CN105985342B - As the pyrimido-pyrimidine derovatives of EGFR inhibitor and its application - Google Patents

As the pyrimido-pyrimidine derovatives of EGFR inhibitor and its application Download PDF

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CN105985342B
CN105985342B CN201510064374.4A CN201510064374A CN105985342B CN 105985342 B CN105985342 B CN 105985342B CN 201510064374 A CN201510064374 A CN 201510064374A CN 105985342 B CN105985342 B CN 105985342B
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cancer
compound
egfr
amino
pyrimidine
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CN105985342A (en
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李洪林
徐玉芳
谢华
赵振江
郝永佳
王霞
丁健
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East China University of Science and Technology
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Priority to PCT/CN2016/073726 priority patent/WO2016124160A1/en
Priority to CN201680008995.3A priority patent/CN107428763B/en
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/54—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one sulfur as the ring hetero atoms, e.g. sulthiame
    • A61K31/541—Non-condensed thiazines containing further heterocyclic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems

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Abstract

The present invention relates to as the pyrimido-pyrimidine derovatives of EGFR inhibitor and its application.Specifically, the purposes the present invention relates to compound shown in Formulas I, pharmaceutical composition containing compound of formula I wherein and the compound in preparing treatment EGFR relevant diseases or inhibiting the drug of EGFR：

Description

As the pyrimido-pyrimidine derovatives of EGFR inhibitor and its application
Technical field
The present invention relates to medicinal chemistry arts；Specifically, the present invention relates to novel pyrimido-pyrimidine derovatives, Its synthetic method and its application as EGFR inhibitor in the drug for preparing tumor-related illness.
Background technology
Cancer is also known as malignant tumour, is the major class disease with the characteristics of abnormal cell proliferation and transfer, has incidence The high and high feature of the death rate is to threaten human health, leads to dead one of malignant disease.Data shows 2008 The whole world has 12,700,000 cancer patients, wherein death toll to be up to more than 700 ten thousand.And the new hair tumour patient in the whole world 20% is in State, 24% tumor mortality patient is in China.If not adopting an effective measure prevention, or take out more preferably therapeutic scheme, it is contemplated that To the year two thousand thirty, will occur 26,000,000 newly-increased cases of cancer in world wide every year, number of cancer deaths is up to 17,000,000.Existing In some cancers, lung cancer is the highest malignant tumour of morbidity and mortality, wherein non-small cell lung cancer in current world wide (NSCLC) 80% or more of patients with lung cancer is accounted for.It is predicted according to the World Health Organization (WHO), by 2025, China increased lung cancer newly every year Case will be more than 1,000,000.Once being diagnosed as lung cancer, patient just only has remote survival prospects, and survival rate is less than 15% within 5 years.
Since the 1980s, with going deep into for oncomolecularbiology research, the molecule machine of tumorigenesis System is increasingly clear.In many factors for inducing cancer, certain albumen of high expression caused by gene mutation swash in cancer cell Enzyme is one of the principal element for leading to its signal transduction pathway exception.Protein tyrosine kinase is the weight in signal transduction process The factor is wanted, a series of cellular activities are participated in, it is closely related with cell growth, differentiation, proliferation.The γ phosphates that it is catalyzed ATP turn It moves on on the tyrosine residue of many key proteins, makes phenolic hydroxyl group phosphorylation, to transmit signal.Therefore, development selectivity Kinases inhibitor come block or regulate and control the disease generated extremely due to these signal paths have been considered as it is antitumor One effective research strategy of drug development.In numerous tyrosine kinase, epidermal growth factor recipient tyrosine kinase (epidermal growth factor receptor tyrosine kinase, EGFR) is indispensable important composition portion Point.EGFR is made of 1186 amino acid, encodes the transmembrane glycoprotein that a molecular weight is 170-kDa.EGFR can mediate more Extracellular signal is transmitted to intracellular by bars Signal Transduction Pathways, is sent out the proliferation of normal cell and tumour cell, differentiation and apoptosis Wave important adjustment effect (Cell, 2000,100,113-127).EGFR is many normal epithelial tissues (such as skin and hair follicle) Constructive expression's ingredient, and in most of solid tumor, EGFR, which exists, to be overexpressed or high expression.For example, in lung cancer, The expression rate of EGFR reaches 40~80%.Therefore selectively inhibit EGFR, the signal transduction pathway for interfering it to mediate, Ke Yida To the purpose for the treatment of lung cancer, a practical way is opened for targeted therapy of lung cancer.
On clinical treatment, in conjunction with traditional radiotherapy, chemotherapy, with EGFR targeted drugs such as Gefitinib (Iressa), E Luo Carry out first-line drug for Buddhist nun (Tarceva) etc. is proved to be very effective in lung cancer therapy.However, clinical practice shows： Most of Patients with Non-small-cell Lung can be occurred acquired after using Gefitinib or Tarceva treatment within the 6-12 months Drug resistance.Wherein mutation (790 Soviet Union's ammonia of the drug resistance of about 50% case and an amino acid residue in EGFR kinase domains Sour residue mutations are methionine, T790M) related (The New England Journal of Medicine, 2005,352, 786-792).T790M mutation generate steric hindrance or increase the affine of EGFR and ATP when inhibitor being caused to be combined with EGFR Power so that the anticancer effect for the competitive inhibitor that this kind of invertibity combines weakens significantly.The generation of drug resistance not only reduces Patient has been greatly reduced the life quality of tumor patient to the sensibility of drug.Drug resistance caused by order to overcome T790M mutation Property, a series of irreversible ATP competitive inhibitors (such as CI-1033, HKI-272, PF00299804) have been enter into clinical research Stage.Irreversible inhibitor michael acceptor segment containing there are one, can be with a conserved amino acid of the ATP-binding site of EGFR Residue (Cys797) forms covalent bond, to obtain EGFR binding affinities more stronger than reversible inhibitor.Nevertheless, Since such drug is selectively poor to wild type and mutant egf R, maximal tolerance dose (MTD) is relatively low, clinical trial Effect is not obvious.
Therefore, research and development selective depression T790M mutation, overcome the third generation EGFR targeted drugs of clinical drug-resistant to have Great clinical meaning and application prospect.
Invention content
The object of the present invention is to provide the pyrimido-pyrimidine derovatives conducts for capableing of selective depression T790M mutation EGFR inhibitor.
It is a further object of the present invention to provide the pharmaceutical compositions comprising above compound.
Further object of the present invention is to provide above compound and is preparing treatment EGFR relevant diseases or inhibiting the medicine of EGFR Purposes in object.
In a first aspect, the present invention provides general formula I compounds represented or its pharmaceutically acceptable salt：
In formula, A is phenyl ring, five yuan or hexa-member heterocycle, C3-C8Naphthenic base；
R1It is each independently selected from hydrogen, halogen, C1-C3Alkoxy, C1-C3Alkyl, C1-C4Alkylamidoalkyl, substituted piperazine Base, substituted high piperazine base, substituted morpholinyl, substituted thio-morpholinyl, 4-N- methyl piperazines base, 4-N- Acetylpiperazines Base, 4-N, N- lupetidines base, substituted piperidyl ,-NRaRb, wherein RaAnd RbAlkyl can be independently selected from and contain azanyl；
R2It is each independently selected from following group：
R3It is selected from the group：Hydrogen, C1-C10Alkyl, substituted C1-C10Alkyl, C2-C6Alkenyl, C2-C6Alkynyl, optionally substitution C3-C8Naphthenic base, the aryl optionally replaced, the benzyl optionally replaced, the heterocycle optionally replaced, the heteroaromatic optionally replaced Base；
B is selected from the group：
M is the integer of 0-7.
