CN103570616B

CN103570616B - N ' straight chains alkanoyl neighbour's pyridine hydrazide derivatives and its preparation method and pharmaceutical composition and purposes

Info

Publication number: CN103570616B
Application number: CN201210249757.5A
Authority: CN
Inventors: 冯志强; 陈晓光; 王克; 李燕; 秦爱芳; 唐克
Original assignee: Institute of Materia Medica of CAMS
Current assignee: Institute of Materia Medica of CAMS
Priority date: 2012-07-18
Filing date: 2012-07-18
Publication date: 2017-10-20
Anticipated expiration: 2032-07-18
Also published as: CN103570616A

Abstract

The present invention relates to the adjacent pyridine hydrazide derivatives of the straight chains of the N ' shown in Formulas I alkanoyl; its officinal salt; its hydrate and solvate; and preparation method thereof; composition containing this one or more compound, the purposes with such compound in terms of the disease relevant with protein kinase in treatment such as immune disorder and tumor disease.

Description

The adjacent pyridine hydrazide derivatives of N '-straight chain alkanoyl and its preparation method and pharmaceutical composition with Purposes

Invention field

The present invention relates to the adjacent pyridine hydrazide derivatives of the N ' shown in Formulas I-straight chain alkanoyl, its officinal salt, its hydrate And solvate, its polycrystalline and eutectic, the precursor or derivative of its same biological function, and preparation method thereof, containing one or The composition of this multiple compound, and such compound is in the treatment disease relevant with protein kinase such as immune disorder and tumour disease Purposes in terms of disease.

Background of invention

Recent years, due to the raising of the understanding of the biomolecule related to disease with some other to enzyme, greatly promote The discovery or development of the new drug for the treatment of disease are entered, protein kinase is exactly an a kind of widely studied important class, and it is one Extended familys, the control with intracellular various signal transduction processes is relevant.Due to they structure and catalysis conservative it Be considered as evolving from a common ancestral gene.Nearly all kinases is all containing a 250-300 similar ammonia Base acid catalysis domain.These protein kinases are divided into multiple families, such as protein tyrosine kinase, egg according to the difference of phosphorylated substrate White serine/threonine kinase, lipoid etc..Typically, protein kinase is by influenceing a phosphoryl to turn from a ribonucleoside triphosphote Move on to a protein receptor related to signal transduction pathway and carry out signal transduction in mediated cell.These phosphorylated events as point Sub switch adjusts the biological function of target protein, is finally excited and various extracellular and other stimulations are reacted.Kinases is present In multilayer signal transduction path, receptor tyrosine kinase is thin positioned at the upstream of Tumor Angiongesis Signal transduction pathway and tumour The upstream of born of the same parents' Signal transduction pathway.Serine/threonine protein kitase is located at tumour and the signal of Tumor Angiongesis cell turns The downstream of guiding path.Study and show, by blocking VEGFR and pdgf receptor in upstream, to block Raf/MEK/ERK in downstream, can The angiogenesis of tumour is reduced simultaneously and suppresses the duplication of tumour cell, so as to hinder the growth of tumour.

Raf kinases is the protein product encoded by proto-oncogene raf, is made up of 648 amino acid, molecular weight is 70000 Containing 3 conserved regions in~74000D, its structure, respectively CR1 (61~194D), CR2 (254~269D), CR3 (335~ 627D).CR1 is located at its molecule possessing amino, rich in cysteine, containing zinc-finger structure, the ligand binding domain with protein kinase C Structure is similar, is the main portions that the Ras of activation is combined with Raf-1 protein kinases.CR2 is also close to aminoterminal, rich in serine And threonine.CR3 is located at the c-terminus of its molecule, is the catalysis area of protein kinase.It is used as Ras/Raf/MEK/ERK paths In a Key kinases, Raf can rely on or independent of Ras by way of play its signal transduction adjustment effect.It is used as Raf The stream substrates of kinases, the MEK phosphorylated CREBs of activation adjust various cell functions.Once excessive activation occurs for the path, then Cell propagation is caused to accelerate to extend with the cells survival phase, so as to lead oncogenic generation.

Research shows, more than 80% oncogene and proto-oncogene are present in the cancer encoding proteins EGFR-TK of people（PTK） In, generation and the development of the various cancers of the mankind be it is relevant with the abnormal cell signal transduction for coming from protein tyrosine kinase, The increase that one of malignant cell is mainly characterized by tyrosine kinase activity.Therefore, the activation or blocking of EGFR-TK are suppressed Its signal transduction path turns into the new way of control tumour.

