl, ethyl. "(1-3C) alkyl" is analogically defined, specific examples include, but are not limited to, methyl, ethyl, 1-propyl, 2-propyl.

In the present invention, "halogen" means fluorine, chlorine, bromine and iodine.

In the present invention, "pharmaceutically acceptable salts" refer to those salts which retain the biological effectiveness and properties of the parent compound. The term "salt" refers to any salt of a compound according to the invention prepared from an inorganic or organic acid or base and an internally formed salt. Typically, such salts have a physiologically acceptable anion or cation.

In the present invention, "administering" or "administering" a compound to an individual refers to providing a compound of the present invention to an individual in need of treatment.

Reference throughout this specification to "an embodiment" or "in another embodiment" or "in certain embodiments" or "in some embodiments of the present application" means that a particular reference element, structure, or feature described in connection with the embodiment is included in at least one embodiment. Thus, the appearances of the phrases "in one embodiment" or "in an embodiment" or "in another embodiment" or "in certain embodiments" or "in some embodiments" in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular elements, structures, or features may be combined in any suitable manner in one or more embodiments.

Throughout the specification and claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be interpreted in an open-ended, inclusive sense, i.e. "including but not limited to.

It should be understood that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

< Compound or pharmaceutically acceptable salt thereof >

Further, the compounds of formula (I) or their salts may be isolated in the form of solvates, and thus any such solvates are within the scope of the invention.

The compounds of formula (I) also include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds of the invention include compounds in which one or more hydrogen atoms are replaced with deuterium or tritium or one or more carbon atoms are replaced with 13C-or 14C-enriched carbon, which are within the scope of the invention.

The compounds of the present invention or pharmaceutically acceptable salts thereof may be formulated as solid formulations for oral administration, including, but not limited to, capsules, tablets, pills, powders, granules, and the like. In these solid dosage forms, the compounds of formula (I) according to the invention are mixed as active ingredient with at least one conventional inert excipient (or carrier), for example with sodium citrate or dicalcium phosphate. Or with the following components: (1) Fillers or solubilisers, for example starch, lactose, sucrose, glucose, mannitol, silicic acid and the like; (2) Binders, for example, hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, acacia, and the like; (3) humectants, for example, glycerin, etc.; (4) Disintegrants, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, sodium carbonate, and the like; (5) a slow solvent such as paraffin wax or the like; (6) absorption accelerators such as quaternary ammonium compounds and the like; (7) Wetting agents such as cetyl alcohol and glycerol monostearate, and the like; (8) adsorbents such as kaolin and the like; (9) Lubricants, for example, talc, calcium stearate, solid polyethylene glycol, sodium lauryl sulfate, and the like, or mixtures thereof. Buffers may also be included in capsules, tablets, pills.

The solid dosage forms, such as tablets, dragees, capsules, pills and granules, may be provided with coatings and shell materials such as enteric coatings and other materials known in the art in the form of crystalline coatings or microencapsulations. They may contain opacifying agents and the release of the active ingredient in such a composition may be released in a delayed manner in a certain part of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes. The active ingredient may also be in the form of microcapsules with one or more of the above excipients, if desired.

The compounds of the present invention or pharmaceutically acceptable salts thereof may be formulated into liquid dosage forms for oral administration, including, but not limited to, pharmaceutically acceptable emulsions, solutions, suspensions, syrups, tinctures, and the like. In addition to the compounds of formula (I) or pharmaceutically acceptable salts thereof as active ingredients, liquid dosage forms may contain inert diluents commonly used in the art such as water and other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1, 3-butylene glycol, dimethylformamide and oils, in particular cottonseed, groundnut, corn, olive, castor, sesame oils and the like or mixtures of these substances and the like. In addition to these inert diluents, the liquid dosage forms of the present invention can also include conventional adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents and the like.

Such suspending agents include, for example, ethoxylated stearyl alcohol, polyoxyethylene sorbitol, and sorbitan, microcrystalline cellulose, agar-agar, and the like, or mixtures of these.

The compounds of the present invention and pharmaceutically acceptable salts thereof may be formulated in dosage forms for parenteral injection, including, but not limited to, physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions and dispersions. Suitable carriers, diluents, solvents, excipients include water, ethanol, polyols and suitable mixtures thereof.

