CN115594671B - Benzothiazole derivative, preparation method and application thereof - Google Patents

Benzothiazole derivative, preparation method and application thereof Download PDF

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CN115594671B
CN115594671B CN202110774091.4A CN202110774091A CN115594671B CN 115594671 B CN115594671 B CN 115594671B CN 202110774091 A CN202110774091 A CN 202110774091A CN 115594671 B CN115594671 B CN 115594671B
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benzothiazol
enamide
amino
ylmethoxy
pyridin
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CN115594671A (en
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刘亚婧
宫平
赵燕芳
侯云雷
秦铭泽
刘默逸
杨帆
马幸幸
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Shenyang Pharmaceutical University
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Abstract

Benzothiazole derivatives shown in general formula I, geometric isomers thereof, pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof, wherein substituent R 1 、R 2 And R 3 Having the meaning given in the claims. The compounds of the general formula I have irreversible tyrosine kinase activation inhibition and can effectively overcome EGFR T790M drug resistance mutation, and the application of the compounds, geometric isomers thereof and pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof in preparing medicines for treating EGFR mutation diseases, in particular to the application in preparing medicines for treating and/or preventing lung cancer, breast cancer, gastric cancer, intestinal cancer and liver cancer.

Description

Benzothiazole derivative, preparation method and application thereof
Technical Field
The invention relates to the field of pharmaceutical chemistry, in particular to benzothiazole derivatives, geometric isomers thereof, pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof, a preparation method thereof and application of a pharmaceutical composition containing the benzothiazole derivatives in treating EGFR mutation diseases, in particular to medicaments for treating lung cancer, breast cancer, gastric cancer, intestinal cancer and liver cancer.
Background
Malignant tumor is a common disease and frequently-occurring disease which seriously threatens human health, and the death rate of human caused by malignant tumor is the second place of all disease death rates, and is next to cardiovascular and cerebrovascular diseases. Among them, lung cancer is one of the malignant tumors that has the highest increase in morbidity and mortality and is the greatest threat to the health and life of people. In recent 50 years, many countries report that the incidence and mortality of lung cancer are obviously increased, and the incidence and mortality of lung cancer in men are the first of all malignant tumors, and the incidence and mortality of lung cancer in women are the second. Whereas Non-small cell lung cancer (Non-small cell lung cancer, NSCLC) accounts for about 80-85% of all lung cancers, about 75% of patients have been found to be in the middle and late stages with 5-year survival rates below 15%.
Methods of treatment for NSCLC include surgical, radiation, drug (chemotherapy) and biological treatments, but chemotherapy remains largely the mainstay of treatment. In recent years, with the rapid development of molecular biology, researchers have more clearly recognized the occurrence and development processes of NSCLC and related targets and signal transduction pathways involved. At present, the molecular targeted therapeutic drug has the characteristics of strong targeting, small toxic and side effects, remarkable curative effect and the like, and has been widely applied in clinic. Whereas the epidermal growth factor receptor tyrosine kinase inhibitor (epidermal growth factor receptor-tyrosine kinase inhibiter, EGFR-TKI) has become the most widely used molecular targeted therapeutic for the clinical treatment of NSCLC.
The epidermal growth factor receptor EGFR (epidermal growth factor receptor) is an expression product of the proto-oncogene HER1, has ligand-induced tyrosine protein kinase activity, and is a member of the HER family. HER families include HER1 (ErbBl, EGFR), HER2 (ErbB 2), HER3 (ErbB 3), and HER4 (ErbB 4), all of which are located on cell membranes.
In recent years, related research has revealed that the occurrence of various malignant tumors is related to the abnormal expression of EGFR, and the abnormal expression of EGFR is closely related to the complex biological behavior process of the tumors and the prognosis of patients. Possible carcinogenic mechanisms are: 1. increased expression of EGFR leads to an enhancement of downstream signaling, possibly due to transcription enhancement at the gene level or amplification of the gene, etc.; 2. an increase in expression of the mutant EGFR receptor or ligand such that the ligand independent self tyrosine kinase region is activated; 3. the increase in ligand overactivates normal receptors, and many tumors can produce excess EGF and/or TGF-Q through both autocrine and paracrine processes, over-amplifying EGFR signaling. Thus, overexpression of EGFR, aberrant activation of downstream signaling pathways, and aberrant function of target genes all contribute to cell immortalization into tumors.
Clinical studies have found that mutation sites of the EGFR gene are mainly concentrated in exons 18, 19, 20 and 21 of the intracellular TK region. Wherein the 19 th exon deletion (amino acid residues 746-750, 45-50% of the total mutation) and the Leu858Arg (L858R) point mutation (40-45% of the mutation) of the 21 st exon are referred to as sensitive mutations, also as common mutations, and the other mutations as rare mutations. Notably, the EGFR gene is subject to drug resistance mutations, the primary reason for which is clinically responsible for the T790M mutation. The T790M mutation replaces the encoded threonine with a bulky methionine, so that the steric hindrance is increased, the binding capacity of a receptor and TKI is reduced, the binding capacity of EGFR and ATP is further improved, EGFR is in an activated state again, the proliferation activity of tumor cells is kept, and finally the activity of an inhibitor is lost, so that drug resistance is generated. This mutation is the primary cause of resistance to the first generation EGFR-TKIs in NSCLC patients, with a mutation frequency of about 50%.
In recent years, aiming at the EGFR target, pharmaceutical chemists have developed a series of therapeutic drugs with different action mechanisms, and the therapeutic drugs can be divided into two major classes of monoclonal antibodies and small molecule tyrosine kinase inhibitors according to different molecular types. The monoclonal antibody can competitively inhibit the combination of EGFR and ligand, thereby inhibiting the activation and autophosphorylation of the receptor, accelerating the endocytosis and degradation of the cell surface receptor, reducing the formation of heterodimer, and inhibiting the activation of a signal transmission system, thereby inhibiting the proliferation of tumor cells; the action mechanism of the small molecule tyrosine kinase inhibitor is that the small molecule tyrosine kinase inhibitor competitively binds with ATP in an intracellular tyrosine kinase region and blocks the interaction between ATP and a tyrosine kinase phosphorylation site, thereby inhibiting the activation of EGFR tyrosine kinase and a series of downstream signal transmission channels.