Endothelial growth factor receptor (EGFR) is a kind of protein tyrosine kinase receptor (RTK), positioned at No. 7 chromosome P13~q22 areas, total length 200kb is made up of 28 extrons, encodes 1 186 amino acid, its glycoprotein molecule amount is about 170kDa, is distributed widely in all histocytes in addition to ripe Skeletal Muscle Cell, parietal endoderm and hematopoietic tissue.EGFR There is the similar acceptor molecule of 4 structures in family：ErbB1 (EGFR), ErbB2 (HER2), ErbB3 (HER3), ErbtM (HER4), Belong to receptor tyrosine kinase (RTKS).They are all containing 1 extracellular ligand binding domains, 1 membrane spaning domain and 1 Cytoplasmic domain with tyrosine kinase activity.Its intracellular region and erbB oncoprotein very high homologies.EGFR activation The Receptor dimerization effect that can be induced by part is realized.In ErbB receptor family, in addition to HER2, other members have it Respective ligand, various parts are that, by corresponding transmembrane protein precursor passes through proteolysis, there is 1 EGF sample Domain.Include EGF (EGF), transforming growth factor α (TGF-α), two-way tune with the part that EGFR is specifically bound Save albumen (AR), beta cell plain (BTC), Heparin-binding EGF like growth factors (HB-EGF), epiregulin (EPR) etc..It is extracellular Cause ErbB2 configurations to change after ligands, EGF (endothelial growth factors) and ErbB2 specific binding, cause Receptor dimerization so as to Activate their cytoplasmic location.After ErbB2 intracellular region tyrosine phosphorylation so activate second messenger transduction, pass through MAPK The activation (regulation kinases Erkl and Er1) of (mitogen protein kinase) approach inducing cell external signal：Pass through PDK (phosphatidyl-4s Alcohol kinases) pathway activation signal transducer JAK；Further start STAT1, STATS3 transcription activators；On the other hand, cell Interior signal activates the ERK (extracellular regulated protein kinase), Jin Erjie in downstream by Grb2 (growth factor receptor binding protein precursor) Lead ATF, NF-kB, Ap-1, C-fos and C-Jun transcription activating.These are all the growths that EGFR is mediated or carcinogenic Basic downstream pathway.Abnormal EGFR activation mechanisms include acceptor amplification in itself, the overexpression of receptors ligand, Activating mutations with And negativity adjusts the shortage of approach, therefore EGFR induced cancers are at least through 3 kinds of mechanism：The overexpression of EGFR parts, EGFR's Amplification or EGFR mutation activation.In this 3 kinds of mechanism, EGFR mutation activation is to cause tumour cell aberrant biological behavior Main factor.Some mutation of EGFR gene can cause the enhancing of acceptor effect and the extension of duration.Lynch etc. is proved Become isoreceptor and have no effect on the stability of receptor protein, activated and found by Tyr1068 phosphorylation assay EGFR, wild type by The activation 15min of body is to lower, and becomes the effect that isoreceptor shows 2 times higher than normal EGFR, and the continuous activation more than 3h.

EGFR mutation do not have an impact the ability that tumour cell is combined with TKI (tyrosine kinase inhibitor).TKI is to those The reason for EGFR is activated is caused to be explained by oncogeneaddiction models because of mutation.Pass through Ras.Raf- MEK.ERK1/ERK2, PI3K.Akt, STAT3/STAT5 path, EGFR discontinuity heights activation downstream signal, start EGFR regulations Anti-apoptotic and survival signaling, cause cancer cell to become dependent upon this signal to maintain it to survive -- i.e. with oncogene (mutation The feature that EG is relied on；After EGFR signals are blocked using specificity T KI, its proliferative effects and output survival signaling will be eliminated, Cause death of neoplastic cells.Result, it is believed that the variation of signal transduction pathway is that the high sensitive basis of medicine occur in cancer cell. On the contrary, the tumour cell (reactionless to Gefitinib, Erlotinib) that normal cell or non-EGFR are relied on is unaffected.Because Existence is also driven by other genes, or can be made up after EGFR suppression by other RTK.In oncogene relies on model, The oncogene that cell carcinomas is relied on can produce the output of 2 signals of apoptosis and existence simultaneously.Under general Sui condition, oncogene is swashed It is living.Survival signaling is occupied an leading position, and apoptotic signal is in low relative levels, cancer cell is maintained growth and is bred.Work as cancer It is that existence significantly weakens rapidly first in crucial window phase after the acute inactivation of gene.And apoptotic signal slowly declines.Cause This causes signal uneven (apoptotic signal accounts for leading), and irreversible apoptosis occurs for active cell.Study discovery tyrosine-kinase Enzyme inhibitor Gefitinib (gefitinib)/Tarceva (Erlotinib) treatment NSCLC patient, about 10% patient performance Go out rapid and satisfied clinical effectiveness, further study show that these patient's overwhelming majority have EGFR genetic mutation.Current The known gene mutation relevant with EGFR-TKI (endothelial growth factor receptor tyrosine kinase inhibitor) is confined to following several Kind：G719X (18 extron), E746-A450 lack (19 extron), L858R (21 extron), L861Q (21 extron), T790M (20 extron) and D770-N771 (20 extron).Wherein E746A450 missings and L858R mutation and TKI curative effect Height correlation.Analysis results of Mitsudomi T, the Yatabe Y to 568 Patients with Non-small-cell Lung：In all non-small cells About 90% EGFR genetic mutation is concentrated in 19 or 21 extrons in patients with lung cancer, wherein the deletion mutation of 19 extrons and The effective percentage that the patient of point mutation in 21 extrons takes EGFR-TKI reaches more than 70%.Recent research prompting, The slotting human nature mutation (D770-N771) of EGFR extron 20s can make acceptor reduce by 100 times to EGFR-TKI sensitiveness, face It has also been found that the patient with this mutation is not obvious to EGFR-TKI therapeutic responses on bed.The amplified production of extron 20 is carried out Subcloning analysis finds that T79OM mutation are that a base-pair occurs from cytidine (C) changing to thymidine (T) Become, the threonine for being exactly the site of EGFR tyrosine kinase domains 790 in protein level is replaced (T790M) by methionine, this Mutation can be such that EGFR in the state that is activated, causes again so that the reason for causing TKI acquired resistance, resistance is mutation EGFR structures change, and make TKI is in connection steric effect occur.