In a particular embodiment, the compound is as shown in general formula II：
In formula, B, R1、R2、R3It is such as defined above；
M is the integer of 0-5.
In a particular embodiment, the compound is as shown in general formula III：
In formula,
R2It is selected from
R3Selected from C1-C6Alkyl, preferably methyl or isopropyl；Or the C of phenyl substitution1-C6Alkyl, preferably benzyl；
R4、R5、R6、R7And R8It is independently selected from the following group：
In a particular embodiment, R5And R8For H；Alternatively, R5、R7And R8For H.
In a particular embodiment, the present invention provides compound selected from the group below or its pharmaceutically acceptable salt：
In second aspect, the present invention provides a kind of pharmaceutical composition, and described pharmaceutical composition contains first aspect present invention The compound or its pharmaceutically acceptable salt and pharmaceutically acceptable carrier or excipient.
In a preferred embodiment, described pharmaceutical composition is adapted for oral dosage form, including but not limited to tablet, molten Liquor, suspension, capsule, granule, pulvis.
In the third aspect, the present invention provides the compound described in first aspect present invention and is preparing treatment or prevention EGFR Jie The disease led, or inhibit the purposes in the drug of EGFR.
In a particular embodiment, the disease that the EGFR is mediated is cancer.
In a particular embodiment, the cancer is selected from the group：Non-small cell lung cancer, Small Cell Lung Cancer, adenocarcinoma of lung, lung Squamous carcinoma, breast cancer, prostate cancer, neurogliocytoma, oophoroma, G. cephalantha, cervical carcinoma, the cancer of the esophagus, liver cancer, kidney Cancer, cancer of pancreas, colon cancer, cutaneum carcinoma, leukaemia, lymthoma, gastric cancer, multiple bone marrow cancer and solid tumor.
In fourth aspect, the present invention is provided treats or prevents EGFR mediations using the compound described in first aspect present invention Disease method.
In a preferred embodiment, the disease that the EGFR is mediated is cancer；Preferably, the cancer is selected from the group： Non-small cell lung cancer, Small Cell Lung Cancer, adenocarcinoma of lung, lung squamous cancer, breast cancer, prostate cancer, neurogliocytoma, oophoroma, It is G. cephalantha, cervical carcinoma, the cancer of the esophagus, liver cancer, kidney, cancer of pancreas, colon cancer, cutaneum carcinoma, leukaemia, lymthoma, gastric cancer, more Hair property bone marrow cancer and solid tumor.
It should be understood that within the scope of the present invention, above-mentioned each technical characteristic of the invention and have in below (eg embodiment) It can be combined with each other between each technical characteristic of body description, to form a new or preferred technical solution.As space is limited, exist This no longer tires out one by one states.
Specific implementation mode
Inventor after extensive and in-depth study, it was unexpectedly found that the pyrimido-pyrimidine diketone that a collection of structure is completely new Derivative, these derivatives can selective depression EGFR T790M mutation, to EGFRT790M/L858RThe IC of kinase inhibiting activity50 Value reaches nM ranks；To cancer cell (EGFRL858R/T790MMutation) proliferation inhibitory activity IC50Value reaches a μM rank.It is basic herein On complete the present invention.
The present inventor has synthesized the candidate compound with EGFR inhibitory activity.Structure is carried out to obtained candidate compound Optimization, has designed and synthesized a series of pyrimido [4,5-d] pyrimidine -2,4 (1H, 3H)-diones chemical combination for having no document report Object, and carry out finishing structure characterization.The active testing that molecular level and cellular level have been carried out to this series compound, obtains a batch Compound with selective depression EGFR T790M mutation.Wherein compound 007 is to EGFRT790M/L858Kinase inhibiting activity IC50For 30.0nM, H1975 (non-small cell lung cancer cell, EGFRL858R/T790M) cell inhibitory effect activity IC50It is 0.23 μM.
Term defines
Some group definitions being referred to herein are as follows：
Herein, " alkyl " refers to the branched-chain or straight-chain alkyl for the saturation that carbon chain lengths are 1-10 carbon atom, preferred alkane Base includes long 2-8,1-6,1-4,3-8, the alkyl of 1-3 carbon atom not etc..The example of alkyl includes but not limited to： Methyl, ethyl, n-propyl, isopropyl, normal-butyl, isobutyl group, heptyl etc..Alkyl can be replaced by one or more substituent groups, example Such as replaced by halogen or halogenated alkyl.For example, alkyl can be by alkyl or alkyl that 1-4 fluorine atom replaces The alkyl replaced by fluoro-alkyl.
Herein, " alkoxy " refers to by alkyl-substituted oxygroup.Preferred alkoxy is the alcoxyl of long 1-6 carbon atom Base, the alkoxy of more preferably long 1-4 carbon atom, the alkoxy of more preferably long 1-3 carbon atom.The example packet of alkoxy It includes but is not limited to：Methoxyl group, ethyoxyl, propoxyl group etc..Alkoxy can be replaced by one or more substituent groups, such as by halogen Or halogenated alkyl substitution.For example, it can be by fluoro that alkoxy, which can be by alkyl or alkyl that 1-4 fluorine atom replaces, Alkyl-substituted alkyl.
Herein, " alkenyl " usually indicates the univalence hydrocarbyl at least one double bond, and it is former to usually contain 2-8 carbon Son preferably comprises 2-6 carbon atom, can be linear chain or branched chain.The example of alkenyl includes but not limited to vinyl, propylene Base, isopropenyl, cyclobutenyl, isobutenyl, hexenyl etc..
Herein, " alkynyl " usually indicates the univalence hydrocarbyl at least one three key, usually contains 2-8 carbon atom, 2-6 carbon atom is preferably comprised, more generally contains 2-4 carbon atom, can be linear chain or branched chain.The example of alkenyl includes second Alkynyl, propinyl, isopropynyl, butynyl, butynyl, hexin base etc..
Herein, " halogen " refers to fluorine, chlorine, bromine or iodine.
Herein, " aryl " refers to the monocyclic, bicyclic or tricyclic aromatic group containing 6 to 14 carbon atoms, including phenyl, naphthalene Base, phenanthryl, anthryl, indenyl, Fluorene bases, tetrahydro naphthyl, indanyl etc..Aryl optionally by 1-5 (for example, 1,2, 3,4 or 5) substituent group substitution selected from the following：Halogen, C1-4Aldehyde radical, C1-6Alkyl, cyano, nitro, amino, hydroxyl, hydroxyl first Alkoxy (such as trifluoromethoxy), carboxyl, the C that base, the alkyl (such as trifluoromethyl) of halogen substitution, halogen replace1-4Alcoxyl Base, ethoxycarbonyl, N (CH3) and C1-4Acyl group etc., heterocycle or heteroaryl etc..
" heterocycle " used herein includes but not limited to heteroatomic 5 yuan or 6 circle heterocyclic rings for being selected from O, S or N containing 1-3 Group, including but not limited to furyl, thienyl, pyrrole radicals, pyrrolidinyl, pyrazolyl, imidazole radicals, triazolyl, oxazolyls, pyrrole It mutters base, pyridyl group, pyrimidine radicals, pyrazinyl, piperidyl, morpholinyl etc..