The EGFR monoclonal antibody brings an effective treatment method and more treatment options for lung cancer patients, and the monoclonal antibody has the advantages of strong action specificity and remarkable curative effect. However, the stability of the medicine is poor, the preparation method and the administration mode are complex, and the cost is high because the biopharmaceutical method is adopted. In addition, in clinical application, the medicines show serious adverse reactions. These reasons limit the clinical application of this type of drug.
Disclosure of Invention
Aiming at the problems existing in the prior art, the invention provides a benzothiazole derivative, a preparation method and application thereof, wherein the benzothiazole derivative has excellent receptor protein tyrosine kinase inhibition activity and anti-tumor cell proliferation cancer activity, and the preparation method is simple and controllable.
The invention relates to a benzothiazole derivative, its geometric isomer, and pharmaceutically acceptable salt, hydrate, solvate or prodrug, its general formula I is:
wherein,
R 1 and R is 2 Identical or different, each independently represents an alkyl or cycloalkyl group, where alkyl is preferably C 1 -C 4 Alkyl, cycloalkyl is preferably C 3 -C 6 Cycloalkyl, or R 1 And R is 2 Together with the attached nitrogen atom, form a 5-10 membered saturated or partially saturated heterocyclic group containing 1-2 heteroatoms selected from N, O, S;
R 3 is a substituent on the benzene ring, preferably 1-3 substituents, independently selected from H, halogen, alkyl, alkoxy, trifluoromethyl, trifluoromethoxy, nitro, 5-10 membered heteroaryl C 1 -C 4 Oxy, alkyl is preferredIs C 1 -C 4 Alkyl, alkoxy is preferably C 1 -C 4 Alkoxy, said heteroaryl containing 1 to 3 heteroatoms selected from N, O, S, 5 to 10 membered heteroaryl C 1 -C 4 The oxy group is preferably pyridylmethylene;
n is any integer selected from 1-3.
The present invention preferably relates to compounds of general formula (I) as defined below, wherein,
R 1 、R 2 identical or different, each independently selected from C 1 -C 4 Alkyl, C 3 -C 6 Cycloalkyl, or R 1 、R 2 Together with the attached nitrogen atom, form a 5-6 membered saturated or partially saturated heterocyclic group containing 1-2 heteroatoms selected from N, O;
R 3 1-3 substituents independently selected from H, halogen, C 1 -C 4 Alkyl, C 1 -C 4 Alkoxy, trifluoromethyl, trifluoromethoxy, nitro, 5-6 membered heteroarylmethyleneoxy, said heteroaryl containing 1-3 heteroatoms selected from N, O, S
n is 1.
The invention also preferably relates to compounds of the general formula I, as defined below, their geometric isomers, and pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,
R 1 、R 2 identical or different, each independently selected from C 1 -C 4 Alkyl, C 3 -C 6 Cycloalkyl, or R 1 、R 2 Together with the attached nitrogen atom, form a 5-6 membered saturated or partially saturated heterocyclic group containing 1-2 heteroatoms selected from N, O;
R 3 1-3 substituents independently selected from H, halogen, C 1 -C 4 Alkyl, C 1 -C 4 Alkoxy, trifluoromethyl, trifluoromethoxy, nitro, 5-6 membered heteroarylmethyleneoxy, said heteroaryl containing 1-3 heteroatoms selected from N, O, S.
The invention particularly preferably relates to compounds of the general formula I, as defined below, their geometric isomers, and their pharmaceutically acceptable salts, hydrates, solvates or prodrugs,
R 1 、R 2 identical or different, each independently selected from C 1 -C 4 Alkyl, or R 1 、R 2 Together with the attached nitrogen atom, form a 5-6 membered saturated heterocyclic group containing 1-2 heteroatoms selected from N, O;
R 3 1-3 substituents independently selected from H, F, cl, methyl, methoxy, trifluoromethyl, trifluoromethoxy, nitro, pyridylmethyleneoxy;
n is 1.
Very particular preference is given according to the invention to the following compounds of the general formula I, the geometrical isomers thereof, and the pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof, which are not meant to be limiting in any way:
(E) -4- (dimethylamino) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E) -4- (dimethylamino) -N- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -4- (dimethylamino) -N- (2- ((3-nitro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -4- (dimethylamino) -N- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -4- (dimethylamino) -N- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) - (morpholin-4-yl) -N- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) -N- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) -N- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide.
Furthermore, benzothiazole derivatives of formula I may form pharmaceutically acceptable salts with acids according to some of the usual methods in the art to which the present invention pertains. Pharmaceutically acceptable salts include inorganic and organic acid addition salts, with the following acid addition salts being particularly preferred: hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, naphthalenedisulfonic acid, acetic acid, propionic acid, lactic acid, trifluoroacetic acid, maleic acid, citric acid, fumaric acid, oxalic acid, tartaric acid, benzoic acid.
In addition, prodrugs of the compounds of the present invention are also encompassed by the present invention. Prodrugs are derivatives of the compounds of formula i according to the invention, which may themselves have a relatively weak activity or even no activity, but are converted to the corresponding biologically active form after administration under physiological conditions (e.g. by metabolism, solvolysis or otherwise).
