CN103910714A - Fluoro cyclobutyl imidazole compound - Google Patents

Fluoro cyclobutyl imidazole compound Download PDF

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Publication number
CN103910714A
CN103910714A CN201310006964.2A CN201310006964A CN103910714A CN 103910714 A CN103910714 A CN 103910714A CN 201310006964 A CN201310006964 A CN 201310006964A CN 103910714 A CN103910714 A CN 103910714A
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compound
formula
alkali
alkyl
preparation
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李晓强
李勇
陈永胜
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TIANJIN TERRABAY PHARMACEUTICALS TECHNOLOGY Co Ltd
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TIANJIN TERRABAY PHARMACEUTICALS TECHNOLOGY Co Ltd
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Priority to CN201310006964.2A priority Critical patent/CN103910714A/en
Priority to PCT/CN2013/089772 priority patent/WO2014108021A1/en
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero 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
    • A61K31/495—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
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D233/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings
    • C07D233/54—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings having two double bonds between ring members or between ring members and non-ring members
    • C07D233/64—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings having two double bonds between ring members or between ring members and non-ring members with substituted hydrocarbon radicals attached to ring carbon atoms, e.g. histidine

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  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The invention relates to a fluoro cyclobutyl imidazole compound shown in the formula (I) and a N-oxide, a pharmaceutically acceptable salt or a prodrug thereof, and relates to a preparation method thereof and intermediates useful in the preparation thereof; the invention also relates to a pharmaceutical composition containing the fluoro cyclobutyl imidazole compound and at least one pharmaceutically acceptable excipients and use of the pharmaceutical composition in the preparation of drugs for treating tyrosine kinase pathway disorder-related diseases, especially cancer. The formula (I) is shown in the specification, and R1, R2, R3, R4, R5, R6, R7, m and n are as defined in the specification.