The compounds of the present invention or pharmaceutically acceptable salts thereof may be formulated into dosage forms for topical administration, including, for example, ointments, powders, suppositories, drops, sprays, inhalants and the like. The compounds of the general formula (I) according to the invention or their pharmaceutically acceptable salts as active ingredients are mixed under sterile conditions with physiologically acceptable carriers and optionally with preservatives, buffers and, if appropriate, propellants.

The pharmaceutical composition of the invention comprises a compound of a general formula (I) or pharmaceutically acceptable salt thereof as an active ingredient, and pharmaceutically acceptable carriers, excipients and diluents. In preparing pharmaceutical compositions, a compound of formula (I) or a pharmaceutically acceptable salt thereof of the present invention is typically admixed with a pharmaceutically acceptable carrier, excipient or diluent. Wherein the content of the compound of the general formula (I) or a pharmaceutically acceptable salt thereof may be 0.01 to 1000mg, for example, 0.05 to 800mg, 0.1 to 500mg, 0.01 to 300mg, 0.01 to 200mg, 0.05 to 150mg, 0.05 to 50mg, etc.

< use >

The invention also provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment of cancer, pain, inflammation, a neurodegenerative disease, trypanosoma cruzi infection or osteolytic disease in a mammal.

A method for treating cancer, pain, inflammation, neurodegenerative diseases, trypanosoma cruzi infection or osteolytic diseases in a mammal, including a human, by inhibiting tropomyosin kinase (TRK) activity, comprising administering to the mammal, including a human, a therapeutically effective amount of a compound of formula I described above, or a pharmaceutically acceptable salt thereof.

A "therapeutically effective amount" is an amount of a compound of the invention effective to produce a biological or medical response (e.g., reduce or inhibit the activity of an enzymatically active protein, or ameliorate symptoms, alleviate a condition, slow or delay the progression of a disease, or prevent a disease) in an individual.

Cancers contemplated by the present invention include melanoma, non-small cell lung cancer, thyroid cancer, acute myelogenous leukemia, glioblastoma, astrocytoma and medulloblastoma, colon cancer, neuroblastoma, ovarian cancer, breast cancer, prostate cancer, pancreatic cancer, multiple myeloma, and large cell neuroendocrine tumor.

Pain referred to in the present invention relates to cancer, surgery, bone fracture, bone pain caused by tumor metastasis, osteoarthritis, psoriatic arthritis, rheumatoid arthritis, interstitial cystitis, chronic pancreatitis, visceral pain, inflammatory pain, migraine, chronic lumbago, backache, bladder pain syndrome, and neuropathic pain.

Inflammation as referred to herein includes asthma, interstitial cystitis, ulcerative colitis, crohn's disease, atopic dermatitis, eczema and psoriasis.

The compounds of the present invention or pharmaceutically acceptable salts thereof may be administered to mammals, including humans, orally, rectally, parenterally (intravenous, intramuscular or subcutaneous), topically (powders, ointments, drops) or intratumorally.

The compounds of the present invention, or pharmaceutically acceptable salts thereof, may be administered alone or in combination with other pharmaceutically acceptable therapeutic agents, in combination with other antitumor agents. The combination therapy may be achieved by the simultaneous, sequential or separate use of the individual components of the therapy. Such therapeutic agents include, but are not limited to: antitumor drugs acting on DNA chemical structures such as cisplatin, antitumor drugs affecting nucleotide synthesis such as methotrexate, 5-fluorouracil and the like, antitumor drugs affecting nucleic acid transcription such as doxorubicin, epirubicin, aclacinomycin and the like, antitumor drugs acting on tubulin synthesis such as paclitaxel, vinorelbine and the like, aromatase inhibitors such as aminoglutethimide, letrozole, ryanodine and the like, cell signal pathway inhibitors such as epidermal growth factor receptor inhibitors Imatinib (Imatinib), gefitinib (Gefitinib), erlotinib (Erlotinib) and the like. Anti-tumor monoclonal antibodies, immunosuppressants PD-1, PD-L1, etc., and the individual components to be combined may be administered simultaneously or sequentially, in a single formulation or in different formulations. The combinations include not only combinations of one or more other active agents of the compounds of the present invention, but also combinations of two or more other active agents of the compounds of the present invention.