"halogen" in the present invention means fluorine or chlorine; "alkyl" refers to a straight or branched chain alkyl group; "cycloalkyl" refers to a substituted or unsubstituted cycloalkyl; "heteroaryl" means a monocyclic or polycyclic ring system containing one or more heteroatoms selected from N, O, S, the ring system being aromatic, such as imidazolyl, pyridyl, pyrazolyl, (1, 2, 3) -and (1, 2, 4) -triazolyl, furyl, thienyl, pyrrolyl, thiazolyl, benzothiazolyl, oxazolyl, isoxazolyl, naphthyl, quinolinyl, isoquinolinyl, benzimidazolyl, benzoxazolyl; "saturated or partially saturated heterocyclyl" refers to a monocyclic or polycyclic ring system containing one or more heteroatoms selected from N, O, S, such as pyrrolidinyl, morpholinyl, piperazinyl, piperidinyl, pyrazolidinyl, imidazolidinyl, phthalimidyl, and thiazolinyl.
The following scheme A describes the preparation of the general formula I of the present invention, all starting materials being prepared by the methods described in these schemes, by methods well known to those of ordinary skill in the art of organic chemistry, or are commercially available. All final compounds of the invention are prepared by the methods described in these schemes or by methods analogous thereto, which are well known to those of ordinary skill in the art of organic chemistry. All variable factors applied in these illustrations are as defined below or as defined in the claims.
Route A synthetic route to the Compounds of formula I
In scheme A, starting from 2-amino-6-nitrobenzothiazole (1), via (Boc) 2 O protection, reduction and condensation with diethoxyphosphoric acetic acid to obtain an intermediate 4, then the intermediate 4 and the intermediate 7 react with Wittig-Horner to obtain an intermediate 5, then deprotection is carried out to obtain an intermediate 6, and finally the intermediate 6 and the intermediate 8 react with substitution to obtain the target compound of the general formula I.
Intermediate 7 is prepared as shown in scheme B by hydrolysis of starting material 9 in concentrated hydrochloric acid.
Scheme B Synthesis of intermediate 7
The pharmaceutical composition of the invention contains benzothiazole derivatives of the general formula I and pharmaceutically acceptable salts thereof as active ingredients, and also contains pharmaceutically acceptable carriers or excipients.
Pharmaceutically acceptable carriers for use in the pharmaceutical compositions of the invention are common types available in the pharmaceutical arts and include: binders, lubricants, disintegrants, cosolvents, diluents, stabilizers, suspending agents, pigment-free agents, flavoring agents and the like for oral preparations; preservatives, solubilizing agents, stabilizers and the like for injectable formulations; matrix for topical formulations, diluents, lubricants, preservatives and the like.
The pharmaceutical compositions of the invention may be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically), and if some of the agents are unstable in gastric conditions, they may be formulated as enteric coated tablets which are more practical than the administration of monoclonal antibody agents, and conventional formulation techniques may be used in preparing such pharmaceutical compositions.
The benzothiazole derivative or the pharmaceutically acceptable salt or the pharmaceutical composition is applied to the preparation of medicines for treating and/or preventing EGFR mutant diseases, particularly the medicines for treating and/or preventing EGFR mutant tumors, and particularly the medicines for treating and/or preventing lung cancer, breast cancer, gastric cancer, intestinal cancer or liver cancer.
The benzothiazole derivative and the preparation method and the application thereof provided by the invention are subjected to inhibition activity screening of EGFR T790M mutant kinase and T790M/L858R double mutant kinase and anti-tumor cell proliferation activity screening of EGFR T790M/L858R double mutant non-small cell lung cancer H1975 cell strain, and the result shows that the benzothiazole derivative and pharmaceutically acceptable salt thereof have excellent receptor protein tyrosine kinase inhibition activity and anti-tumor cell proliferation cancer activity. Therefore, the compound has good application prospect.
Detailed Description
The examples and preparations provided below further illustrate and exemplify the compounds of the invention and methods of preparing the same. It should be understood that the scope of the following examples and preparations is not intended to limit the scope of the present invention in any way.
The reagents used in the experiment are analytically pure or chemically pure; the melting points of the compounds are all measured by an MP420/430 full-automatic melting point meter manufactured by Jinan He energy instruments Co., ltd, and the temperature is not corrected; the mass spectrum is measured by an Agilent 1100 type four-stage rod liquid chromatograph; the nuclear magnetic resonance hydrogen spectrum was measured with a Bruker ARX-400MHz nuclear magnetic resonance analyzer.
Example 1 preparation of (E) -4- (dimethylamino) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
Step 1: preparation of tert-butyl (6-nitrobenzothiazol-2-yl) carbamate (2)
10.0g (50 mmol) of 2-amino-6-nitrobenzothiazole are dissolved in 150mL of DMF, 16.8 (g) (77 mmol) of Boc anhydride and 3.1g (25 mmol) of DMAP are added successively at room temperature and the temperature is raised to 90℃for 4h. After the reaction, the reaction solution was cooled to room temperature, poured into cold water, a large amount of yellow solid was precipitated, suction-filtered, and the cake was washed with water and dried to obtain 14.8g of yellow solid with a yield of 97.8%. MS (ESI) m/z 296.1[ M+H ]] + 。
Step 2: preparation of tert-butyl (6-aminobenzothiazol-2-yl) carbamate (3)
At room temperature, 10.0g of34 mmol) of intermediate 2 was added to 100mL of dioxane and the temperature was raised to 70 ℃. 1.4g (5 mmol) of FeCl are added in succession 3 ·6H 2 O and 0.1g (10 mmol) of active carbon, 25.4g (508 mmol) of 80% hydrazine hydrate are added dropwise, and the temperature is raised to 80 ℃ for reaction for 2h after the dripping is finished. After the reaction, the filter cake was washed with hot dioxane, the filtrate was concentrated under reduced pressure, 200mL of water was added to the residue, stirred at room temperature for 30 minutes, suction filtered, and the filter cake was washed with water to give 6.3g of pale brown solid, yield 70.2%. MS (ESI) m/z 266.3[ M+H ]] + 。
Step 3: preparation of tert-butyl (6- (2- (diethoxyphosphoryl) acetamido) benzothiazol-2-yl) carbamate (4)
10.0g (38 mmol) of intermediate 3 was added to 100mL of ethyl acetate at room temperature, the temperature was raised to 70℃and a 50% mass fraction of diethoxyphosphoric acid in ethyl acetate was added dropwise to react for 3 hours at 70 ℃. After the completion of the reaction, the reaction mixture was concentrated under reduced pressure, and 100mL of ethyl acetate, water (3X 30 mL), saturated brine (2X 30 mL) and anhydrous Na were added to the residue 2 SO 4 Drying, suction filtration and vacuum concentration gave 9.1g of grey solid in 54.5% yield. MS (ESI) m/z 444.1[ M+H ]] + 。
Step 4: preparation of tert-butyl (E) - (6- (4- (dimethylamino) but-2-enoylamino) benzothiazol-2-yl) carbamate (5)
5.5g (34 mmol) of dimethylaminoacetaldehyde diethyl acetal were dissolved in 20mL of concentrated hydrochloric acid at room temperature and reacted at 120℃for 4h. After the reaction, the reaction solution was concentrated under reduced pressure to give a brown oil for use without further purification.