" aromatic heterocyclic " used herein refers to and having 6,10 or 14 electronics in ring body containing 5-14 annular atom It fastens shared.And institute's ontaining annular atoms are carbon atom and optional 1-3 hetero atom from oxygen, nitrogen, sulphur.Useful aromatic heterocyclic Including piperazinyl, morpholinyl, piperidyl, pyrrolidinyl, thienyl, furyl, pyranose, pyrrole radicals, imidazole radicals, pyrazolyl, Pyridyl group, including but not limited to 2- pyridyl groups, 3- pyridyl groups and 4- pyridyl groups, pyrazinyl, pyrimidine radicals etc..
Aromatic heterocyclic is optionally replaced by 1-5 (for example, 1,2,3,4 or 5) substituent groups selected from the following：Halogen, C1-4Aldehyde radical, C1-6Linear or branched alkyl group, cyano, nitro, amino, hydroxyl, methylol, halogen substitution alkyl (such as trifluoro Methyl), halogen substitution alkoxy (such as trifluoromethoxy), carboxyl, C1-4Alkoxy, ethoxycarbonyl, N (CH3) and C1-4 Acyl group.
Herein, " acyloxy " refer to structural formula be "-O-C (O)-R " group, wherein R can be selected from alkyl, alkenyl and Alkynyl.The R is optionally substituted.
Herein, " acylamino- " refers to the group that structural formula is "-R '-NH-C (O)-R ", wherein and R ' can be selected from hydrogen or alkyl, R can be selected from alkyl, alkenyl, alkynyl, by NRcRdSubstituted alkyl, by NRcRdSubstituted alkenyl and NRcRdSubstituted alkynyl, The alkyl being optionally substituted by halogen, the alkenyl replaced by cyano, wherein RcAnd RdIt can be selected from alkyl and alkenyl.
Herein, " optionally replace " and refer to substituent group that it is modified optionally by 1-5 (for example, 1,2,3,4 or 5 It is a) substituent group substitution selected from the following：Halogen, C1-4Aldehyde radical, C1-6Linear or branched alkyl group, cyano, nitro, amino, hydroxyl, hydroxyl Alkoxy (such as trifluoromethoxy), carboxyl, the C that methyl, the alkyl (such as trifluoromethyl) of halogen substitution, halogen replace1-4Alkane Oxygroup, ethoxycarbonyl, N (CH3) and C1-4Acyl group.
The compound of the present invention
The compound of the present invention is following general formula I compounds represented or its pharmaceutically acceptable salt：
In formula, A is phenyl ring, five yuan or hexa-member heterocycle, C3-C8Naphthenic base；
R1It is each independently selected from hydrogen, halogen, C1-C3Alkoxy, C1-C3Alkyl, C1-C4Alkylamidoalkyl, substituted piperazine Base, substituted high piperazine base, substituted morpholinyl, substituted thio-morpholinyl, 4-N- methyl piperazines base, 4-N- Acetylpiperazines Base, 4-N, N- lupetidines base, substituted piperidyl ,-NRaRb, wherein RaAnd RbAlkyl can be independently selected from and contain azanyl；
R2It is each independently selected from following group：
R3It is selected from the group：Hydrogen, C1-C10Alkyl, substituted C1-C10Alkyl, C2-C6Alkenyl, C2-C6Alkynyl, optionally substitution C3-C8Naphthenic base, the aryl optionally replaced, the benzyl optionally replaced, the heterocycle optionally replaced, the heteroaromatic optionally replaced Base；
B is selected from the group：
M is the integer of 0-7.
In a particular embodiment, A rings are phenyl ring, to which the compound of the present invention is as shown in following general formula II：
In formula, B, R1、R2、R3As described above；With the integer that m is 0-5.
In a preferred embodiment, the above-mentioned phenyl ring in the compound of the present invention can be substituted or unsubstituted, example Such as, the compound of the present invention can be as shown in following general formula III：
In formula,
R2It is selected from
R3Selected from C1-C6Alkyl, preferably methyl or isopropyl；Or the C of phenyl substitution1-C6Alkyl, preferably benzyl；
R4、R5、R6、R7And R8It is independently selected from the following group：
In further embodiment, the above-mentioned phenyl ring in the compound of the present invention can make ortho position substitution, meta position takes Generation and/or contraposition substitution.In a preferred embodiment, the above-mentioned phenyl ring in the compound of the present invention is ortho position substitution and contraposition Substitution.In another preferred embodiment, the above-mentioned phenyl ring in the compound of the present invention be ortho position substitution, meta position substitution and Contraposition substitution.In a particular embodiment, the R in formula above III5And R8For H；Alternatively, R5、R7And R8For H.
The present inventor synthesize to have obtained a series of structures have no document report pyrimido [4,5-d] pyrimidine -2,4 (1H, 3H)-cyclohexadione compounds, specific compound are as follows：
On the basis of the compound of the present invention, the present invention provides a kind of pharmaceutical composition, and the composition, which contains treatment, to be had The compound of the present invention of effect amount or its pharmaceutically acceptable salt and pharmaceutically acceptable carrier or excipient.
The example of the pharmaceutically acceptable salt of the compounds of this invention includes but not limited to inorganic and acylate, such as salt Hydrochlorate, hydrobromate, sulfate, citrate, lactate, tartrate, maleate, fumarate, mandelate and grass Hydrochlorate；And it is formed with alkali such as sodium hydroxyl, three (hydroxymethyl) aminomethanes (TRIS, amine butantriol) and N-METHYL-ALPHA-L-GLUCOSAMINE Inorganic and organic alkali salt.
Although each Man's Demands are different, those skilled in the art can determine that each in pharmaceutical composition of the present invention is lived The optimal dose of property ingredient.Under normal circumstances, the compound of the present invention or its pharmaceutically acceptable salt, it is daily to mammal Oral medication, dose is according to about 0.0025 to 50 mg kg of body weight.It is preferred that about 0.01 to 10 milli of per kilogram oral medication Gram.For example, unit oral doses may include about 0.01 to 50 milligrams, preferably about 0.1 to 10 milligrams of the compounds of this invention. Unit dose can be given one or many, be daily one or more pieces, and every contains about 0.1 to 50 milligrams, and eligibly about 0.25 To 10 milligrams of the compounds of this invention or its solvate.
The pharmaceutical composition of the present invention can be formulated into the dosage form of suitable various administration routes, including but not limited to quilt It is configured to for parenteral, subcutaneously, vein, muscle is intraperitoneal, transdermal, and oral cavity is intrathecal, encephalic, nasal cavity or topical route administration Form, for treating tumour and other diseases.Dosage is to effectively improve or eliminate the dose of one or more illnesss.For The treatment of specified disease, effective quantity are the doses for being enough to improve or in some manner mitigate symptom related with disease.It is such Dose can be used as single dose application, or can be administered according to effective therapeutic scheme.Dosage also permits healing disease, still It is administered typically to the symptom for improving disease.Repetitively administered is generally required to realize that required symptom improves.The dosage of medicine will According to the age of patient, health and weight, the type of concurrent treatment, the frequency for the treatment of and required treatment benefit determine.
The pharmaceutical preparation of the present invention can give any mammal, as long as they can obtain the treatment of the compounds of this invention Effect.The most importantly mankind in these mammals.