In combination with one or more other drugs that act by the same or different mechanism of action. The combination therapy may be achieved by the simultaneous, sequential or separate use of the individual components of the therapy. Such therapeutic agents include, but are not limited to: steroid dexamethasone, cortisone, fluticasone; analgesic, aspirin, ibuprofen, indomethacin and opioids.

The following examples illustrate, but do not limit, the synthesis of compounds of formula (I).

The temperatures are in degrees celsius. Reagents were purchased from commercial suppliers and used without further purification, if not otherwise indicated. The structure of the end products, intermediates and starting materials is confirmed by standard analytical methods, such as spectroscopic characterization, MS, NMR. Abbreviations used are conventional in the art.

Preparation of intermediate a:

preparation of (5R) -5-methyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (intermediate a):

step a: preparation of tert-butyl (R) - (2-hydroxypropyl) carbamate

(R) -1-amino-2-propanol (30 g,0.4 mol) was dissolved in DCM (700 mL), di-tert-butyl dicarbonate (96 g,0.44 mol) and TEA (40 g,0.4 mol) were added at 0deg.C and stirred at room temperature for 20h. Water (400 mL) was added, saturated aqueous sodium bicarbonate (600 mL) was added, the pH was adjusted to 8, the mixture was extracted three times with DCM (600 mL), the saturated aqueous sodium bicarbonate (400 mL) was washed once, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at 20℃to give a colorless oily liquid (68.9 g, 97.5% yield). MS (ESI) M/z 176.1 (M+1) ⁺ 。

Step b: preparation of (5R) -5-methyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2-oxide

Imidazole (104.6 g,1.54 mol) was dissolved in DCM (1100 mL), evacuated and nitrogen blanketed. Thionyl chloride (51.3 g,0.43 mol) was added dropwise with stirring after cooling to-60℃and a solution of tert-butyl (R) - (2-hydroxypropyl) carbamate (66.2 g,0.38 mol) in DCM (500 mL) was added dropwise. Stirring was continued for 3h and then allowed to stir at room temperature overnight. After completion of the reaction, water was added at 0 ℃ and extracted three times with DCM (500 mL), and the organic phase was collected and washed three times with saturated brine (500 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to give the crude product as a yellow oil (78.5 g, 93.4% yield). MS (ESI) M/z 222.1 (M+1) ⁺ 。

Step c: preparation of (5R) -5-methyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (A)

(5R) -5-methyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2-oxide (70 g,0.32 mol) was dissolved in acetonitrile (1300 mL), ruthenium trichloride (72 mg,0.32 mmol) was added, and the mixture was evacuated, nitrogen-protected, and cooled to 0 ℃. Then sodium periodate (135.9 g,0.64 mol) was added dropwiseWater (1300 mL) solution. Remove to room temperature and stir overnight. After completion of the reaction, water (700 mL) was added at 0 ℃ and extracted three times with DCM (1500 mL), and the organic phase was collected and washed three times with saturated brine (1000 mL). Drying over anhydrous sodium sulfate, filtration and concentration gave the crude product as a brown oil, which was column chromatographed to give (5R) -5-methyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (A) as a white solid (46 g,0.194mol, 60.6% yield). MS (ESI) M/z 238.1 (M+1) ⁺ 。

Preparation of intermediate B:

a process for the preparation of tert-butyl 1,2, 3-oxathiazolidine-3-carboxylate 2, 2-dioxide (B).

Step d: preparation of tert-butyl 2-hydroxyethyl carbamate

Ethanolamine (12.2 g,0.2 mol) was dissolved in DCM (300 mL), di-tert-butyl dicarbonate (48 g,0.22 mol) and potassium carbonate (55.2 g,0.4 mol) were added at 0deg.C and stirred overnight at room temperature. Water (400 mL) was added, extracted three times with DCM (300 mL), and the saturated brine solution (200 mL) was washed once, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at low temperature to give a colorless oily liquid (30.2 g, 93.8% yield). MS (ESI) M/z 162.1 (M+1) ⁺ 。