5.4g (135 mmol) of sodium hydroxide are dissolved in a mixed solvent of ethanol/water (volume ratio, ethanol: water=10:1), 5.0g (11 mmol) of intermediate 4 are added, after the solution is clear, the brown oil is slowly added dropwise, and the reaction is carried out for 4h at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure, 100mL of water was added, the mixture was extracted with n-butanol (3X 50 mL), the organic layers were combined, washed with water (2X 50 mL), saturated brine (2X 50 mL), and anhydrous Na 2 SO 4 Drying, suction filtration and vacuum concentration gave 2.9g of pale yellow solid with a yield of 67.4%. MS (ESI) m/z 377.4[ M+H ]] + 。
Step 5: (E) Preparation of (E) -N- (2-aminobenzothiazol-6-yl) -4- (dimethylamino) but-2-enamide (6)
2.0g (5.3 mmol) of intermediate 5 are dissolved in 15mL of dichloromethane at room temperature. Trifluoroacetic acid 15 and mL are added dropwise in ice bath, and the reaction is carried out for 3 hours at room temperature after the addition of the trifluoroacetic acid. After the reaction, the reaction solution was concentrated under reduced pressure, 20mL of water was added, the pH was adjusted to 9-10 with saturated sodium carbonate solution, existing solids were precipitated, suction filtered, and the cake was washed with water to give 0.7g of an off-white solid, with a yield of 47.8%. MS (ESI) m/z 275.4[ M-H ]] - 。
Step 6: preparation of 2- ((4-bromo-2-fluorophenoxy) methyl) pyridine (8)
1.7g (10 mmol) of chloromethylpyridine hydrochloride and 2.9g (20 mmol) of potassium carbonate were added to 40mL of DMF at room temperature, stirred at room temperature for 30min, followed by the sequential addition of 2g (10 mmol) of 2-fluoro-4-bromophenol and 0.1g (0.6 mmol) of potassium iodide, and the reaction was carried out at 60℃for 2h. After the reaction, the reaction solution was cooled to room temperature, poured into 200mL of cold water, a large amount of white solid was precipitated, suction-filtered, and the cake was washed with water and dried to give 2.9g of white solid with a yield of 99.4%. MS (ESI) m/z 282.2[ M+H ]] + 。
Step 7: (E) Preparation of (E) -4- (dimethylamino) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
0.15g (0.5 mmol) of intermediate 6, 0.31g (1.1 mmol) of intermediate 8, 0.03g (0.16 mmol) of cuprous iodide and 0.38g (2.7 mmol) of potassium carbonate were added to 10mL of DMSO at room temperature and reacted at 120℃for 2 hours. After the reaction, the reaction mixture was cooled to room temperature, 20mL of water was added, the mixture was extracted with methylene chloride (3X 30 mL), the organic layers were combined, washed with water (2X 30 mL), saturated brine (2X 30 mL), and anhydrous Na 2 SO 4 Drying, suction filtration and vacuum concentration gave 0.1g of a pale brown solid with a yield of 41.9%. 133.1-134.5 ℃, MS (ESI) m/z 476.3[ M-H ]] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.08(s,1H),8.58(d,J＝4.4Hz,1H),7.85(td,J＝7.7,1.6 Hz,1H),7.74–7.66(m,1H),7.63(s,1H),7.61(s,1H),7.52(d,J＝7.8Hz,1H),7.36(dd,J＝7.1, 5.3Hz,1H),7.26(t,J＝8.8Hz,1H),7.17(dd,J＝11.5,1.9Hz,1H),7.10(d,J＝8.4Hz,1H),7.02 (d,J＝8.6Hz,1H),6.68(dt,J＝15.2,6.1Hz,1H),6.23(d,J＝15.3Hz,1H),5.24(s,2H),3.23(d, J＝6.0Hz,2H),2.30(s,6H).
The compounds of examples 2-19 were prepared by the procedure of example 1, with appropriate choice of starting materials and reagents, respectively. When specific reaction starting materials are mentioned, it is understood that one skilled in the art can select appropriate starting materials and reagents as desired for the example.
Benzothiazole derivatives of the general formula I and pharmaceutically acceptable salts thereof:
the structural formulas of examples 1 to 19 of the present invention are shown in the following table 1:
TABLE 1 structural formulas of examples 1-19
Example 2 preparation of (E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
m.p.:133.4～134.6℃，MS(ESI)m/z:492.3[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.09(s,1H),8.58(d,J＝4.5Hz,1H),7.87(t,J＝8.0Hz, 1H),7.70–7.58(m,2H),7.55(d,J＝7.7Hz,1H),7.38(s,2H),7.26(d,J＝8.6Hz,1H),7.20(d,J＝ 8.2Hz,1H),7.09(s,1H),6.77–6.57(m,1H),6.23(d,J＝15.4Hz,1H),5.27(s,2H),3.24(d,J＝ 6.0Hz,2H),2.31(s,6H).