The compound of the present invention or its pharmaceutical composition can be used for treating various by epidermal growth factor receptor kinase (EGFR) disease mediated.Herein, it is various cancers by the disease that EGFR is mediated.The cancer includes but not limited to：It is non-small Cell lung cancer, Small Cell Lung Cancer, adenocarcinoma of lung, lung squamous cancer, breast cancer, prostate cancer, neurogliocytoma, oophoroma, neck It is portion's squamous carcinoma, cervical carcinoma, the cancer of the esophagus, liver cancer, kidney, cancer of pancreas, colon cancer, cutaneum carcinoma, leukaemia, lymthoma, gastric cancer, multiple Bone marrow cancer and solid tumor.
The pharmaceutical preparation of the present invention can manufacture in a known manner.For example, by traditional mixing, granulation, ingot processed, dissolving, Or freezing dry process manufacture.When manufacturing oral preparation, in combination with solid adjuvant material and reactive compound, selective ground and mixed Object.After if necessary or appropriate amount of addition agent being added when necessary, granulate mixture is processed, obtains tablet or pastille core.
Suitable auxiliary material especially filler, such as carbohydrate such as lactose or sucrose, mannitol or sorbierite；Cellulose preparation or Calcium phosphate, such as tricalcium phosphate or calcium monohydrogen phosphate；And binder, such as gelatinized corn starch, including cornstarch, wheaten starch, Rice starch, potato starch, gelatin, tragacanth, methylcellulose, hydroxypropyl methyl cellulose, sodium carboxymethylcellulose, or Polyvinylpyrrolidone.If desired, can increase disintegrant, than starch as mentioned above and carboxymethyl starch, crosslinking is poly- Vinylpyrrolidone, agar or alginic acid or its salt, such as sodium alginate.Adjuvant especially flowing regulator and lubricant, example Such as, silica, talcum, stearates, such as magnesium calcium stearate, stearic acid or polyethylene glycol.If desired, Ke Yi Give pastille cores The suitable coating of gastric juice can be resisted by providing.For this purpose, concentration saccharide solution can be applied.This solution can contain Arabic tree Glue, talcum, polyvinylpyrrolidone, polyethylene glycol and/or titanium dioxide paint solution and suitable organic solvent or solvent mixing Object.In order to prepare the coating of resistant to gastric juice, cellulose solution appropriate, such as cellulose acetate phthalic acid or hydroxypropyl can be used Ylmethyl cellulose phthalic acid.Dyestuff or pigment can be added to the coating of tablet or pastille core.For example, for identification or In order to characterize the combination of active constituent dosage.
Based on above compound and pharmaceutical composition, the present invention further provides a kind of sides of disease that treating EGFR mediations Method, this method include giving the object of needs with the compound of the present invention or pharmaceutical composition.
Medication includes but not limited to various medications well known in the art, can be subject to according to the actual conditions of patient It determines.These methods are including but not limited to parenteral, subcutaneous, vein, muscle, intraperitoneal, transdermal, oral cavity, intrathecal, encephalic, nasal cavity Or topical route administration.
The present invention also includes that the compounds of this invention is preparing the disease for preventing or treating EGFR mediations or inhibiting activity of EGFR Drug in purposes.
Advantages of the present invention：
1. compound provided by the invention is a kind of pyrimido that structure is completely new [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone Class compound；
2. compound provided by the invention has excellent inhibitory activity to the cancer cell that mutant egf R or EGFR are mutated；
It is mutated 3. compound provided by the invention is exploitation energy selective depression T790M, clinical drug-resistant can be overcome EGFR targeted drugs are laid a good foundation, and have great industrialization and commercialization foreground and market value, remarkable in economical benefits.
Technical scheme of the present invention is further described below in conjunction with specific implementation case, but following case study on implementation is not constituted Limitation of the present invention, the various method of administration that all principles and technological means according to the present invention use, belongs to the present invention Range.In the following examples, the experimental methods for specific conditions are not specified, usually according to normal condition, or is built according to manufacturer The condition of view.Unless otherwise stated, otherwise percentage and number are calculated by weight.
Materials and methods
The synthesis of pyrimido [4,5-d] pyrimidine -2,4 (1H, 3H)-cyclohexadione compounds of the present invention is as follows：
Reagent and condition：(a)DIPEA,CH3CN, reflux；(b)ArNH2,CH3CN, reflux；(c) NaOH, THF, reflux； (d)R3NH2,HATU,DIPEA,DMF；(e)K2CO3, CDI, THF, reflux；(f) trifluoroacetic acid, CH2Cl2, 0 DEG C-room temperature；(g) acyl Chlorine, Et3N,CH2Cl2, 0 DEG C-room temperature.
Embodiment 1
The specific synthetic method of above-mentioned steps a-g is as follows：
The synthesis of 1.4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- chlorine pyrimidine -5- Ethyl formates
Bis- chloro- 5- nitro-pyrimidines (2.210g, 10mmol) of 2,4-, DIPEA (1.290g, 10mmol) are weighed in 50mL single port The dissolving of 15mL acetonitriles is added in flask.(3- aminophenyls) t-butyl carbamate (2.080g, 10mmol) is separately taken to be dissolved in 10mL second Nitrile is added drop-wise in above-mentioned reaction solution, drips reflux 2h.TLC tracks to raw material conversion, is cooled to room temperature, and filters, and acetonitrile is washed It washs, filter cake drying obtains 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- chlorine pyrimidine -5- Ethyl formate 3.371g, Yield 86%.1H NMR(400MHz,DMSO-d6)δ10.23(s,1H),9.50(s,1H),8.80(s,1H),7.70(s,1H), 7.35 (d, J=8.0Hz, 1H), 7.29 (t, J=8.0Hz, 1H), 7.23 (d, J=8.0Hz, 1H), 4.38 (q, J=7.2Hz, 2H), 1.49 (s, 9H), 1.36 (t, J=7.2Hz, 3H) .LC-MS:m/z:393.1(M+H)+.
2.4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- (phenyl amino) pyrimidine -5- Ethyl formates Synthesis
Weigh 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- chlorine pyrimidine -5- Ethyl formates (1.960g, 5mmol), the dissolving of 40mL acetonitriles, temperature rising reflux 3h is added in 100mL single-necked flasks in aniline (0.558g, 6mmol).TLC is tracked Raw material converts, and is cooled to room temperature, and filters, washing, and filter cake drying obtains 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) ammonia Base) -2- (phenyl amino) pyrimidine -5- Ethyl formate 1.930g, yield 86%.1H NMR(400MHz,CDCl3)δ10.46(s, 1H), 8.76 (s, 1H), 7.57 (d, J=8.0Hz, 2H), 7.36 (t, J=7.6Hz, 3H), 7.23 (d, J=8.0Hz, 1H), 7.16 (t, J=7.2Hz, 1H), 7.04 (d, J=7.6Hz, 1H), 6.36 (s, 1H), 4.37 (q, J=7.2Hz, 2H), 1.54 (s, 9H), 1.41 (t, J=7.2Hz, 3H) .LC-MS:m/z:450.4(M+H)+.