There is the reason for research prompting KRAS is probably Gefitinib, Erlotinib initial drug-resistant.Helena The TKI therapeutic effects of 1008 NSCLC patients are summarized in linardou Meta analyses, in occur K-ras mutation 165 In patient, 94% patient is treated without significant reaction to TKI.In general, KRAS and EGFR mutation NSCLC is excluded each other There is notable difference in different tumors subtypes：EGFR mutation are mainly seen in non-smoker, and KRAS is more often seen The related cancer of smoking.Because KRAS is always betided in the NSCLC with Wild type EGFR, it is difficult to differentiate between pair EGFR-TKI insensitive is mutated because of KRAS, or because of without EGFR on earth.

Vascular endothelial growth factor receptor (vascular endothelial growth factor receptor, VEGFR) family includes 3 kinds of hypotypes, i.e.,:VEGFR-1 (while Flt-1 can also be write), VEGFR-2 (KDR/Flk-1) and VEGFR-3 (Flt 1), in addition, also 1 and 2 two cooperative expert systems of neuropilin (neuropilin).Wherein VEGFR-1 Vascular endothelial cell, candidate stem cell, macrophage and monocyte are mainly distributed on, can be with VEGF-A, VEGF-B and P1GF With reference to mainly relevant with the growth regulating of candidate stem cell.VEGFR-2 is mainly distributed on vascular endothelial cell and lymphatic endothelia is thin In born of the same parents, it can be combined with VEGF-A, VEGF-C, VEGF-D, VEGF-E.VEGF stimulating endothelial cells propagation, increase vascular permeability Property and new vascular generation effect it is main by combining and activation VEGFR-2 realizes compared with VEGFR-2, VEGFR-1 with VEGF affinity is high 10 times, but adjusts the active much lower of endothelial cell, it may be possible to have negative regulation to VEGFR-2 activity Effect.VEGFR-3 is mainly expressed in lymphatic endothelial cells, can be combined with VEGF-C and VEGF-D, regulates and controls lymphatic endothelium Growth.

Research shows：When diameter of tumor is more than 2mm, it is desirable to have new vessels is useless to provide nutriment and excretion metabolism Thing.VEGF/VEGFR signal paths serve in tumor vascular generation it is key, can pass through block or interference VEGF/ VEGFR signal paths suppress the new life of blood vessel, with the curative effect for the growth for reaching control tumour.With traditional cytotoxic drug phase Than the antineoplastic using VEGF/VEGFR-2 as target has very big advantage under normal physiological conditions, and angiogenesis only exists Worked in the physiological activity such as wound healing and menstrual cycle, so tumour is treated using anti-angiogenic medicaments, to human body poison Property effect it is small, vascular endothelial cell is directly contacted with blood, make medicine be more prone to reach action site pass through it is current right The understanding of VEGF/VEGFR signal path mechanism of action, can obtain following several possible inhibitor research directions：A. utilize Monoclonal antibody suppresses VEGF or VEGFR, prevents it from specifically binding, disabling signal conduction.Gene can certainly be utilized Technology suppresses their expression, weakens its activity.B. specific micromolecular inhibitor is designed, the extracellular VEGF knots of VEGFR are attached to Close region, competitive antagonism VEGF similarly or is attached to the particular combination domain of VEGFR on VEGF, competitive antagonism VEGFR.C. VEGFR intracellular kinase domain, mainly ATP binding site are suppressed, competitively antagonism ATP, makes it not carry Phosphorus supply acidic group.D. the critical proteins for suppressing the VEGFR downstream signals of intracellular considers the compliance of patient, can be oral it is small Molecule inhibitor may have good prospect.

Platelet derived growth factor (platelet.derived growth factor, PDGF) is induction and promotes blood One of most strong, most single-minded angiogenesis factor of pipe formation effect.PDGF mainly by being combined with pdgf receptor (PDGFR), enters And activated protein kinase signal transduction pathway and play a role.PDGFR is made up of two kinds of subunits of α and β, has 3 kinds of dimers (PDGFR- α α, α β, β β), wherein β β dimerization receptor body (PDGFR- β) are mostly important, and its molecular weight is about 180~190ku, category In tyrosine kinase receptor (receptor tyrosine kinase, RTK) family.PDGFR is in tumour formation and development process In also play an important role.PDGFR- β overexpression or overactivity can stimulate intratumoral vasculature to generate, and promote tumour Growth.PDGFR- β are one of molecular markers of tumor vascular endothelial cell, the high expression in endothelial cells in tumor neogenetic blood vessels, And it is closely related with the growth, transfer and prognosis of some tumours.So PDGFR- β are an ideal neoplasm targeted therapies Target.

Raf kinases and its Raf/MEK/ERK paths of mediation have remarkable effect in tumour progression and transfer process, and Include EGF (EGF), VEGF (VEGF) and platelet growth factor with many growth factors (PDGF) it is etc. closely related.People have thought a variety of methods to adjust this path, including the farnesyl for suppressing Ras albumen Change, suppress the expression of Rat "-I kinases (also referred to as C-RAF kinases), suppress the activity of Raf kinases and MEK kinases.Above-mentioned method Not only inhibit ERK signal transduction but also successfully inhibit the growth of xenograft tumours.In addition, existing evidence is shown, greatly Partial tumors are not dominated by single signal conduction path, and bigger curative effect may be obtained by carrying out suppression for Mutiple Targets.