Example 3 preparation of (E) -4- (dimethylamino) -N- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
m.p.:131.7～132.9℃，MS(ESI)m/z:460.3[M+H] + 。
1 H NMR(400MHz,DMSO-d 6 )δ10.04(s,1H),8.58(d,J＝4.7Hz,1H),7.83(t,J＝6.9Hz, 1H),7.59(s,2H),7.51(d,J＝7.8Hz,1H),7.35(dd,J＝6.9,5.2Hz,1H),7.27(d,J＝8.5Hz,2H), 7.06(d,J＝8.7Hz,2H),6.76–6.60(m,1H),6.21(d,J＝15.2Hz,1H),5.17(s,2H),3.18(d,J＝6.0 Hz,2H),2.27(s,6H).
Example 4 preparation of (E) -4- (dimethylamino) -N- (2- ((3-nitro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
m.p.:138.2～140.1℃，MS(ESI)m/z:505.6[M+H] + 。
1 H NMR(600MHz,DMSO-d 6 )δ10.09(s,1H),8.57(d,J＝4.9Hz,1H),8.06(s,1H),7.87(t, J＝8.4Hz,1H),7.77(s,1H),7.62(s,1H),7.58(d,J＝9.0Hz,1H),7.53(d,J＝7.7Hz,1H),7.45(s, 1H),7.40–7.33(m,1H),7.04(d,J＝9.1Hz,1H),6.67(dt,J＝12.6,6.1Hz,1H),6.25(d,J＝15.5 Hz,1H),5.35(s,2H),3.25(s,2H),2.31(s,6H).
Example 5 preparation of (E) -4- (dimethylamino) -N- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
m.p.:137.9～138.5℃，MS(ESI)m/z:474.3[M+H] + 。
1 H NMR(600MHz,DMSO-d 6 )δ8.58(s,1H),8.07(s,1H),7.85(td,J＝7.7,1.6Hz,1H), 7.64(s,1H),7.57(s,1H),7.54(d,J＝7.8Hz,1H),7.35(dd,J＝6.9,5.2Hz,1H),7.21(s,1H),7.12 (s,2H),7.04(d,J＝8.6Hz,1H),6.67(d,J＝15.6Hz,1H),6.23(d,J＝15.4Hz,1H),5.19(s,2H), 3.51(s,2H),2.32(s,6H),2.22(s,3H).
Example 6 preparation of (E) -4- (dimethylamino) -N- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
m.p.:135.4～136.7℃，MS(ESI)m/z:490.3[M+H] + 。
1 H NMR(400MHz,DMSO-d 6 )δ10.07(s,1H),8.57(d,J＝5.3Hz,1H),7.84(td,J＝7.7,1.7 Hz,1H),7.60(d,J＝10.5Hz,2H),7.52(d,J＝7.7Hz,1H),7.35(dd,J＝7.0,5.2Hz,1H),7.09(d, J＝8.7Hz,1H),7.06(d,J＝8.5Hz,1H),7.03(d,J＝2.0Hz,1H),6.79(d,J＝8.5Hz,1H),6.67(dt, J＝12.7,6.1Hz,1H),6.22(d,J＝15.4Hz,1H),5.15(s,2H),3.77(s,3H),3.20(d,J＝6.0Hz,2H), 2.28(s,6H).
Example 7 preparation of (E) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
m.p.:133.1～134.5℃，MS(ESI)m/z:518.4[M+H] + 。
1 H NMR(400MHz,DMSO-d 6 )δ10.06(s,1H),8.58(d,J＝4.0Hz,1H),7.93–7.75(m,1H), 7.62(d,J＝5.8Hz,1H),7.52(d,J＝7.3Hz,2H),7.41–7.31(m,2H),7.30–7.22(m,1H),7.17(d, J＝11.5Hz,1H),7.09(d,J＝6.9Hz,1H),7.01(d,J＝7.4Hz,1H),6.68(dd,J＝15.6,5.5Hz,1H), 5.24(s,2H),3.21(s,2H),2.47(s,4H),1.54(d,J＝3.0Hz,4H),1.41(s,2H).
Example 8 preparation of (E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
m.p.:135.7～136.9℃，MS(ESI)m/z:534.6[M+H] + 。
1 H NMR(600MHz,DMSO-d 6 )δ8.57(s,1H),8.06(s,1H),7.86(d,J＝14.7Hz,1H),7.62(s, 1H),7.54(d,J＝7.7Hz,1H),7.36(d,J＝11.3Hz,2H),7.25(d,J＝8.6Hz,1H),7.19(d,J＝8.8Hz, 1H),7.08(d,J＝8.2Hz,1H),6.68(d,J＝15.6Hz,1H),6.22(t,J＝15.2Hz,1H),5.29(s,2H),3.20 (s,2H),2.47(s,4H),1.54(s,4H),1.41(s,2H).
Example 9 preparation of (E) -N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:140.3～141.8℃，MS(ESI)m/z:518.5[M-H] - 。
1 H NMR(600MHz,DMSO-d 6 )δ8.66–8.49(m,1H),8.06(s,1H),7.85(t,J＝8.4Hz,1H), 7.71(d,J＝8.7Hz,1H),7.53(dd,J＝16.8,8.1Hz,1H),7.48–7.43(m,1H),7.39–7.32(m,1H), 7.26(t,J＝10.5Hz,1H),7.22–7.15(m,1H),7.13(d,J＝8.5Hz,1H),7.01(d,J＝9.0Hz,1H),6.68 (d,J＝15.2Hz,1H),5.36–5.08(s,2H),3.58(d,J＝4.0Hz,4H),3.20–2.96(m,2H),2.37(s,4H).