Step e: preparation of tert-butyl 1,2, 3-oxathiazolidine-3-carboxylate 2-oxide

Imidazole (51.7 g,0.76 mol) was dissolved in DCM (500 mL), evacuated and nitrogen blanketed. Thionyl chloride (26.3 g,0.22 mol) was added dropwise with stirring after cooling to-60℃and a solution of tert-butyl 2-hydroxyethylcarbamate (30.2 g,0.19 mol) in DCM (250 mL) was added dropwise. Stirring was continued for 3h and then allowed to stir at room temperature overnight. After completion of the reaction was monitored, water was added at 0 ℃ and extracted three times with DCM (250 mL), and the organic phase was collected and washed twice with saturated brine (250 mL). The organic phase is subjected to anhydrous sulfuric acidSodium was dried, filtered, and concentrated to give the crude product as a yellow oil (29.1 g, 73.4% yield). MS (ESI) M/z207.1 (M+1) ⁺ 。

Step f: preparation of tert-butyl 1,2, 3-oxathiazolidine-3-carboxylate 2, 2-dioxide (B)

Tert-butyl 1,2, 3-oxathiazolidine-3-carboxylate 2-oxide (29.1 g,0.14 mol) was dissolved in acetonitrile (500 mL), ruthenium trichloride (36 mg,0.16 mmol) was added, evacuated, nitrogen-protected, and cooled to 0 ℃. A solution of sodium periodate (68.1 g,0.32 mol) in water (500 mL) was added dropwise. Remove to room temperature and stir overnight. After completion of the reaction, water (300 mL) was added at 0 ℃ and extracted three times with DCM (500 mL), and the organic phase was collected and washed twice with saturated brine (500 mL). Drying over anhydrous sodium sulfate, filtration, and concentration gave the crude product as a brown oil, which was subjected to column chromatography to give 1,2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (B) as a white solid (18.7 g,0.08mol, yield 57.1%). MS (ESI) M/z 223.1 (M+1) ⁺ 。

Preparation of intermediate C:

a process for preparing 4, 4-dimethyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (C).

Step g: preparation of tert-butyl 1-hydroxy-2-methylpropan-2-ylcarbamate

2-amino-2-methyl-1-propanol (17.8 g,0.2 mol) was dissolved in DCM (300 mL), di-tert-butyl dicarbonate (48 g,0.22 mol) and TEA (20.0 g,0.2 mol) were added at 0deg.C and stirred overnight at room temperature. Water (400 mL) was added, extracted three times with DCM (300 mL), and the saturated brine solution (200 mL) was washed once, and the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at low temperature to give a colorless oily liquid (34.4 g, 91.0% yield). MS (ESI) M/z 190.1 (M+1) ⁺ 。

Step h: preparation of 4, 4-dimethyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2-oxide

Imidazole (49.0 g,0.72 mol) was dissolved in DCM (500 mL), evacuated and nitrogen blanketed. Thionyl chloride (25.1 g,0.21 mol) was added dropwise with stirring after cooling to-60℃and a solution of tert-butyl 1-hydroxy-2-methylpropan-2-ylcarbamate (34.4 g,0.18 mol) in DCM (250 mL) was added dropwise. Stirring was continued for 3h and then allowed to stir at room temperature overnight. After completion of the reaction was monitored, water was added at 0 ℃ and extracted three times with DCM (250 mL), and the organic phase was collected and washed twice with saturated brine (250 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to give the crude product as a yellow oil (32.9 g, yield 77.6%). MS (ESI) M/z 236.1 (M+1) ⁺ 。

Step i: preparation of 4, 4-dimethyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (C)

4, 4-dimethyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2-oxide (32.9 g,0.14 mol) was dissolved in acetonitrile (500 mL), ruthenium trichloride (36 mg,0.16 mmol) was added, and the mixture was evacuated, nitrogen-protected and cooled to 0 ℃. A solution of sodium periodate (68.1 g,0.32 mol) in water (500 mL) was added dropwise. Remove to room temperature and stir overnight. After completion of the reaction, water (300 mL) was added at 0 ℃ and extracted three times with DCM (500 mL), and the organic phase was collected and washed twice with saturated brine (500 mL). Drying over anhydrous sodium sulfate, filtration and concentration gave the crude product as a brown oil, which was subjected to column chromatography to give 4, 4-dimethyl-1, 2, 3-oxathiazolidine-3-carboxylic acid tert-butyl ester 2, 2-dioxide (C) as a white solid (22.7 g,0.09mol, 64.3% yield). MS (ESI) M/z 252.1 (M+1) ⁺ 。