Many diseases are that the abnormal cell effect triggered with protein kinase mediated event is associated.These disease bags Include, but be not limited to, tumour, inflammation disease, immunological diseases, bone disease, metabolic disease, sacred disease, cardiovascular and cerebrovascular disease, hormone Related disease etc..Consequently found that being very important with searching kinases inhibitor as medicine.Although many hairs It is bright that very big contribution has been made to this area, but to improve medication effect, this area still is continuing to study.

The content of the invention

It is an object of the invention to provide the adjacent pyridine hydrazide derivatives of the N ' shown in formula I-straight chain alkanoyl, its is pharmaceutically acceptable Salt, its solvate, its prodrug, its polycrystalline or eutectic.

Another object of the present invention is to provide the system of the adjacent pyridine hydrazide derivatives of N '-straight chain alkanoyl shown in formula I Preparation Method.

Spread out it is still another object of the present invention to provide a kind of adjacent pyridine hydrazides of N ' containing shown in formula I-straight chain alkanoyl Biological pharmaceutical composition.

A further object of the present invention be provide such compound in anticancer, and with the medicine of protein kinase related disorder Purposes.

In order to complete the purpose of the present invention, it can adopt the following technical scheme that：

The present invention is related to the adjacent pyridine hydrazide derivatives of the having structure N ' shown in formula I-straight chain alkanoyl：

Or its officinal salt, its hydrate and solvate, its polycrystalline and eutectic, the precursor of its same biological function or spread out It is biological.

The invention also discloses the method for preparing the compounds of this invention, including following route steps：

The method for preparing the compound of claim 1, comprises the following steps：

Route 1

Step（a）In with hydrazides 1 be raw material, be readily available with common method and acyl chlorides or acid anhydrides or acid reaction N '- Straight chain alkanoyl hydrazide derivatives 2.

Step（b）In, with para hydroxybenzene amine under alkaline environment by obtaining compound to the chlorine substituted ether in hydrazides 2 3。

Step（c）In can pass through the chloro- 3- 5-trifluoromethylanilines condensation generation urea derivative I of CDI and 4-;Also can be chloro- with 4- 3- trifluoromethylbenzenes based isocyanate obtains urea derivative I by nucleophilic addition；Also can be with the chloro- 3- trifluoromethyls phenylaminos of 4- Formic acid 4- nitros phenyl ester obtains urea derivative I by nucleophilic substitution.

Route 2

Step（a）In, it is raw material with ester 4 or acyl chlorides 5, bishydrazide derivative 2 is obtained with N '-reaction of straight chain alkane hydrazides 6.

Step（b）In, PAP is under alkaline environment by being obtained to the chlorine substituted ether in bishydrazide derivative 2 Compound 3.

Route 3

Step（a）In, phenolic compounds 7 is under alkaline environment by being obtained to the chlorine substituted ether in bishydrazide derivative 2 Urea derivative I.Step（b）In, ester compounds 8 and N '-reaction of straight chain alkane hydrazides 6 is similarly obtained urea derivative I.

Route 4

The route is raw material with hydrazides 9, is held it very much with straight chain fatty acid or acyl chlorides or anhydride reaction with common method It is easy to get to urea derivative I.

In addition, the initiation material and intermediate in above-mentioned reaction are readily obtained, or to those skilled in the art Synthesis can be easy to the conventional method in organic synthesis.

The adjacent pyridine hydrazide derivatives of N ' described in Formulas I-straight chain alkanoyl can be deposited in the form of solvate or non-solvent compound Crystallization, which is being carried out, using different solvents is likely to be obtained different solvates.Pharmaceutically acceptable salt described in Formulas I is included not The acid-addition salts of same acid-addition salts, such as following inorganic acid or organic acid：Hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, methanesulfonic acid, to toluene Sulfonic acid, trifluoroacetic acid, matrimony vine acid, maleic acid, tartaric acid, fumaric acid, citric acid, lactic acid.Within the scope of the present invention it is all this A little salt can all be prepared using conventional method.In the adjacent pyridine hydrazide derivatives of described N '-straight chain alkanoyl and its solvate and In the preparation process of its salt, different crystallization conditions are likely to occur polycrystalline or eutectic.

The invention further relates to the pharmaceutical composition using the compounds of this invention as active ingredient.The pharmaceutical composition can basis It is prepared by method well known in the art.Can be by by the compounds of this invention and one or more pharmaceutically acceptable solids or liquid Excipient and/or assistant agent are combined, and any formulation used suitable for human or animal is made.The compounds of this invention is in its pharmaceutical composition In content be usually 0.1-95 weight %.

The compounds of this invention or pharmaceutical composition containing it can be administered in a unit, and method of administration can be enteron aisle Or non-bowel, such as oral, intravenous injection, intramuscular injection, hypodermic injection, nasal cavity, oral mucosa, eye, lung and respiratory tract, skin, Vagina, rectum etc..