Description

Fluoro tetramethylene base glyoxaline compound
Technical field
The present invention relates to formula (I) fluoro tetramethylene base glyoxaline compound and N-oxide compound, its pharmacologically acceptable salt or its prodrug, relate to its preparation method and in the described compound of preparation useful intermediate, the invention still further relates to comprise compound of the present invention and optionally at least one pharmaceutically acceptable vehicle pharmaceutical composition with and for the preparation for the treatment of and the tyrosine kinase pathway relevant disease-particularly purposes of the medicine of cancer of lacking of proper care.
Background technology
Protein kinase is the enzyme that a class regulates multiple important bioprocess, and it can be divided into receptor type and non-receptor type.It participates in controlling multiple important cell function for example signal transduction, Differentiation and proliferation.Except participating in the function of normal tissue/organ, many protein kinases also produce material impact to the human body of suffering from for example cancer of disease.The in the situation that of imbalance, carcinogenic protein kinases can cause that tumour forms and growth, and further encourages maintaining and developing of tumour.
Kinases inhibitor, as the common medicine of a class in molecular targeted property medicine, has good application prospect in the treatment of malignant tumour.The kinases inhibitor of early stage listing is mainly the specific inhibitor for single target spot, but is easy to produce resistance after target spot kinase mutant because its target spot is single.Broad spectrum kinase inhibitor is due to its simultaneously multiple kinase molecules of target and many signal paths, therefore not only can avoid the single target spot resistance causing of suddenling change, and can significantly expand its antitumor spectra.By the sudden change of tumor type on a large scale, the kinases of sudden change mark has been promoted to the research (Futreal for the broad spectrum kinase inhibitor of kinases group as drug target, P.A.et al.A census of human cancer genes.Nature Rev.Cancer.4,177-183(2004)).
Many Tyrosylprotein kinases (comprising receptor type and non-receptor type) are relevant to cancer (referring to Jianming Zhang, Priscilla L Yang and Nathanael S Gray.Targeting cancer with small molecular kinase inhibitors.Nature Review Cancer, 2009,9 (1): 28-39).Receptor tyrosine kinase (RTK) has extracellular part, membrane-spanning domain and intracellular portion, and the entirety of nonreceptor tyrosine kinase is positioned at cell.The sudden change of c-Kit Tyrosylprotein kinase declines relevant with the survival rate in the tumour of stomach and intestine interstitial.In acute myeloid granulocytic leukemia, Flt-3 sudden change is indicating that disease free survival declines.The expression of the VEGFR that blood vessel is played an important role is relevant with the lower survival rate of lung cancer.It is the important predictor that chronic myelocytic leukemia is replied that Bcr-Ab1 expresses, and Src Tyrosylprotein kinase is the telltale of prognosis mala in all stages of colorectal carcinoma.Therefore, in antitumor research, there is good application prospect for the broad spectrum kinase inhibitor of Tyrosylprotein kinase group.
Tyrosine kinase inhibitor can be used as the competitive inhibitor that Triphosaden (ATP) is combined with Tyrosylprotein kinase; Also can be used as the analogue of tyrosine, the activity of blocking-up Tyrosylprotein kinase, suppresses cell proliferation.At present the most frequently used micromolecular inhibitor for BCR-ABL Tyrosylprotein kinase comprises first-generation medicine imatinib clinically, s-generation medicine Dasatinib and nilotinib, and it is mainly by suppressing BCR-ABL fusion rotein, thus performance leukemia resisting action.
In January, 2012, U.S. FDA has been ratified the small molecule tyrosine kinase inhibitors Axitinib listing of Pfizer, how the small molecule tyrosine kinase inhibitors handkerchief that in December, 2012, U.S. FDA was ratified Ya Ruiyade company is for Buddhist nun's listing, this is indicating another new upsurge of taking turns antineoplastic target drug research, carries out medicament research and development become the focus of antitumor drug research in the world taking Tyrosylprotein kinase as target spot.
Summary of the invention
By the further R and D to tyrosine kinase inhibitor, the new fluoro tetramethylene base glyoxaline compound of contriver's discoverable type (I) can be used for treatment and the tyrosine kinase pathway relevant disease-particularly cancer of lacking of proper care.Compare with common three kinds of tyrosine kinase inhibitor imatinibs, nilotinib and Dasatinibs, the compounds of this invention has significantly more excellent pharmacology and property of medicine index, and has advantages of that target is clear and definite, toxicity is low, drug effect is high.
Therefore, in one aspect, the invention provides fluoro tetramethylene base glyoxaline compound and N-oxide compound, its pharmacologically acceptable salt or its prodrug of a kind of formula (I),
Wherein
R 1, R 2, R 4, R 6or R 7represent independently of one another H, halogen, C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many;
R 3and R 5represent independently of one another H, C 1-C 8alkyl, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) R awith-PhR asubstituting group replace one or many; And
R arepresent independently H or C 1-C 8alkyl;
M represents 0,1 or 2;
N represents 0,1,2,3 or 4.
In yet another aspect, the present invention also provides the method for the compound of preparation formula (I), comprising:
(A) under the existence of thinner and optional under the existence of catalyzer and alkali, make the compound of formula (II) and the compound generation linked reaction of formula (III),
Wherein
R 1, R 2, R 3, R 4, m and n as above define, and
X represents leavings group;
Wherein
R 5, R 6, R 7, m and n as above define.
Or
(B) under the existence of thinner and optional under the existence of alkali, make compound and Buddhist nun sieve's ethyl ester or Buddhist nun sieve's methyl esters generation condensation reaction of formula (IV),
Wherein
R 1, R 2, R 3, m and n as above define;
(Buddhist nun sieve's ethyl ester or methyl esters)
In yet another aspect, the present invention is also included in useful intermediate-formula (II) and compound of formula (IV) and preparation method thereof in the compound of preparation formula of the present invention (I) and the compound and preparation method thereof of useful intermediate-formula (VI) in the compound of preparation formula (IV).
In yet another aspect, the present invention also provides the compound of formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug for suppressing the purposes of receptor type or non-receptor type Tyrosylprotein kinase.
In yet another aspect, the present invention also provides the compound of use formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug for the preparation of the purposes of medicine that suppresses tyrosine kinase pathway.
In yet another aspect, the present invention also provides the purposes of compound and the medicine that N-oxide compound, its pharmacologically acceptable salt or its prodrug are bred for the preparation of inhibition tumor cell thereof of use formula (I).