Preparation of intermediate D:

preparation method of 5-chloro-6-fluoro-pyrazolo [1,5-a ] pyrimidine-3-ethyl formate (D)

Step j: preparation of ethyl 6-fluoro-5, 7-dihydroxypyrazolo [1,5-a ] pyrimidine-3-carboxylate

To a solution of ethyl 3-aminopyrazole-4-carboxylate (15.0 g,74.3 mmol) and diethyl 2-fluoromalonate (18.3 g,117.9 mmol) in ethanol (250 mL) was added sodium ethoxide (12.63 g,187.8 mmol) and stirred at 90℃for 6 hours. After completion of the reaction, the mixture was cooled to room temperature, diluted with toluene (180 mL) and concentrated under reduced pressure. Toluene (180 mL) was further added to the residue, and the residue was concentrated again under reduced pressure to give the crude ethyl 6-fluoro-5, 7-dihydroxypyrazolo [1,5-a ] pyrimidine-3-carboxylate, which was used in the next step without purification.

Step k: preparation of ethyl 5, 7-dichloro-6-fluoropyrazolo [1,5-a ] pyrimidine-3-carboxylate

The aforementioned 6-fluoro-5, 7-dihydroxypyrazolo [1,5-a ]]The crude pyrimidine-3-carboxylic acid ethyl ester (74.3 mmol) was placed in a three-necked flask and phosphorus oxychloride (294 g,180mL,1938 mmol) was added to the reaction flask at-10 ℃. Vacuumizing and protecting with nitrogen. Stirring was carried out for 24 hours at 100℃in an oil bath. After the completion of the monitoring reaction, the reaction mixture was cooled to room temperature, and phosphorus oxychloride was distilled off. The residue was taken up in water (300 mL) at 0deg.C, pH adjusted to 7-8, and extracted three times with DCM (300 mL). The organic phase was collected, washed once with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to give a brown solid crude product, which was chromatographed to give 5, 7-dichloro-6-fluoropyrazolo [1,5-a ] as a pale yellow solid ]Pyrimidine-3-carboxylic acid ethyl ester (15.5 g,55.8mmol, 75.1% combined two steps). MS (ESI) M/z 280.0 (M+1) ⁺ 。

Step l: preparation of 5-chloro-6-fluoro-pyrazolo [1,5-a ] pyrimidine-3-carboxylic acid ethyl ester (D)

Taking 5, 7-dichloro-6-fluoropyrazolo [1,5-a ]]Pyrimidine-3-carboxylic acid ethyl ester (15.5 g,55.8 mmol) and ammonium chloride (16.2 g,300 mmol) were dissolved in THF (195 mL), water (390 mL), ethanol (585 mL) and placed in a reaction flask. Zinc powder (16.02 g,245.1 mmol) was added to the reaction system with stirring at 0deg.C. Stirring for 3h at 0deg.C. After monitoring the reaction was complete, the zinc powder was removed by filtration and the filter cake was washed with DCM (300 mL). The filtrate was extracted three times with DCM (500 mL), the organic phases were combined, dried over anhydrous sodium sulfate and concentrated to give a brown solid crude product which was column chromatographed to give 5-chloro-6 fluoro-pyrazolo [1,5-a ] as a white solid]Pyrimidine-3-carboxylic acid ethyl ester (D) (9.66 g,39.8mmol, yield)Rate 71.3%). MS (ESI) M/z 244.0 (M+1) ⁺ 。

Example 1

(1 ³ R,1 ^4a E,1 ^5a E, 5S) -34-fluoro-13, 5-dimethyl-13, 14-dihydro-12H-4-oxa-7-aza-1 (4, 6) -pyrazolo [1',5':1,2]Pyrimido [5,4-b][1,4]Oxazin-3 (1, 2) -benzocyclooctan-8-one

Structural formula:

(1 ³ R,1 ^4a E,1 ^5a e, 5S) -34-fluoro-13, 5-dimethyl-13, 14-dihydro-12H-4-oxa-7-aza-1 (4, 6) -pyrazolo [1',5':1,2]Pyrimido [5,4-b ][1,4]Preparation of oxazin-3 (1, 2) -benzocyclooctan-8-one (example 1):

step m: preparation of tert-butyl (S) - (2- (5-fluoro-2-formylphenoxy) propyl) carbamate

To a solution of 4-fluoro-2-hydroxybenzaldehyde (2.8 g,20 mmol) and intermediate A (5.52 g,22 mmol) in DMF (80 mL) was added potassium carbonate (6.9 g,50 mmol) and stirred at 45℃for 15 hours. After monitoring the completion of the reaction, cool to room temperature, add DCM (60 mL), filter with celite and collect the filtrate. The filtrate was extracted three times with DCM (80 mL) and the organic phase was collected and washed three times with saturated brine (40 mL). The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated to give tert-butyl (S) - (2- (5-fluoro-2-formylphenoxy) propyl) carbamate (5.5 g,17.7mmol, 88.5% yield) as a yellow viscous material. MS (ESI) M/z298.1[ M+1 ]] ⁺

Step n: preparation of tert-butyl ((S) 2- (5-fluoro-2- ((((R) -1-hydroxypropan-2-yl) amino) methyl) phenoxy) propyl) carbamate

To a solution of tert-butyl (S) - (2- (5-fluoro-2-formylphenoxy) propyl) carbamate (450 mg,1.45 mmol) in dry methanol (10 mL) was added D-aminopropanol (128 mg,1.75 mmol) and returned at 65 ℃Stream 2h. Cooled to room temperature, sodium borohydride (165 mg,4.34 mmol) was added and stirred overnight at room temperature. After completion of the monitored reaction, it was cooled to 0deg.C, quenched with water (15 mL), extracted three times with DCM (45 mL), and the organic phase was collected and washed twice with saturated brine (15 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to give the crude product as a pale yellow solid, which was column chromatographed to give tert-butyl ((S) 2- (5-fluoro-2- ((((R) -1-hydroxypropan-2-yl) amino) methyl) phenoxy) propyl) carbamate as an off-white solid (250 mg,0.68mmol, 47.0% yield). MS (ESI) m/z 357.2[ M+1 ] ] ⁺

Step o: preparation of ethyl 5- ((2- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) ((R) -1-hydroxypropan-2-yl) amino) -6-fluoro-pyrazolo [1,5-a ] pyrimidine-3-carboxylate

Tert-butyl ((S) 2- (5-fluoro-2- ((((R) -1-hydroxypropan-2-yl) amino) methyl) phenoxy) propyl) carbamate (250 mg,0.676 mmol) was weighed out in n-butanol (9 mL) and intermediate D (165 mg,0.679 mmol) and DIPEA (440 mg,3.41 mmol) were added. Vacuumizing and protecting with nitrogen. Stir overnight at 90 degrees celsius under an oil bath. After completion of the reaction, the reaction mixture was cooled to room temperature, water (10 mL) was added, the mixture was extracted three times with EA (15 mL), and the organic phase was collected and washed three times with saturated brine (10 mL). The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated to give the crude product 5- ((2- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) ((R) -1-hydroxypropan-2-yl) amino) -6-fluoro-pyrazolo [1, 5-a) as a yellow oil]Pyrimidine-3-carboxylic acid ethyl ester (377 mg,0.653mmol, crude yield 96.6%). MS (ESI) m/z 564.3[ M+1 ]] ⁺

Step p: preparation of ethyl (R) -4- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) -3-methyl-3, 4-dihydro-2H-pyrazolo [1',5':1,2] pyrimido [5,4-b ] [1,4] oxazine-6-carboxylate