The synthesis of 3.4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- (phenyl amino) pyrimidine -5- formic acid
Weigh 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- (phenyl amino) pyrimidine -5- Ethyl formates (1.796g, 4mmol) is added the dissolving of 30mL tetrahydrofurans, 1M NaOH solution 10mL is added dropwise, are warming up in 100mL single-necked flasks 50 DEG C, stir 3h.TLC tracks to raw material conversion, is cooled to room temperature, and dilute hydrochloric acid is added and adjusts pH to acidity, filters, washes, filter Cake is dried, and 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- (phenyl amino) pyrimidine -5- formic acid 1.499g are obtained, Yield 89%.1H NMR(400MHz,DMSO-d6)δ10.44(s,1H),9.82(s,1H),9.36(s,1H),8.70(s,1H), 7.69 (d, J=8.0Hz, 2H), 7.63 (s, 1H), 7.43 (s, 1H), 7.26-7.21 (m, 4H), 6.98 (t, J=6.8Hz, 1H),1.47(s,9H).HRMS(ESI):Calculated value C22H24N5O4(M+H)+422.1828 experiment value 422.1823.
4.3- (5- (methylcarbamoyl -2- (phenyl amino) pyrimidine -4- amino) phenyl) t-butyl carbamate Synthesis
Weigh 4- ((3- ((tert-butoxycarbonyl) amino) phenyl) amino) -2- (phenyl amino) pyrimidine -5- formic acid The dissolving of 10mL anhydrous DMFs is added in 50mL flasks in (1.263g, 3mmol), DIPEA (0.774g, 6mmol).Separately take HATU (1.368g, 3.6mmol) is added portionwise in above-mentioned reaction solution, 1h is stirred at room temperature.Addition methylamine hydrochloride (0.396g, 6mmol), it is stirred overnight at room temperature.TLC tracks raw material conversion, and ice water is added, and filters, and washing obtains 3- (5- (methylcarbamoyls Base -2- (phenyl amino) pyrimidine -4- amino) phenyl) t-butyl carbamate 0.612g, yield 47%.1H NMR(400MHz, DMSO-d6) δ 11.33 (s, 1H), 9.59 (s, 1H), 9.31 (s, 1H), 8.64 (s, 1H), 8.50 (d, J=4.8Hz, 1H), 7.68 (d, J=7.8Hz, 2H), 7.62 (s, 1H), 7.49 (br, 1H), 7.23 (t, J=7.8Hz, 2H), 7.19 (d, J= 8.0Hz, 1H), 7.15 (d, J=8.0Hz, 1H), 6.96 (t, J=7.2Hz, 1H), 2.79 (d, J=4.4Hz, 3H), 1.47 (s, 9H).LC-MS:m/z:435.2(M+H)+.
5. (3- (3- methyl -2,4- dioxos -7- (phenyl amino) -3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidines -1 (2H) - Base) phenyl) t-butyl carbamate
Weigh 3- (5- (methylcarbamoyl -2- (phenyl amino) pyrimidine -4- amino) phenyl) t-butyl carbamate (0.434g,1mmol)、K2CO3The dissolving of 5mL anhydrous tetrahydro furans, temperature rising reflux is added in 25mL flasks in (0.276g, 2mmol) Overnight.TLC tracks raw material conversion, is cooled to room temperature, and ice water is added, and EA is extracted, and collected organic layer, rotary evaporation removes solvent, Crude product detaches (EA/PE=2 through silica gel column chromatography:1, v/v) 193mg, yield 42%, are obtained.1H NMR(400MHz,DMSO-d6) δ 9.62 (s, 1H), 8.92 (s, 1H), 7.66 (s, 1H), 7.52 (d, J=8.0Hz, 1H), 7.45 (t, J=8.0Hz, 1H), 7.36-7.32 (m, 2H), 7.02 (d, J=8.0Hz, 3H), 6.90 (d, J=7.6Hz, 1H), 3.29 (s, 3H), 1.45 (s, 9H).HRMS(ESI):Calculated value C24H25N6O4(M+H)+461.1937 experiment value 461.1939.
6.1- (3- aminophenyls) -3- methyl -7- (phenyl amino) pyrimido [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone Synthesis
Weigh (3- (3- methyl -2,4- dioxos -7- (phenyl amino) -3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidines -1 (2H)-yl) phenyl) t-butyl carbamate (0.193g, 0.42mmol) is in 50mL flasks, and addition 5mL dichloromethane dissolves, slowly It is slow that 1mL trifluoroacetic acids are added dropwise, it is stirred overnight at room temperature.TLC tracks raw material conversion, and saturation NaHCO is added3Solution neutralizes, dichloromethane Alkane extracts, collected organic layer, and rotary evaporation removes solvent, obtains 1- (3- aminophenyls) -3- methyl -7- (phenyl amino) pyrimido [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone 0.136g, yield 90%, product is not purified to be directly used in the next step.
7.N- (3- (3- methyl -2,4- dioxos -7- (phenyl amino) -3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidines -1 (2H)-yl) phenyl) acrylamide synthesis (compound 001)
Weigh 1- (3- aminophenyls) -3- methyl -7- (phenyl amino) pyrimido [4,5-d] pyrimidines -2,4 (1H, 3H)-two The dissolving of 3mL dichloromethane, ice bath is added in ketone (0.136g, 0.38mmol), DIPEA (0.097g, 0.76mmol) and 25mL flasks Lower stirring 15 minutes.Separately acryloyl chloride (0.041g, 0.46mmol) is taken to be dissolved in 1mL dichloromethane, is added drop-wise in above-mentioned reaction solution, It is stirred overnight at room temperature.TLC tracks raw material conversion, is spin-dried for, and crude product detaches (EA/PE=1.5 through silica gel column chromatography:1, v/v) N-, is obtained (3- (3- methyl -2,4- dioxos -7- (phenyl amino) -3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidine -1 (2H)-yl) phenyl) third Acrylamide 83mg, yield 53%.1H NMR(400MHz,DMSO-d6) δ 10.38 (s, 1H), 8.93 (s, 1H), 7.82 (d, J= 3.2Hz, 1H), 7.80 (s, 1H), 7.53 (t, J=8.0Hz, 1H), 7.33-7.29 (m, 2H), 7.15 (d, J=7.8Hz, 1H), 7.02-6.96 (m, 2H), 6.89 (t, J=6.0Hz, 1H), 6.44 (dd, J=17.2Hz, J=10.4Hz, 1H), 6.26 (dd, J =17.2Hz, J=1.6Hz, 1H), 5.76 (dd, J=10.4Hz, J=2.0Hz, 1H), 3.30 (s, 3H) .HRMS (ESI):Meter Calculation value C22H19N6O3(M+H)+415.1519 experiment value 415.1512.
Following compound (compound 002-010) synthesizes to obtain according to the method for above-mentioned steps a-g：
((7- ((4- methoxyphenyls) amino) -3- methyl -2,4- dioxo -3,4- dihydro-pyrimidins are simultaneously [4,5-d] by 3- by N- Pyrimidine -1 (2H)) phenyl) acrylamide (compound 002)
1H NMR(400MHz,DMSO-d6) δ 10.37 (s, 1H), 10.23 (s, 1H), 8.88 (s, 1H), 7.82 (d, J= 8.0Hz, 1H), 7.79 (s, 1H), 7.53 (t, J=8.0Hz, 1H), 7.22 (d, J=7.6Hz, 2H), 7.12 (d, J=8.0Hz, 1H), 6.56 (d, J=7.6Hz, 2H), 6.44 (dd, J=16.8Hz, J=10.0Hz, 1H), 6.24 (dd, J=16.8Hz, J= 1.6Hz, 1H), 5.76 (dd, J=10.0Hz, J=1.6Hz, 1H), 3.65 (s, 3H), 3.29 (s, 3H) .HRMS (ESI) are calculated Value C23H21N6O4[M+H]+445.1624 experiment value 445.1627.