Example 10 preparation of (E) -N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:138.5～139.9℃，MS(ESI)m/z:536.4[M+H] + 。
1 H NMR(600MHz,DMSO-d 6 )δ10.11(s,1H),8.58(s,1H),7.86(t,J＝6.7Hz,1H),7.77(s, 1H),7.69(d,J＝7.8Hz,1H),7.54(d,J＝7.4Hz,2H),7.37(d,J＝14.7Hz,2H),7.26(d,J＝8.4 Hz,1H),7.22–7.11(m,2H),6.69(d,J＝15.2Hz,1H),5.27(s,2H),3.59(s,4H),3.12(s,2H),2.38 (s,4H).
Example 11 preparation of (E) - (morpholin-4-yl) -N- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
m.p.:137.2～138.8℃，MS(ESI)m/z:500.4[M-H] - 。
1 H NMR(600MHz,DMSO-d 6 )δ10.08(s,1H),8.57(d,J＝4.4Hz,1H),7.83(t,J＝7.7Hz, 1H),7.72–7.62(m,2H),7.51(s,1H),7.50(s,1H),7.34(dd,J＝11.2,6.0Hz,1H),7.26(d,J＝8.7 Hz,2H),7.14(d,J＝8.6Hz,1H),7.06(d,J＝8.8Hz,2H),6.74–6.60(m,1H),5.17(s,2H),3.71– 3.49(m,4H),3.11(d,J＝5.5Hz,2H),2.39(s,4H).
Example 12 preparation of (E) -N- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:129.5～130.7℃，MS(ESI)m/z:514.4[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.09(s,1H),8.57(d,J＝4.5Hz,1H),7.85(t,J＝7.7Hz, 1H),7.71–7.62(m,2H),7.54(d,J＝14.4Hz,1H),7.40–7.30(m,2H),7.20(s,1H),7.13(t,J＝ 8.5Hz,2H),7.04(d,J＝8.6Hz,1H),6.82–6.56(m,1H),5.19(s,2H),3.87–3.47(m,4H),3.15 (d,J＝4.2Hz,2H),2.42(d,J＝2.8Hz,4H),2.21(s,3H).
Example 13 preparation of (E) -N- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:133.5～134.8℃，MS(ESI)m/z:532.4[M+H] + 。
1 H NMR(400MHz,DMSO-d 6 )δ10.07(s,1H),8.56(d,J＝4.7Hz,1H),7.83(t,J＝7.7Hz, 1H),7.73–7.62(m,2H),7.51(d,J＝7.8Hz,1H),7.42–7.29(m,1H),7.13(d,J＝8.6Hz,1H), 7.05(d,J＝8.5Hz,2H),7.02(d,J＝2.0Hz,1H),6.78(d,J＝8.4Hz,1H),6.68(d,J＝15.4Hz, 1H),5.14(s,2H),3.77(s,3H),3.67–3.47(m,4H),3.10(d,J＝5.5Hz,2H),2.38(s,4H).
Example 14 preparation of (E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
m.p.:151.2～152.6℃，MS(ESI)m/z:492.3[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.11(s,1H),8.58(d,J＝4.2Hz,1H),7.85(td,J＝7.7,1.5 Hz,1H),7.79(d,J＝1.2Hz,1H),7.67(dd,J＝8.4,1.1Hz,1H),7.50(d,J＝7.8Hz,1H),7.40– 7.33(m,1H),7.23(dd,J＝8.7,2.4Hz,1H),7.17(d,J＝9.3Hz,1H),7.15(d,J＝8.9Hz,1H),6.69 (dt,J＝15.1,6.1Hz,1H),6.54(d,J＝2.2Hz,1H),6.23(d,J＝15.4Hz,1H),5.28(s,2H),3.21(d, J＝5.8Hz,2H),2.28(s,6H).
Example 15 preparation of (E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
m.p.:161.5～162.4℃，MS(ESI)m/z:534.5[M+H] + 。
1 H NMR(600MHz,DMSO-d 6 )δ10.13(s,1H),8.58(d,J＝4.3Hz,1H),8.07(s,1H),7.84(t, J＝7.7Hz,1H),7.81(s,1H),7.68(d,J＝5.0Hz,1H),7.49(d,J＝7.7Hz,1H),7.42–7.33(m,1H), 7.23(dd,J＝8.7,2.2Hz,1H),7.19(d,J＝5.7Hz,1H),7.15(d,J＝8.7Hz,1H),6.70(dd,J＝15.4, 6.1Hz,1H),6.54(s,1H),5.27(s,2H),3.26(s,2H),2.47(s,4H),1.72–1.48(m,4H),1.41(s,2H).
Example 16 preparation of (E) -N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:162.1～163.8℃，MS(ESI)m/z:514.4[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.16(s,1H),8.58(d,J＝4.3Hz,1H),7.90–7.81(m,2H), 7.75(dd,J＝8.8,2.1Hz,1H),7.48(d,J＝7.8Hz,1H),7.40–7.33(m,1H),7.28–7.24(m,2H), 7.24(d,J＝2.3Hz,1H),7.17(d,J＝8.8Hz,1H),6.70(dt,J＝15.4,5.9Hz,1H),6.52(d,J＝2.4 Hz,1H),5.28(s,2H),3.64–3.51(m,4H),3.12(d,J＝5.2Hz,2H),2.39(s,4H).
Example 17 preparation of (E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
m.p.:168.2～168.9℃，MS(ESI)m/z:538.4[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.13(s,1H),8.56(d,J＝4.3Hz,1H),7.92–7.81(m,2H), 7.76(dd,J＝8.8,2.1Hz,1H),7.49(d,J＝7.8Hz,1H),7.41–7.35(m,1H),7.30–7.25(m,2H), 7.21(d,J＝2.3Hz,1H),7.19(d,J＝8.8Hz,1H),6.75(dt,J＝15.4,5.9Hz,1H),6.53(d,J＝2.4 Hz,1H),5.88(s,2H),3.61–3.52(m,4H),3.19(d,J＝5.2Hz,2H),2.38(s,4H).