Form of administration can be liquid dosage form, solid dosage forms or semisolid dosage form.Liquid dosage form can be solution（Including True solution and colloidal solution）, emulsion（Including o/w types, w/o types and emulsion）, supensoid agent, injection（Including liquid drugs injection, powder-injection And transfusion）, eye drops, nasal drop, lotion and liniment etc.；Solid dosage forms can be tablet（Including ordinary tablet, enteric coatel tablets, lozenge, Dispersible tablet, chewable tablets, effervescent tablet, oral disnitegration tablet）, capsule（Including hard shell capsules, soft capsule, capsulae enterosolubilis）, granule, dissipate Agent, micropill, dripping pill, suppository, film, paster, gas（Powder）Mist agent, spray etc.；Semisolid dosage form can be ointment, gel Agent, paste etc..

It is sustained release preparation, controlled release preparation, targeting preparation and various that the compounds of this invention, which can be made ordinary preparation, also be made, Particulate delivery system.

In order to which the compounds of this invention is made into tablet, various excipient well known in the art can be widely used, including it is dilute Release agent, binder, wetting agent, disintegrant, lubricant, glidant.Diluent can be starch, dextrin, sucrose, glucose, breast Sugar, mannitol, sorbierite, xylitol, microcrystalline cellulose, calcium sulfate, calcium monohydrogen phosphate, calcium carbonate etc.；Wetting agent can be water, second Alcohol, isopropanol etc.；Adhesive can be starch slurry, dextrin, syrup, honey, glucose solution, microcrystalline cellulose, Arabic gum Slurry, gelatine size, sodium carboxymethylcellulose, methylcellulose, hydroxypropyl methyl cellulose, ethyl cellulose, acrylic resin, card Ripple nurse, polyvinylpyrrolidone, polyethylene glycol etc.；Disintegrant can be dried starch, microcrystalline cellulose, low substituted hydroxy-propyl fiber Element, PVPP, Ac-Di-Sol, sodium carboxymethyl starch, sodium acid carbonate and citric acid, polyoxy second Alkene sorbitan fatty acid ester, dodecyl sodium sulfate etc.；Lubricant and glidant can be talcum powder, silica, tristearin Hydrochlorate, tartaric acid, atoleine, polyethylene glycol etc..

Tablet can also be further made to coating tablet, such as sugar coated tablet, thin membrane coated tablet, enteric coated tablets, or double Synusia and multilayer tablet.

In order to which administration unit is made into capsule, active ingredient the compounds of this invention can be mixed with diluent, glidant Close, mixture is placed directly within hard shell capsules or soft capsule.Also can active ingredient the compounds of this invention is first and diluent, bonding Particle or micropill is made in agent, disintegrant, then is placed in hard shell capsules or soft capsule.For preparing each dilute of the compounds of this invention tablet Release the capsule that agent, binder, wetting agent, disintegrant, glidant kind can also be used for preparing the compounds of this invention.

For the compounds of this invention is made into injection, water, ethanol, isopropanol, propane diols or their mixture can be used Make solvent and add appropriate solubilizer commonly used in the art, cosolvent, pH adjustments agent, osmotic pressure regulator.Solubilizer or hydrotropy Agent can be poloxamer, lecithin, hydroxypropyl-β-cyclodextrin etc.；PH adjustment agent can be phosphate, acetate, hydrochloric acid, hydrogen Sodium oxide molybdena etc.；Osmotic pressure regulator can be sodium chloride, mannitol, glucose, phosphate, acetate etc..Such as prepare freeze-dried powder Injection, can also add mannitol, glucose etc. as proppant.

In addition, if desired, colouring agent, preservative, spices, flavouring or other additions can also be added into pharmaceutical preparation Agent.

To reach medication purpose, strengthen therapeutic effect, medicine of the invention or pharmaceutical composition known can be given with any Prescription method is administered.

The dosage of the compounds of this invention pharmaceutical composition is according to the property and serious journey to be prevented or treated disease The individual instances of degree, patient or animal, method of administration and formulation etc. can have large-scale change.In general, the present inventionization The daily Suitable dosage ranges of compound are 0.001-150mg/Kg body weight, preferably 0.01-100mg/Kg body weight.Above-mentioned dosage With a dosage unit or several dosage unit administrations can be divided into, this depends on the clinical experience of doctor and including with other The dosage regimen for the treatment of means.

The compound or composition of the present invention can individually be taken, or merge with other treatment medicine or symptomatic drugs and use. When the compound of the present invention exists with other medicines to act synergistically, its dosage should be adjusted according to actual conditions.

The compounds of this invention is Mutiple Targets kinases inhibitor or its precursor, and these protein kinases are according to phosphorylated substrate Difference be divided into multiple families, such as protein tyrosine kinase, Protein Serine/threonine kinase, lipoid etc..Typically, albumen Kinases is transferred to a protein receptor related to signal transduction pathway by one phosphoryl of influence from a ribonucleoside triphosphote Carry out signal transduction in mediated cell.These phosphorylated events adjust the biological function of target protein as molecular switch, are finally swashed Hair is reacted to various extracellular and other stimulations.Kinases is present in multilayer signal transduction path, receptor tyrosine kinase Positioned at the upstream of Tumor Angiongesis Signal transduction pathway and the upstream of tumour cell Signal transduction pathway.Serine/threonine Protein kinase is located at the downstream of the Signal transduction pathway of tumour and Tumor Angiongesis cell.Research shows by blocking in upstream VEGFR and pdgf receptor, block Raf/MEK/ERK in downstream, and the angiogenesis of tumour can be reduced simultaneously and to suppress tumour thin The duplication of born of the same parents, so as to hinder the growth of tumour.The compounds of this invention has higher bioavilability, is disliked available for a variety of mankind The treatment of property tumour, including described tumor disease is liver cancer, kidney, lung cancer, cancer of pancreas, colorectal cancer, carcinoma of urinary bladder and mammary gland Cancer, oophoroma, squamous cell carcinoma, glioma, leukaemia, incidence cancer.