((7- ((2- methoxyphenyls) amino) -3- methyl -2,4- dioxo -3,4- dihydro-pyrimidins are simultaneously [4,5-d] by 3- by N- Pyrimidine -1 (2H)) phenyl) acrylamide (compound 003)
1H NMR(400MHz,DMSO-d6)δ10.34(s,1H),8.89(s,1H),8.68(s,1H),7.80(s,1H), 7.76 (d, J=8.0Hz, 1H), 7.50 (t, J=8.0Hz, 1H), 7.41 (d, J=8.0Hz, 1H), 7.12 (d, J=7.6Hz, 1H), 6.95 (d, J=7.6Hz, 2H), 6.44 (dd, J=16.8Hz, J=10.0Hz, 1H), 6.25 (dd, J=17.2Hz, J= 2.0Hz, 1H), 5.76 (dd, J=10.4Hz, J=2.0Hz, 1H), 3.78 (s, 3H), 3.29 (s, 3H) .HRMS (ESI) are calculated Value C23H21N6O4[M+H]+445.1624 experiment value 445.1631.
N- (3- (7- ((2- methoxyl group -4- morphlinophenyls) amino) -3- methyl -2,4- dioxo -3,4- dihydro-pyrimidins And [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 004)
1H NMR(400MHz,DMSO-d6)δ10.36(s,1H),8.80(s,1H),8.44(s,1H),8.03(s,1H), 7.85 (d, J=8.0Hz, 1H), 7.69 (t, J=2.0Hz, 1H), 7.52 (t, J=8.4Hz, 1H), 7.31 (d, J=8.8Hz, 1H), 7.09 (d, J=7.6Hz, 1H), 6.57 (d, J=2.0Hz, 1H), 6.45 (dd, J=16.8Hz, J=10.0Hz, 1H), 6.26 (dd, J=17.2Hz, J=2.0Hz, 1H), 5.77 (dd, J=10.4Hz, J=2.0Hz, 1H), 3.77 (s, 3H), 3.58-3.54 (m, 4H), 3.06-2.99 (m, 4H), 2.05 (s, 3H) .HRMS (ESI) calculated values C27H28N7O5[M+H]+ 528.1995 experiment value 528.1993.
N- (3- (7- ((2- methoxyl group -4- thiomorpholines are for phenyl) amino) -3- methyl -2,4- dioxo -3,4- dihydros Pyrimido [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 005)
1H NMR(400MHz,DMSO-d6)δ10.35(s,1H),8.84(s,1H),8.60(s,1H),8.85(s,1H), 7.70 (s, 1H), 7.48 (t, J=8.0Hz, 1H), 7.17 (d, J=8.4Hz, 1H), 7.09 (d, J=7.6Hz, 1H), 6.44 (dd, J=17.2Hz, J=10.0Hz, 1H), 6.27 (dd, J=16.8Hz, J=2.0Hz, 1H), 5.97 (s, 1H), 5.77 (dd, J=10.0Hz, J=1.6Hz, 1H), 3.75 (s, 3H), 3.42-3.39 (m, 4H), 3.28 (s, 3H), 2.66-2.63 (m, 4H) .HRMS (ESI) calculated values C27H28N7O4S[M+H]+546.1923 experiment value 546.1924.
N- (3- (7- ((4- (4- acetyl group -3-) -2- methoxyphenyls) amino) -3- methyl -2,4- dioxos -3,4- two Hydrogen pyrimido [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 006)
1H NMR(400MHz,DMSO-d6)δ10.34(s,1H),8.83(s,1H),8.59(s,1H),7.86(s,1H), 7.69 (s, 1H), 7.48 (t, J=8.4Hz, 1H), 7.17 (d, J=8.8Hz, 1H), 7.09 (d, J=8.0Hz, 1H), 6.55 (s, 1H), 6.44 (dd, J=17.2Hz, J=10.4Hz, 1H), 6.26 (dd, J=17.2Hz, J=1.6Hz, 1H), 5.99 (s, 1H), 5.76 (dd, J=10.0Hz, J=1.2Hz, 1H), 3.76 (s, 3H), 3.58-3.54 (m, 4H), 3.28 (s, 3H), 3.05-2.99 (m, 4H), 2.05 (s, 3H) .HRMS (ESI) calculated values C29H31N8O5[M+H]+571.2417, experiment value 571.2417。
N- (3- (7- ((4- (4- (dimethylamino) piperidines) -2- methoxyphenyls) amino) -3- methyl -2,4- dioxos - 3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 007)
1H NMR(400MHz,DMSO-d6)δ10.36(s,1H),8.83(s,1H),8.54(s,1H),7.88-7.86(m, 1H), 7.70 (s, 1H), 7.47 (t, J=8.0Hz, 1H), 7.16 (d, J=8.4Hz, 1H), 7.08 (d, J=8.0Hz, 1H), 6.50-6.43 (m, 2H), 6.26 (dd, J=16.8Hz, J=1.6Hz, 1H), 5.98 (s, 1H), 5.76 (dd, J=10.0Hz, J =1.2Hz, 1H), 3.75 (s, 3H), 3.59-3.58 (m, 2H), 3.28 (s, 3H), 2.60-2.55 (m, 2H), 2.22 (s, 6H), 1.82-1.79 (m, 2H), 1.47-1.40 (m, 2H) .HRMS (ESI) calculated values C30H35N8O4[M+H]+571.2781, experiment value 571.2780。
N- (3- (7- ((4- (4- acetylpiperazines base) -2- methoxyphenyls) amino) -3- ethyl -2,4- dioxos -3,4- Dihydro-pyrimidin simultaneously [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 008)
1H NMR(400MHz,DMSO-d6) δ 10.38 (s, 1H), 8.85 (s, 1H), 8.64 (s, 1H), 7.88 (d, J= 4.0Hz, 1H), 7.69 (s, 1H), 7.49 (t, J=8.0Hz, 1H), 7.18 (d, J=8.4Hz, 1H), 7.12 (d, J=7.6Hz, 1H), 6.55 (s, 1H), 6.46 (dd, J=17.2Hz, J=10.0Hz, 1H), 6.27 (dd, J=16.8Hz, J=1.6Hz, 1H), 6.00-5.97 (m, 1H), 5.78 (dd, J=10.0Hz, J=2.0Hz, 1H), 3.94 (q, J=6.4Hz, 2H), 3.77 (s, 3H), 3.58-3.54 (m, 4H), 3.05-2.98 (m, 4H), 2.06 (s, 3H), 1.19 (t, J=6.8Hz, 3H) .HRMS (ESI) calculated value C30H33N8O5[M+H]+585.2574 experiment value 585.2572.
N- (3- (7- ((4- (4- acetylpiperazines base)-2- methoxyphenyls) amino) dioxo-3-3- isopropyl-2,4-, 4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidine -1 (2H)) phenyl) acrylamide (compound 009)
1H NMR(400MHz,DMSO-d6) δ 10.37 (s, 1H), 8.83 (s, 1H), 8.61 (s, 1H), 7.86 (d, J= 8.0Hz, 1H), 7.67 (s, 1H), 7.48 (t, J=8.0Hz, 1H), 7.17 (d, J=7.6Hz, 1H), 7.10 (d, J=7.6Hz, 1H), 6.55 (s, 1H), 6.45 (dd, J=16.8Hz, J=10.0Hz, 1H), 6.27 (dd, J=16.8Hz, J=1.6Hz, 1H), 6.00-5.97 (m, 1H), 5.78 (dd, J=10.0Hz, J=2.0Hz, 1H), 5.15-5.11 (m, 1H), 3.76 (s, 3H), 3.57-3.54 (m, 4H), 3.05-2.97 (m, 4H), 2.05 (s, 3H), 1.44 (d, J=6.4Hz, 6H) .HRMS (ESI) Calculated value C31H35N8O5[M+H]+599.2730 experiment value 599.2731.