Example 18 preparation of (E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
m.p.:139.1～141.5℃，MS(ESI)m/z:578.5[M+H] + 。
1 H NMR(400MHz,DMSO-d 6 )δ10.08(s,1H),8.58(d,J＝4.0Hz,1H),7.93–7.75(m,1H), 7.65(d,J＝5.8Hz,1H),7.52(d,J＝7.3Hz,2H),7.41–7.31(m,2H),7.30–7.22(m,1H),7.17(d, J＝11.5Hz,1H),7.09(d,J＝6.9Hz,1H),7.01(d,J＝7.4Hz,1H),6.68(dd,J＝15.6,5.5Hz,1H), 5.21(s,2H),3.25(s,2H),2.49(s,4H),1.55(d,J＝3.0Hz,4H),1.41(s,2H).
Example 19 preparation of (E) -N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl-2-enamide
m.p.:152.7～153.4℃，MS(ESI)m/z:580.4[M-H] - 。
1 H NMR(400MHz,DMSO-d 6 )δ10.16(s,1H),8.55(d,J＝4.3Hz,1H),7.90–7.81(m,2H), 7.75(dd,J＝8.8,2.1Hz,1H),7.48(d,J＝7.8Hz,1H),7.40–7.33(m,1H),7.28–7.24(m,2H), 7.24(d,J＝2.3Hz,1H),7.20(d,J＝8.8Hz,1H),6.71(dt,J＝15.4,5.9Hz,1H),6.54(d,J＝2.4 Hz,1H),5.29(s,2H),3.66–3.52(m,4H),3.12(d,J＝5.2Hz,2H),2.37(s,4H).
EXAMPLE 20 pharmacological investigation of the products of the invention
In vitro anti-tumor cell proliferation Activity test
The benzothiazole derivatives of the formula I are subjected to in vitro anti-tumor activity screening.
(1) The frozen tube was removed from the liquid nitrogen and placed in a 37℃water bath for rapid thawing. Sucking the liquid in the freezing tube, placing in a centrifuge tube, adding 2-3mL of culture solution, centrifuging at 800rpm for 8min, collecting cells, and placing in 10% fetal bovine serum culture solution for culturing. After 24 hours, the culture solution in the flask is removed, 4mL of digestive juice is added, and 5% CO at 37 ℃ is added 2 In the incubator, after 1min, the flask was knocked down, the cells were digested, poured into a centrifuge tube, and the digestion was stopped by adding about 8mL of a culture solution containing 10% fetal bovine serum. Centrifuging at 800rpm for 8min, discarding supernatant, adding 4mL of culture solution, blowing, mixing, and passaging at (1/4) ratio. The cells after passage were placed in CO at 37 ℃ 2 The cells are cultured in an incubator, and generally can be used for experiments after 2 generations.
(2) Digesting cells cultured in a culture flask to logarithmic cell growth phase with pancreatin, stopping digestion with 10% serum culture solution, loading into a centrifuge tube, sealing, centrifuging, discarding supernatant, blowing with 10% serum culture solution, adjusting cell suspension concentration with culture solution, adding into 96-well plate, adding 100 μl of each well (A1 well is not added, and is set to zero-setting well; 1 and 12 columns are not added as drug self absorbance value contrast), placing cell concentration about 1×104/well at 37deg.C, and 5% CO 2 Culturing in incubator for 24 hr to adhere cells, and performing microscopic examinationPreferably, the cells are about 30% of the cells per well.
(3) The test drugs (1-2 mg) are firstly mixed uniformly by 50 mu L of DMSO in a vortex way, then added with culture solution containing 950 mu L of 10% fetal bovine serum for dilution and uniform mixing, 50 mu L of the diluted liquid medicine is taken out from the first row of holes of a 24-hole plate, 950 mu L of culture solution containing 10% serum is added into the holes of the liquid medicine, uniform mixing is carried out by a pipette, 200 mu L of culture solution containing 10% serum is taken out, the second hole is added with 800 mu L of culture solution containing 10% serum for 5 times dilution, and the like, and the total of 24-hole plates are diluted into 5 drugs with different concentrations for standby. The culture solution in the 96-well plate with embedded cells is thrown out forcefully, the 96-well plate is divided into four areas by cross, the upper row and the lower row of the 96-well plate are used as blank control holes, the previously prepared liquid medicine is respectively added into the 96-well plate according to the sequence from low concentration to high concentration (the 96-well plate is added according to the sequence from right to left), 170 mu L of each hole is added, 3 holes are added per concentration, 170 mu L of each hole culture solution is added, the highest concentration of each adjacent medicine is respectively added into the left row and the right row of holes without cells, and the mixture is placed at 37 ℃ and 5% CO 2 Culturing in an incubator for 72 hours.
(4) From CO 2 The 96-well plate is taken out from the incubator, the prepared 0.5% MTT solution is added into the 96-well plate, and then the mixture is placed at 37 ℃ and 5% CO 2 Culturing in an incubator for 4 hours. And taking out the 96-well plate, forcibly throwing out the liquid in the 96-well plate, adding 100 mu L of DMSO into each well, vibrating on a magnetic vibrator for 3min, fully dissolving the crystal, and measuring the absorbance value of each well on an enzyme-labeled instrument by a double-wavelength method (490 nm,630 nm).
Calculation of IC for each drug by Bliss method based on absorbance 50 Values.
IR% = (control OD-sample OD)/(control OD-blank OD) ×100%.
The in vitro antitumor activity results of the compounds are shown in Table 2.
Table 2 in vitro antitumor cell Activity of the example Compounds
In vitro protein kinase inhibition Activity assay
(1) Compounds were tested for their inhibitory activity against FLT3 and JAK2 kinase by a shift change method (mobility shift assay). 1 Xkinase buffer consisted of 50mM HEPES (pH 7.5), 0.0015% Brij-35; the stop buffer consisted of 100mM HEPES (pH 7.5), 0.015% Brij-35, 0.2%Coating Reagent#3 and 50mM EDTA.