Embodiment

Invention is described further below with reference to embodiment, but not limit the scope of the invention.Determining instrument：Core Nuclear magnetic resonance spectroscopy Vaariaan Mercury300 or 400 type NMRs.Mass spectrum ZAD-2F and VG300 mass spectrographs.

Embodiment 1.1- (the chloro- 3- trifluoromethyls of 4-) -3- (4- (2- (2- (positive butyryl) hydrazine carbonyl) pyridine -4- oxygen Base) phenyl) urea

The synthesis of N '-positive bytyry -4- chloropyridine -2- hydrazides

Butyric acid 0.6g (7.0mmol) is dissolved in 3mLDMF, raw material 4- chloropyridine -2- hydrazides 2.1g (5.8mmol) are added, HATU2.7g (7.0mmol), triethylamine 1.8g (17.4mmol), is stirred at room temperature, and TLC monitoring raw material reactions are complete, add 100mL Water, has a large amount of off-white powders to separate out, obtains product 1.39g.¹H NMR(400MHz,DMSO-d₆):10.51(s,1H,-CONH-), 9.98(s,1H,-CONH-),8.66(d,1H,Ar-H),8.03(d,1H,Ar-H),7.81(dd,1H,Hz,Ar-H),2.16(t, 2H,-COCH₂-),1.56(m,2H,-CH₂-Me),0.92(t,3H,-CH₃).MS(FAB):(M⁺+1=242).

The synthesis of N '-positive bytyry -4- (p-aminophenyl epoxide) pyridine -2- hydrazides

Para-aminophenol 295mg (2.7mmol) is dissolved in DMF5mL, nitrogen protection is lower to add potassium tert-butoxide 416mg (3.4mmol), is stirred at room temperature after 3h, adds intermediate N '-positive bytyry -4- chloropyridine -2- hydrazides 0.5g (2.1mmol), carbon Sour potassium 138mg (1.3mmol), outer 80 °C of bath after TLC monitoring intermediate reactions are complete, adds 100mL water, ethyl acetate extraction 3 It is secondary, merge organic phase, saturated common salt is washed once, and anhydrous sodium sulfate drying is filtered afterwards, filtrate concentration obtains black solid 0.47g。¹H NMR(400MHz,DMSO-d₆):8.66(d,1H,Ar-H),8.03(s,1H,Ar-NH),7.81(dd,1H,Ar- H),7.71(d,2H,Ar-H),6.90(d,2H,Ar-H),2.16(t,2H,-CO-CH₂-),1.57(m,2H,-CH₂-Me),0.92 (t,3H,-CH₃).MS(FAB):(M⁺+1=315).

1- (the chloro- 3- trifluoromethyls of 4-) -3- (4- (2- (2- (positive butyryl) hydrazine carbonyl) pyridine -4- epoxides) phenyl) urea

By 1.12g（6.9mmol）CDI is dissolved in the dichloromethane of 7mL dryings, and beginnings solution is white opacity, will be dissolved with 1.2g（6.2mmol）The 10mL dichloromethane solutions of the chloro- 3- 5-trifluoromethylanilines of 4-, are instilled in above-mentioned solution, solution gradually becomes Clarification, is stirred at room temperature after 8h, adds dissolved with 0.66g（2.1mmol）N '-positive bytyry -4- (p-aminophenyl epoxide) pyridine -2- acyls The dichloromethane solution 5mL of hydrazine, is heated to reflux after 10h, stops reaction, and column chromatography separates object 0.26g.¹H NMR (300MHz,DMSO-d₆):δ(ppm):10.38(s,1H,-NHCO),9.94(s,1H,-NHCO-),9.27(s,1H,- NHCO-),9.05(s,1H,-NHCO-),8.55(d,1H,ArH),8.12(s,1H,ArH),7.68~7.63(m,2H,ArH), 7.60(d,2H,ArH),7.37(d,1H,ArH),7.21~7.19(m,3H,ArH),2.13(t,2H,-CH₂),1.59~1.49 (m,2H,-CH₂),0.90(q,3H,-CH₃).MS(FAB):(M⁺+1=536).

Or by 200mg（0.43mmol）Compound 1- (the chloro- 3- trifluoromethyls of 4-) -3- (4- (2- (hydrazine carbonyl) pyrroles Pyridine -4- epoxides) phenyl) urea is dissolved in 7mlTHF, adds 0.09mL（0.64mmol）TEA, is added dropwise to 2ml dissolved with 0.05ml （0.52mmol）After the THF solution of butyl chloride, backflow 4h, there is white solid generation, stop reaction, filtering, a small amount of tetrahydrochysene furan Mutter and wash, wash, drying is similarly obtained 1- (the chloro- 3- trifluoromethyls of 4-) -3- (4- (2- (2- (positive butyryl) hydrazine carbonyl) pyrroles Pyridine -4- epoxides) phenyl) urea, white solid 80mg.