N- (3- (7- ((4- (4- Acetylpiperazine -1- bases) -2- methoxyphenyls) amino) -3- benzyl -2,4- dioxos - 3,4- dihydro-pyrimidins simultaneously [4,5-d] pyrimidine -1 (2H)-yl) phenyl) acrylamide (compound 010)
1H NMR(400MHz,DMSO-d6) δ 10.36 (s, 1H), 8.87 (s, 1H), 8.69 (s, 1H), 7.87 (d, J= 5.6Hz, 1H), 7.68 (s, 1H), 7.48 (t, J=8.0Hz, 1H), 7.38 (d, J=6.8Hz, 2H), 7.33 (t, J=7.2Hz, 2H), 7.26 (t, J=7.2Hz, 1H), 7.17 (d, J=8.0Hz, 1H), 7.12 (d, J=7.8Hz, 1H), 6.55 (s, 1H), 6.44 (dd, J=16.8Hz, J=10.0Hz, 1H), 6.26 (dd, J=17.2Hz, J=2.0Hz, 1H), 5.98 (s, 1H), 5.77 (dd, J=10.0Hz, J=1.6Hz, 1H), 5.09 (s, 2H), 3.76 (s, 3H), 3.58-3.55 (m, 4H), 3.05- 2.97 (m, 4H), 2.05 (s, 3H) .HRMS (ESI) calculated values C35H35N8O5[M+H]+647.2730 experiment value 647.2735.
The specific synthetic method of compound 011 is as follows：
Reagent and condition：(a)-(e) is as previously described；(f) N, N, N'- trimethyl ethylenediamine, K2CO3,DMSO；(n)Zn, NH4Cl,CH3OH；(o) acyl chlorides, Et3N,CH2Cl2, 0 DEG C-room temperature.
7- ((4- ((2- (dimethylamino) ethyl) (methyl) amino -2- methoxyl group -5- nitrobenzophenones) amino) -3- first The synthesis of base -1- phenyl pyrimidines simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone
Weighing 7- ((the fluoro- 2- methoxyl groups -5- nitrobenzophenones of 4-) amino), simultaneously [4,5-d] is phonetic for -3- methyl-1s-phenyl pyrimidine Pyridine -2,4 (1H, 3H)-diketone (438mg, 1mmol), K2CO3(207mg, 1.5mmol), N, N, N'- trimethyl ethylenediamines (153mg, 1.5mmol) is added 4mL DMSO dissolvings, is warming up to 90 DEG C, stirs 4h in 10mL single-necked flasks.TLC tracks to original Material conversion, is cooled to room temperature, and ice water is added, and ethyl acetate extraction is collected organic phase, is spin-dried for, crude product is detached through silica gel column chromatography (DCM/CH3OH=20:1, v/v) 7- ((4- ((2- (dimethylamino) ethyl) (methyl) amino -2- methoxyl group -5- nitros, are obtained Phenyl) amino) -3- methyl-1s-phenyl pyrimidine simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone 358mg, yield 69%.1H NMR(400MHz,CDCl3)δ8.97(s,1H),7.88(s,1H),7.63(s,1H),7.46-7.43(m,3H),7.19-7.16 (m, 2H), 6.52 (s, 1H), 3.83 (s, 3H), 3.39 (s, 3H), 3.08 (t, J=6.4Hz, 2H), 2.71 (s, 3H), 2.41 (t, J=6.8Hz, 2H), 2.17 (s, 6H) .LC-MS:m/z:521.3(M+H)+.
7- ((5- amino -4- ((2- (dimethylamino) ethyl) (methyl) amino) -2- methoxyphenyls) amino) -3- first The synthesis of base -1- phenyl pyrimidines simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone
Weigh 7- ((4- ((2- (dimethylamino) ethyl) (methyl) amino -2- methoxyl group -5- nitrobenzophenones) amino) - 3- methyl-1s-phenyl pyrimidine simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone (312mg, 0.6mmol) in 50mL single-necked flasks, The dissolving of 8mL methanol is added, instills 1mL saturated ammonium chloride solutions, zinc powder (390mg, 6mmol) is added and is stirred overnight at room temperature.TLC with Track to raw material converts, and filters out zinc powder, and filtrate is spin-dried for, and crude product detaches (DCM/CH through silica gel column chromatography3OH=15:1, v/v) it, obtains 7- ((5- amino -4- ((2- (dimethylamino) ethyl) (methyl) amino) -2- methoxyphenyls) amino) -3- methyl-1s-benzene Yl pyrimidines simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone 180mg, yield 61%.1H NMR(400MHz,CDCl3)δ8.94(s, 1H), 8.11 (s, 1H), 7.60 (t, J=7.6Hz, 2H), 7.52 (t, J=7.2Hz, 1H), 7.33 (d, J=7.6Hz, 2H), 6.89 (s, 1H), 6.51 (s, 1H), 3.70 (s, 3H), 3.40 (s, 3H), 2.81 (t, J=6.4Hz, 2H), 2.52 (s, 3H), 2.29 (t, J=6.8Hz, 2H), 2.19 (s, 6H) .LC-MS:m/z:491.3(M+H)+.
N- (2- ((2- (dimethylamino) ethyl) (methyl) amino) -4- methoxyl groups -5- ((6- methyl -5,7- dioxos - 8- phenyl -5,6,7,8- tetrahydropyrimidines simultaneously [4,5-d] pyrimidine -2-base) amino) phenyl) and acrylamide synthesis (compound 011)
Weigh 7- ((5- amino -4- ((2- (dimethylamino) ethyl) (methyl) amino) -2- methoxyphenyls) amino) - 3- methyl-1s-phenyl pyrimidine simultaneously [4,5-d] pyrimidine -2,4 (1H, 3H)-diketone (147mg, 0.3mmol), DIPEA (77mg, 0.6mmol) in 25mL flasks, the dissolving of 3mL dichloromethane is added, is stirred 15 minutes under ice bath.It is another take acryloyl chloride (33mg, It 0.36mmol) is dissolved in 1mL dichloromethane, is added drop-wise in above-mentioned reaction solution, is stirred overnight at room temperature.TLC tracks raw material conversion, rotation Dry, crude product detaches (DCM/CH through silica gel column chromatography3OH=15:1, v/v) N- (2- ((2- (dimethylamino) ethyl) (first, is obtained Base) amino) -4- methoxyl groups -5- ((6- methyl -5,7- dioxo -8- phenyl -5,6,7,8- tetrahydropyrimidines simultaneously [4,5-d] pyrimidines - 2- yls) amino) phenyl) acrylamide 80mg, yield 49%.1H NMR(400MHz,CDCl3)δ9.36(s,1H),9.08(s, 1H), 7.88 (s, 1H), 7.50-7.46 (m, 5H), 6.58 (dd, J=17.2Hz, J=10.0Hz, 1H), 6.48 (dd, J= 17.2Hz, J=1.6Hz, 1H), 5.83 (dd, J=10.0Hz, J=2.0Hz, 1H), 3.83 (s, 3H), 3.48 (s, 3H), 3.23 (t, J=6.0Hz, 2H), 3.06 (t, J=6.4Hz, 2H), 2.77 (s, 6H), 2.68 (s, 3H) .HRMS (ESI):Calculated value C28H33N8O4(M+H)+545.2625 experiment value 545.2630.