(2) The test compound was precisely weighed and diluted with 100% dmso to 50 times the target concentration to prepare a stock solution. Before testing, adding 10 μl stock solution into 90 μl 1 Xkinase buffer, oscillating for 10min, and mixing to obtain 5 Xcompound solution; adding a certain amount of kinase into a1 Xkinase buffer solution (the concentration of different kinase needed to be configured is different) to prepare a 2.5 Xkinase solution for standby; the FAM-labeled polypeptide substrate and ATP were added to 1 x kinase buffer (different concentrations of the different kinase configuration needed) to make a 2.5 x substrate solution for use.
(3) In 384-well plates, 5 μl of 5×compound solution, 10 μl of 2.5×kinase solution were added sequentially to each well, and incubated at room temperature for 10 minutes; then 10. Mu.L of 2.5 Xsubstrate solution was added and incubated for 1h at 28 ℃; after the incubation has ended 25. Mu.L of stop buffer is added. Reading data on a microplate reader, and calculating the kinase inhibition rate according to a formula:
percent inhibition% = (max-version)/(max-min) ×100% in the formula, max: values for wells without compound added; min: no kinase well values were added.
The in vitro results of the compounds on EGFR T790M mutant and on EGFR T790M/L858R double mutant protein kinase inhibition activity are shown in Table 3.
TABLE 3 in vitro kinase inhibitory Activity of the example Compounds
As is clear from the above test results, the compounds of the general formula I to be protected according to the present invention have excellent receptor protein tyrosine kinase inhibitory activity and antitumor activity against proliferation of tumor cells. Therefore, the compound has good application prospect.

Claims (9)

1. Benzothiazole derivatives shown in general formula I or pharmaceutically acceptable salts thereof,
；
wherein,
in the formula (I) of the present invention,
R 1 and R is 2 Identical or different, each independently represents alkyl, or R 1 And R is 2 Together with the attached nitrogen atom, form a 5-10 membered saturated heterocyclic group containing 1-2 heteroatoms selected from N, O, S;
R 3 is 1-3 substituents on the benzene ring, independently selected from halogen, alkyl, alkoxy, trifluoromethyl, trifluoromethoxy, nitro, 5-10 membered heteroaryl C 1 -C 4 An oxy group, said heteroaryl group containing 1 to 3 heteroatoms selected from N, O, S;
n is any integer selected from 1-3.
2. A benzothiazole derivative of the general formula I or a pharmaceutically acceptable salt thereof as claimed in claim 1, R 1 And R is 2 Identical or different, each independently represents C 1 -C 4 Alkyl, R 3 Independently selected from C 1 -C 4 Alkyl or C 1 -C 4 An alkoxy group.
3. A benzothiazole derivative of the general formula I or a pharmaceutically acceptable salt thereof as claimed in claim 1, wherein,
R 1 and R is 2 Identical or different, each independently selected from C 1 -C 4 Alkyl, or R 1 And R is 2 Together with the attached nitrogen atom, form a 5-6 membered saturated heterocyclic group containing 1-2 heteroatoms selected from N, O;
R 3 is 1-3 substituents on benzene ring, independently selected from halogen, C 1 -C 4 Alkyl, C 1 -C 4 Alkoxy, trifluoromethyl, trifluoromethoxy, nitro, 5-6 membered heteroarylmethyleneoxy, said heteroaryl containing 1-3 heteroatoms selected from N, O, S;
n is 1.
4. A benzothiazole derivative of the general formula I or a pharmaceutically acceptable salt thereof as claimed in claim 3, wherein,
R 1 and R is 2 Identical or different, each independently selected from C 1 -C 4 Alkyl, or R 1 And R is 2 Together with the attached nitrogen atom, form a 5-6 membered saturated heterocyclic group containing 1-2 heteroatoms selected from N, O;
R 3 1-3 substituents independently selected from F, cl, methyl, methoxy, trifluoromethyl, trifluoromethoxy, nitro, pyridylmethyleneoxy;
n is 1.
5. Benzothiazole derivatives or pharmaceutically acceptable salts thereof according to claims 1-4 selected from the group consisting of:
(E) 4- (dimethylamino) materialN- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E)-N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E) 4- (dimethylamino) materialN- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) 4- (dimethylamino) materialN- (2- ((3-nitro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) 4- (dimethylamino) materialN- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E) 4- (dimethylamino) materialN- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E)-N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E)-N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E)-N- (2- ((3-fluoro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E)-N- (2- ((3-chloro-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E) Morpholin-4-ylN- (2- ((4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) but-2-enamide
(E)-N- (2- ((3-methyl-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E)-N- (2- ((3-methoxy-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E)-N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E)-N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E)-N- (2- ((5-chloro-2- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholin-4-yl) butyl but-2-enamide
(E)-N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (dimethylamino) but-2-enamide
(E)-N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) -4- (piperidin-1-yl) butyl-2-enamide
(E)-N- (2- ((3-bromo-4- (pyridin-2-ylmethoxy) phenyl) amino) benzothiazol-6-yl) - (morpholino-4-yl) butyl but-2-enamide.
6. A pharmaceutical composition comprising the benzothiazole derivative of any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof as an active ingredient and a pharmaceutically acceptable excipient.
7. Use of a benzothiazole derivative according to any one of claims 1-5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 for the manufacture of a medicament for the treatment and/or prevention of EGFR mutation diseases.
8. Use of a benzothiazole derivative according to any of claims 1-5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 for the manufacture of a medicament for the treatment and/or prevention of EGFR mutant tumors.
9. Use of a benzothiazole derivative according to any one of claims 1-5 or a pharmaceutically acceptable salt thereof or a pharmaceutical composition according to claim 6 for the preparation of a medicament for the treatment and/or prevention of lung cancer, breast cancer, stomach cancer, intestinal cancer or liver cancer.
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