Embodiment 2.1- (4- (2- (2- acethydrazides carbonyl) pyridine -4- epoxides) phenyl) -3- (chloro- 3- trifluoromethylbenzenes of 4- Base) urea

N-butyric acie is replaced using acetic acid, is carried out with reference to the operation of embodiment 1, target compound is obtained for white solid 175mg。¹HNMR(300MHz,DMSO-d₆):δ(ppm):10.38(s,1H,-NHCO-CH₃),9.99(s,1H,-NHCO-), 9.21(s,1H,-NHCO-),9.00(s,1H,-NHCO-),8.53(d,1H,ArH),8.11(s,1H,ArH),7.67~7.62 (m,2H,ArH),7.59(d,2H,ArH),7.36(d,1H,ArH),7.20~7.17(m,3H,ArH),1.88(s,3H,-CH₃) .MS(FAB)（M⁺+1=508）

Embodiment 3.1- (4- (2- (2- propionyl hydrazines carbonyl) pyridine -4- epoxides) phenyl) -3- (chloro- 3- trifluoromethylbenzenes of 4- Base) urea

N-butyric acie is replaced using n Propanoic acid, is carried out with reference to the operation of embodiment 1, target compound is obtained for white solid 135mg。¹H NMR(400MHz,DMSO-d₆):δ(ppm):10.37(s,1H,-NHCO-CH₂-),9.94(s,1H,-NHCO-), 9.22(s,1H,-NHCO-),9.01(s,1H,-NHCO-),8.55(d,1H,ArH),8.12(s,1H,ArH),7.68~7.63 (m,2H,ArH),7.60(d,2H,ArH),7.37(d,1H,ArH),7.20~7.18(m,3H,ArH),2.16(q,2H,- CH₂-),1.04(t,3H,-CH₃).MS(FAB)（M⁺+1=522）

Embodiment 4.1- (4- (2- (2- valeryl hydrazines carbonyl) pyridine -4- epoxides) phenyl) -3- (chloro- 3- trifluoromethylbenzenes of 4- Base) urea

N-butyric acie is replaced using positive valeric acid, is carried out with reference to the operation of embodiment 1, target compound is obtained for white solid 145mg。¹H NMR(400MHz,DMSO-d₆):δ(ppm):10.37(s,1H,-NHCO-CH₂-),9.94(s,1H,-NHCO-), 9.34(s,1H,-NHCO-),9.10(s,1H,-NHCO-),8.54(d,1H,ArH),8.12(s,1H,ArH),7.68~7.63 (m,2H,ArH),7.61(d,2H,ArH),7.37(d,1H,ArH),7.20~7.18(m,3H,ArH),2.16(t,2H,- CH₂-),1.51(m,2H,-CH₂-),1.32(m,2H,-CH₂-),0.89(t,3H,-CH₃).MS(FAB)（M⁺+1=550）

Embodiment 5.1- (4- (2- (the pungent formylhydrazine carbonyls of 2-) pyridine -4- epoxides) phenyl) -3- (chloro- 3- trifluoromethyls of 4- Phenyl) urea

N-butyric acie is replaced using caprylic acid, is carried out with reference to the operation of embodiment 1, target compound is obtained for white solid 115mg。¹H NMR(400MHz,DMSO-d₆):δ(ppm):10.37(s,1H,-NHCO-CH₂-),9.94(s,1H,-NHCO-), 9.22(s,1H,-NHCO-),9.00(s,1H,-NHCO-),8.54(d,1H,ArH),8.12(s,1H,ArH),7.67~7.63 (m,2H,ArH),7.60(d,2H,ArH),7.37(d,1H,ArH),7.20~7.18(m,3H,ArH),2.14(t,2H,- CH₂-),1.54~1.51(m,2H,-CH₂-),1.26(brs,8H,4×-CH₂-),0.86(t,3H,-CH₃).MS(FAB)（M⁺+1= 592）

Embodiment 6.1- (4- (2- (the own formylhydrazine carbonyls of 2-) pyridine -4- epoxides) phenyl) -3- (chloro- 3- trifluoromethyls of 4- Phenyl) urea

N-butyric acie is replaced using n-caproic acid, is carried out with reference to the operation of embodiment 1, target compound is obtained for white solid 155mg。¹H NMR(400MHz,DMSO-d₆):δ(ppm):10.37(s,1H,-NHCO-CH₂-),9.94(s,1H,-NHCO-), 9.24(s,1H,-NHCO-),9.02(s,1H,-NHCO-),8.54(d,1H,ArH),8.12(s,1H,ArH),7.67~7.63 (m,2H,ArH),7.60(d,2H,ArH),7.37(d,1H,ArH),7.20~7.18(m,3H,ArH),2.15(t,2H,- CH₂-),1.54~1.51(m,2H,-CH₂-),1.26(brs,4H,2×-CH₂-),0.86(t,3H,-CH₃).MS(FAB)（M⁺+1= 564）