2. biological activity test of embodiment
Compound provided by the invention is following to the extracorporeal extracorporeal suppression experiment of EGFR kinase activity to be carried out：
External enzyme activity assay：Wild type and various saltant types (T790M, L858/T790M) EGFR be purchased from Invitrogen.It is provided with from 5.1 × 10 for all compounds to be tested-11Mol/L to 1.0 × 10-610 of mol/L are dense Spend gradient.
The concentration of different kinases is determined by Optimal Experimental, corresponding a concentration of：EGFR(PV3872,Invitrogen) 0.287μg/μL,EGFR-T790M(PV4803,Invitrogen)0.174μg/μL,EGFR-L858R/T790M(PV4879, Invitrogen)0.055μg/μL.Compound is in DMSO from 5.1x10-9M to 1x10-4M dilutes three times.4 μ L compounds are dissolved in 96 μ L water, obtain the compound solution of 4x.40 μM of ATP are dissolved in 1.33x kinase buffer liquids, and kinases/peptide mixer swashs comprising 2x Enzyme, 4 μM of trorsine 14 peptides are ready to for use.10 μ L kinase reactions include 2.5 μ L compound solutions, 5 μ L kinases/peptide mixer, 2.5 μ L ATP solution.5 μ L Phosphorylated Peptide solution replace kinases/peptide mixer to be compareed as 100% phosphorylation.2.5μL1.33x Kinase buffer liquid replaces ATP solution to be used as 100% inhibition control, and 2.5 μ L 4%DMSO are used as 0% instead of compound solution and inhibit Control.It is cultivated at room temperature after solution is sufficiently mixed in plate 1.5 hours.After 5 μ L Development Solution are added per hole Continue to cultivate 1 hour at room temperature, non-phosphorylated peptide is cleaved within this time.Finally, 5 μ L are added and terminate preparation (Stop Reagent) reaction was completed.Orifice plate is measured with EnVisionMultilabel Reader (Perkin Elmer).Test number It is calculated according to GraphPad Prism version 4.0 are used.Experiment is repeated 3 times above every time.
Cell Proliferation and growth inhibition analysis：H1975 (non-small cell lung cancer cell, EGFRL858R/T790M), A431 it is (non-small Cell lung cancer cell, EGFR wild types), cell is obtained from ATCC.Cell-proliferation activity is assessed using MTS analytic approach. Cell exposure 72 hours under processing conditions, the used cell number of each experiment of each cell line is according to absorbance value (at 490nm Absorbance value be 1.3-2.2) be adjusted.6 concentration gradients (0.1nM-10 μM) are provided with for compound to be tested, each Concentration value at least uses 6 groups of parallel controls.
H1975, A431 cell are cultivated in corresponding culture medium, and cell at least passes on twice after recovery, is subsequently used for Experiment uses.The cell of logarithmic phase is by trypsin acting and settling flux in the medium.H1975 (per 1000 cell of hole), A431 is seeded in 96 orifice plates, 100 μ L of volume (per 2000 cell of hole)；6 groups of parallel and 7 row are set.Orifice plate is put in 37 DEG C 5% 2 In the incubator of carbonoxide overnight.Compound is dissolved in DMSO, compound concentration is 10 μM every liter, then gradually by compound concentration It is respectively every liter 10 μM, 1 μM, 0.1 μM, 0.01 μM, 0.001 μM, 0.0001 μM to dilute obtained compound concentration.2 μ L chemical combination Object solution is added in the culture medium of 998 μ L, and mixture is adequately mixed.The mixture of 100 μ L is added in 96 orifice plates.2μL DMSO It is used as 0% instead of compound solution to inhibit to compare.After culture 68 hours, 20 μ L MTT (5mg/mL) are added.4 as a child, throws It abandons supernatant and 150 μ L DMSO is added.After shake 10 minutes, orifice plate is read with Synergy HT (Bio TeK) (OD490) Data.Data are calculated using GraphPad Prism version 4.0, IC50Value is by using the non-of dose-effect curve Linear regression model (LRM) adjusts to obtain.
Test result is as follows shown in table 1.
Table 1
*：>100 indicate substantially to can't detect the inhibiting effect to cell.
It discusses：
Inventor after extensive and in-depth study, designs and synthesizes to have obtained a series of pyrimidos for having no document report [4,5-d] pyrimidine -2,4 (1H, 3H)-cyclohexadione compounds has carried out molecular level and cellular level to obtained compound Active testing obtains the compound that a batch is capable of selective depression EGFR T790M mutation.Present inventors have further discovered that this hair Proliferation Ability capacity variance of the bright compound to EGFR saltant types cancer cell (H1975) and EGFR wild types cancer cell (A431) Higher than to mutant egf R and Wild type EGFR kinase activity rejection ability difference, to prompt the compound of the present invention in vivo There is better difference toxicity, it is possible to become selective depression T790M mutation, overcome the third generation EGFR targeting medicines of clinical drug-resistant Object, or obtain activity more preferably and/or the basis of difference toxicity more preferably compound as through further modification.
All references mentioned in the present invention is incorporated herein by reference, independent just as each document It is incorporated as with reference to such.In addition, it should also be understood that, after reading the above teachings of the present invention, those skilled in the art can To be made various changes or modifications to the present invention, such equivalent forms equally fall within model defined by the application the appended claims It encloses.

Claims (9)

1. compound shown in general formula III or its pharmaceutically acceptable salt：
In formula,
R2It is selected from
R3Selected from C1-C6Alkyl；Or the C of phenyl substitution1-C6Alkyl；
R4、R5、R6、R7And R8It is independently selected from the following group：
2. compound as described in claim 1 or its pharmaceutically acceptable salt, which is characterized in that R3Selected from methyl, isopropyl Or benzyl.
3. compound as claimed in claim 1 or 2 or its pharmaceutically acceptable salt, which is characterized in that R5And R8For H；Or Person, R5、R7And R8For H.
4. compound selected from the group below or its pharmaceutically acceptable salt：
5. a kind of pharmaceutical composition, described pharmaceutical composition contains compound or its medicine described in any one of claim 1-4 Acceptable salt and pharmaceutically acceptable carrier or excipient on.
6. the compound described in any one of claim 1-4 is preparing the disease for treating or preventing EGFR and mediating, or inhibits Purposes in the drug of EGFR.
7. purposes as claimed in claim 6, which is characterized in that the disease that the EGFR is mediated is cancer.
8. purposes as claimed in claim 7, which is characterized in that the cancer is selected from the group：Non-small cell lung cancer, cellule lung Cancer, adenocarcinoma of lung, lung squamous cancer, breast cancer, prostate cancer, oophoroma, G. cephalantha, cervical carcinoma, the cancer of the esophagus, liver cancer, kidney, pancreas Gland cancer, colon cancer, cutaneum carcinoma, leukaemia, gastric cancer, multiple bone marrow cancer and solid tumor.
9. purposes as claimed in claim 8, which is characterized in that the solid tumor is neurogliocytoma or lymthoma.
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