Weighing 5- ((2- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) ((R) -1-hydroxypropan-2-yl) amino) -6-fluoro-pyrazolo [1, 5-a)]Pyrimidine-3-carboxylic acid ethyl ester (450 mg,0.82 mmol) was dissolved in DMF (30 mL) and placed in a reaction flask. A solution of Kot-Pent (310 mg,2.46 mmol) in toluene (2 mL) was added to the reaction system under stirring. Vacuumizing and protecting nitrogen. Stirring was carried out at room temperature (25 ℃) for 2h. After completion of the monitoring reaction, it was quenched with saturated aqueous ammonium chloride at-20℃and extracted three times with dichloromethane (100 mL). The organic phase was collected, washed twice with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to give a brown oily crude product, which was column chromatographed to give (R) -4- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) -3-methyl-3, 4-dihydro-2H-pyrazolo [1',5':1, 2) as a pale yellow solid]Pyrimido [5,4-b][1,4]Oxazine-6-carboxylic acid ethyl ester (220 mg,0.41mmol, 48.8% yield). MS (ESI) m/z 544.3[ M+1 ]] ⁺

Step q: (1 ³ R,1 ^4a E,1 ^5a E, 5S) -34-fluoro-13, 5-dimethyl-13, 14-dihydro-12H-4-oxa-7-aza-1 (4, 6) -pyrazolo [1',5':1,2]Pyrimido [5,4-b][1,4]Preparation of oxazin-3 (1, 2) -benzocyclooctan-8-one (example 1)

To (R) -4- (2- (((S) -1- ((tert-butoxycarbonyl) amino) propan-2-yl) oxy) -4-fluorophenyl) -3-methyl-3, 4-dihydro-2H-pyrazolo [1',5':1, 2) ]Pyrimido [5,4-b][1,4]To a solution of oxazine-6-carboxylic acid ethyl ester (220 mg,0.41 mmol) in methanol (54 mL) and tetrahydrofuran (18 mL) was added an aqueous solution of lithium hydroxide (2.0M, 18 mL). The mixture was stirred at 60 ℃ overnight. After completion of the reaction, the mixture was monitored to cool to-20℃and then rendered acidic with dilute hydrochloric acid (2.0M) and extracted three times with dichloromethane (90 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, concentrated and dried under reduced pressure overnight. To the residue was added dichloromethane (72 mL), followed by a methanol solution of hydrogen chloride (4.0 m,54 mL). Stirred overnight at room temperature, concentrated under reduced pressure and dried overnight under reduced pressure. To the residue were added dichloromethane (72 mL), DMF (36 mL) and FDPP (616 mg,1.6 mmol), followed by DIPEA (3.28 g,25.2 mmol). Stir at room temperature overnight. After completion of the monitored reaction, quench by adding aqueous sodium carbonate (2M, 90 mL) and extract four times with dichloromethane (180 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, concentrated and dried under reduced pressure overnight. The crude product was obtained as a yellow oil, which was column chromatographed to give a white solid (1 ³ R,1 ^4a E,1 ^5a E, 5S) -34-fluoro-13, 5-dimethyl-13, 14-dihydro-12H-4-oxa-7-aza-1 (4, 6) -pyrazolo [1',5':1,2 ]PyrimidineAnd [5,4-b ]][1,4]Oxazin-3 (1, 2) -benzocyclooctan-8-one (example 1) (24 mg,0.06mmol, 14.6% yield). ). MS (ESI) m/z 398.2[ M+1 ]] ⁺ 。 ¹ H NMR(400MHz,Chloroform-d)δ9.78(d,J＝8.0Hz,1H),8.21(s,1H),8.03(s,1H),7.20(dd,J＝8.2,6.7Hz,1H),6.67(q,J＝2.5Hz,1H),6.64(s,1H),5.57(d,J＝14.8Hz,1H),4.58–4.53(m,1H),4.30(dd,J＝8.5,2.3Hz,1H),4.26–4.22(m,1H),4.16–4.13(m,1H),3.98(d,J＝14.8Hz,1H),3.31(ddd,J＝13.7,8.5,2.5Hz,1H),1.56(d,J＝6.2Hz,3H),1.54(d,J＝6.6Hz,3H). ¹³ C NMR(101MHz,CDCl3)δ164.08,162.61,161.63,156.89,146.04,143.29,132.03,129.28,121.74,119.86,107.85,102.07,100.91,76.13,69.10,54.04,43.60,42.30,29.68,18.48,16.82.

Examples 2-10 (see Table 1) were synthesized following procedures similar to the basic procedure of example 1 to give the desired products.