Pharmacological activity

External activity is evaluated：

Mtt assay determines tumor cell survival

It is 0.8 ~ 2 × 10 that concentration is configured to after the cell of exponential phase is digested with pancreatin⁴Cell/ml cell liquid, 96 orifice plates are inoculated in by 1000/hole, add 100 μ l per hole.It is new that next day addition medicine containing various concentrations and coordinative solvent are compareed Fresh culture medium, adds 100 μ l per hole（DMSO final concentrations<0.5%）, set 5~7 dosage groups per medicine, every group at least set three it is parallel Hole, continues to cultivate after 120hr in 37 DEG C, abandons supernatant, and the MTT containing 0.5mg/ml of 100 μ l Fresh serum-free training is added per hole Base is supported, continues to cultivate 4hr, abandons culture supernatant, adds 200 μ l DMSO to dissolve MTT first hairpin precipitation per hole, is vibrated with microoscillator Mix, OD value (OD) is determined under the conditions of reference wavelength 450nm, Detection wavelength 570nm with MK3 types ELIASA, with solvent The tumour cell of control treatment is control group, and inhibiting rate of the medicine to tumour cell is calculated with formula below, and by middle efficacious prescriptions journey Calculate IC₅₀：

MTT the selection results

Activity in vivo is evaluated：

Influence to Renca tumor-bearing mice tumour growths

Experimental animal enters in SPF grades of environment that breeding observing is after 24 hours, experiment without exception of being allowed for access.It will exist in advance The Renca knurl liquid of Balb/c kinds mouse peritoneal recovery, 1 is pressed with sterile physiological saline：3 dilution proportions.Dilution is inoculated in Experiment mice left fore is subcutaneous, every injection dilution posterior tuberosity liquid 0.2mL.After all animal injections are finished, empirically require random Packet, every group 8.

Start administration after being inoculated with 24 hours, once a day, gavage is administered 12 times altogether, is put to death in being dislocated after last dose Animal, strips tumor tissues, and weigh.

To mice-transplanted tumor Renca growth inhibition effect

*P<005;**P<001;***P<0001, compared with negative control group.

Claims

1. compound and its officinal salt shown in Formulas I,

In formula:N is selected from 2,4,6,8,10,12,14,16.

2. the compound according to claim 1, it is characterised in that described officinal salt is selected from：Hydrochloride, hydrobromate, phosphorus Hydrochlorate, sulfate, mesylate, tosilate, acetate, trifluoroacetate, salicylate, amino-acid salt, matrimony vine acid Salt, maleate, tartrate, fumarate, citrate, lactate.

3. preparing the method for the compound of formula I of claim 1, comprise the following steps：

1) route 1

2) route 2

3) route 3

4) route 4

4. according to the preparation method of claim 3 route 1, it is characterised in that with hydrazides 1 be raw material in step (a), with acyl chlorides or Acid anhydrides or acid reaction obtain N '-straight chain alkanoyl hydrazide derivatives 2；In step (b), lead to para hydroxybenzene amine under alkaline environment Cross and compound 3 is obtained to the chlorine substituted ether in hydrazides 2；It is condensed in step (c) by the chloro- 3- 5-trifluoromethylanilines of CDI and 4- Compound shown in production I；Or obtained with the chloro- 3- trifluoromethylbenzenes based isocyanates of 4- by nucleophilic addition shown in Formulas I Compound；Or chemical combination shown in Formulas I obtained by nucleophilic substitution with 4- chloro- 3- trifluoromethyls phenylamino formic acid 4- nitros phenyl esters Thing.

5. it is raw material with ester 4 or acyl chlorides 5 according to the preparation method of claim 3 route 2, it is characterised in that in step (a), with N '-reaction of straight chain alkane hydrazides 6 obtains bishydrazide derivative 2；In step (b), PAP is under alkaline environment by double Chlorine substituted ether in hydrazide derivatives 2 obtains compound 3；Contracted in step (c) by the chloro- 3- 5-trifluoromethylanilines of CDI and 4- Compound shown in symphysis into Formulas I；Or Formulas I institute obtained by nucleophilic addition with 4- chloro- 3- trifluoromethylbenzenes based isocyanates Show compound；Or chemical combination shown in Formulas I obtained by nucleophilic substitution with 4- chloro- 3- trifluoromethyls phenylamino formic acid 4- nitros phenyl esters Thing.

6. according to the preparation method of claim 3 route 3, it is characterised in that in step (a), phenolic compounds 7 is under alkaline environment By obtaining compound shown in Formulas I to the chlorine substituted ether in bishydrazide derivative 2；In step (b), ester compounds 8 and N '-straight The reaction of alkane hydrazides 6 is similarly obtained compound shown in Formulas I.

7. according to the preparation method of claim 3 route 4, it is characterised in that with hydrazides 9 be raw material, by itself and straight chain fatty acid Or acyl chlorides or anhydride reaction obtain compound shown in Formulas I.

8. a kind of composition of medicine, it is characterised in that the acceptable carrier of compound and galenic pharmacy containing claim 1.

9. application of the compound of claim 1 in the medicine for preparing the prevention and treatment disease relevant with protein kinase.

10. application of the compound of claim 1 in the medicine for preparing the prevention and treatment disease relevant with EGFR-TK.

11. application of the compound of claim 1 in the medicine for preparing the prevention and treatment disease relevant with Raf kinases.

12. application according to claim 11, it is characterised in that the disease relevant with Raf kinases is tumour, immune mistake Tune, sacred disease.

13. application according to claim 12, it is characterised in that described tumor disease be liver cancer, kidney, lung cancer, cancer of pancreas, Stomach cancer, colorectal cancer, carcinoma of urinary bladder and breast cancer, oophoroma, squamous cell carcinoma, glioma, incidence cancer.