TABLE 1 Structure and map data for examples 2-10

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EXAMPLE 11 biological Activity test

1. TRKA, TRKB, TRKC ELISA determination

TRKA, TRKB, TRKC kinase activity was assessed using an enzyme-linked immunosorbent assay (ELISA) in the presence of inhibitors. Taking the TRKA kinase activity test as an example: immulon4HBX 384-well microtiter plates (Thermo) were coated with 0.025mg/mL solutions of poly (Glu, ala, tyr;6:3:1;Sigma P3899). Each concentration of test compound, 2.5nM TRKA (Invitrogen, histidine-tagged recombinant human TRK, cytoplasmic domain) and 500 μmol ATP were incubated in the coated plates for 25 minutes at ambient temperature while shaking.The assay buffer consisted of 15mM MOPS pH7.5,0.005% (v/v) Triton X-100 with 5mM MgCl ₂ Composition is prepared. The reaction mixture was removed from the plate by washing with PBS containing 0.1% (v/v) Tween (Tween) 20. The phosphorylated reaction product was detected using 0.2 μg/mL phosphotyrosine specific monoclonal antibody coupled to horseradish peroxidase (clone PY 20) in combination with TMB peroxidase material system (KPL). After the addition of 1M phosphoric acid, the color intensity of the color-developing substance was quantified via absorbance at 450 nm. IC50 values were calculated using 4 or 5-parameter log (logistic) curve fitting. TRKB, TRKC kinase activity was determined using a procedure similar to the basic procedure for TRKA kinase activity assay.

The compounds were tested for kinase inhibitory activity using the same kinase activity test principle for WT-ALK and WT-ROS 1. Table 2 provides specific ICs tested in this assay for the compounds of the invention and the commercially available compound Repotrectinib ₅₀ Values.

In the above activity assay, the compounds of the invention have an average IC of less than 1000nM ₅₀ . Certain compounds have an average IC of less than 100nM ₅₀ The structure of the compound is simpler than that of the commercial compound Repotrectinib, and part of the compounds can reach or be lower than the average IC of the commercial compound Repotrectinib ₅₀ . Table 2 provides specific ICs tested in this assay for the compounds of the invention and the commercially available compound Repotrectinib ₅₀ Values.

TABLE 2 specific IC's for the compounds of the invention and commercially available compounds ₅₀ Value of

Analysis of the above results may yield: compared with repotrectrinib, the compound disclosed by the invention has better selectivity on pan-TRK and does not have obvious inhibition effect on wild ALK and ROS 1. It is speculated that the compounds of the present invention may have a broader therapeutic window when applied.

While the invention has been illustrated by the foregoing specific examples, it should not be construed as being limited thereto; but rather the invention encompasses the generic aspects previously disclosed. Various modifications and embodiments can be made without departing from the spirit and scope of the invention.

Claims

1. A compound having the structure or a pharmaceutically acceptable salt thereof,

2. the compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the pharmaceutically acceptable salt is an inorganic salt or an organic salt, and the inorganic salt is selected from the group consisting of hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, nitrate, phosphate, and acid phosphate; the organic salt is selected from acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, ethanesulfonate, benzenesulfonate, salicylate.

3. A pharmaceutical composition comprising a compound according to any one of claims 1-2 or a pharmaceutically acceptable salt thereof.

4. A composition according to claim 3, further comprising one or more of the following components: pharmaceutically acceptable carriers, excipients, diluents.

5. Use of a compound according to any one of claims 1-2, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the treatment of cancer, pain, inflammation, neurodegenerative disease, trypanosoma cruzi infection or osteolytic disease in a mammal.

6. The use of claim 5, wherein the cancer is selected from the group consisting of melanoma, non-small cell lung cancer, thyroid cancer, acute myelogenous leukemia, glioblastoma, astrocytoma and medulloblastoma, colon cancer, neuroblastoma, ovarian cancer, breast cancer, prostate cancer, pancreatic cancer, multiple myeloma, and large cell neuroendocrine tumor; wherein the pain is selected from the group consisting of bone pain, visceral pain, inflammatory pain, migraine, chronic back pain, bladder pain syndrome and neuropathic pain caused by tumor metastasis; wherein the inflammation is selected from asthma, interstitial cystitis, ulcerative colitis, crohn's disease, atopic dermatitis, eczema and psoriasis.