In yet another aspect, the present invention also provides the compound of contained (I) and the pharmaceutical composition of N-oxide compound, its pharmacologically acceptable salt or its prodrug and at least one pharmaceutically acceptable vehicle thereof.
In yet another aspect, the present invention also provide the compound of use formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug and optionally at least one pharmaceutically acceptable vehicle in conjunction with the purposes of the medicine of the relevant disease of lacking of proper care for the preparation for the treatment of and tyrosine kinase pathway.
In yet another aspect, the present invention also provide the compound of use formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug and optionally at least one pharmaceutically acceptable vehicle in conjunction with the purposes of the medicine for the preparation of the treatment disease relevant to tumor cell proliferation.
In yet another aspect, the present invention also provide the compound of formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug and optionally at least one pharmaceutically acceptable vehicle in conjunction with the purposes of the medicine for the preparation for the treatment of cancer.
Brief description of the drawings
Fig. 1 shows TEB-415 and staurosporine is measured the IC50 of FLT3.
Fig. 2 shows TEB-415 and staurosporine is measured the IC50 of PDGFR α.
Fig. 3 shows TEB-415 and staurosporine is measured the IC50 of PDGFR β.
Fig. 4 shows TEB-415 and staurosporine is measured the IC50 of AKT1.
Fig. 5 shows TEB-415 and staurosporine is measured the IC50 of EGFR.
Fig. 6 shows TEB-415 and staurosporine is measured the IC50 of C-KIT.
Fig. 7 shows TEB-415 and staurosporine is measured the IC50 of JAK2.
Fig. 8 shows TEB-415 and staurosporine is measured the IC50 of ABL.
Fig. 9 show TEB-415 in L1210 cell IC50 measure.
Figure 10 show TEB-415 in Jurkat E6-1 cell IC50 measure.
Figure 11 show TEB-415 in HL-60 cell IC50 measure.
Figure 12 show TEB-415 in A549 cell IC50 measure.
Figure 13 show TEB-415 in K562 cell IC50 measure.
Figure 14 show TEB-415 in RPMI8226 cell IC50 measure.
Figure 15 show TEB-415 in THP-1 cell IC50 measure.
Figure 16 show TEB-415 in Ba/F3 cell IC50 measure.
Figure 17 shows animal pattern in the body weight change of accepting after TEB-415 and control drug treatment.
Figure 18 shows laboratory animal at administration 14 days (inoculating 22 days) the administration body weight change percentage of the 1st day relatively.
Figure 19 shows the growth curve of the K562 people source chronic leukemia transplantation model tumour after treatment.
Embodiment
Term as used herein " alkyl " and for example, alkyl group in combination definition (alkoxyl group) can be straight or branched, and it preferably has rudimentary carbon skeleton, for example, contain 1-8 carbon atom, preferred 1-6 carbon atom, more preferably 1-4 carbon atom.Described alkyl can be, for example methyl, ethyl, propyl group (as n-propyl or sec.-propyl), butyl (as normal-butyl, isobutyl-or the tertiary butyl), amyl group (as n-pentyl, isopentyl or neo-pentyl), hexyl are (as n-hexyl, isohexyl, 3-methyl amyl, 2,2-dimethylbutyl or 2,3-dimethylbutyl) and heptyl (as n-heptyl, 1-methyl hexyl or Isosorbide-5-Nitrae-dimethyl amyl group).
Term as used herein " alkoxyl group " refers to that alkyl is connected the group obtaining with Sauerstoffatom.It preferably has rudimentary carbon skeleton, for example, contain 1-8 carbon atom, preferred 1-6 carbon atom, more preferably 1-4 carbon atom.Described alkyl can be, for example methoxyl group, oxyethyl group, propoxy-or isopropoxy
Term as used herein " halogen " is fluorine, chlorine, bromine or iodine.The group being replaced by halogen represent by halogen, preferably by fluorine, chlorine and/or bromine, more preferably replaced or replace the group of (for example monosubstituted, two replace or three replace), for example CF by fluorine and/or cl part 3, CHF 2, CH 2f, CF 3cF 2, CH 2fCHCl, CCl 3, CHCl 2, CH 2cH 2cl, OCF 3, OCHF 2, OCH 2f, CF 3cF 2o, OCH 2cF 3and OCH 2cH 2cl etc.
Group of the present invention is not preferably substituted or mono-to trisubstituted, more preferably by monosubstituted to two replacements.
In the present invention, the key table of on-fixed position shows on the position that may connect arbitrarily of this key mapping in marked region.
In the present invention, the all possible steric isomer being limited by its concrete spatial form is contained in the definition of formula (I), as the mixture of enantiomer, diastereomer or cis-trans isomer and aforementioned stereoisomers, if suitable, the present invention had both comprised pure steric isomer, comprised again the mixture of its isomer.
For the various substituent possible combination of formula (I), must follow the rule of compound structure, formula (I) does not comprise the compound of running counter to natural law well known by persons skilled in the art.
The restriction of in the context of the invention, the group of formula (I) being made is also correspondingly applicable to other theme of the present invention, comprises intermediate, precursor, pharmacologically acceptable salt and the pharmaceutical composition of its preparation method, intermediate, intermediate.
The compound of formula of the present invention (I) and N-oxide compound thereof, its pharmacologically acceptable salt or its prodrug,
Wherein
R 1, R 2, R 4, R 6or R 7represent independently of one another H, halogen, C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many;
R 3and R 5represent independently of one another H, C 1-C 8alkyl, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many; And
R arepresent independently H or C 1-C 8alkyl;
M represents 0,1 or 2;
N represents 0,1,2,3 or 4.
Formula (I) above provides the broad definition to the compounds of this invention, as follows about preferred, preferred, most preferred definition or range specification:
The compound of formula of the present invention (I) and N-oxide compound thereof, its pharmacologically acceptable salt or its prodrug, wherein
R 1, R 2, R 4, R 6or R 7preferably represent independently of one another H, halogen, C 1-C 6alkyl, C 1-C 6alkoxyl group, wherein said C 1-C 6alkyl and C 1-C 6alkoxyl group is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-phR asubstituting group replace one or many;
R 3and R 5preferably represent independently of one another H or C 1-C 6alkyl, wherein said C 1-C 6alkyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-phR asubstituting group replace one or many; And
R apreferably represent independently H or C 1-C 6alkyl;
M preferably represents 0 or 1;
N preferably represents 0,1,2 or 3.
The compound of formula of the present invention (I) and N-oxide compound thereof, its pharmacologically acceptable salt or its prodrug, wherein
R 1, R 2, R 4, R 6or R 7more preferably represent independently of one another H, fluorine, chlorine, bromine and be selected from following C 1-C 4alkyl and C 1-C 4alkoxyl group, described C 1-C 4alkyl and C 1-C 4alkoxyl group is optionally replaced by one or more halogens that are selected from fluorine and chlorine: methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl, sec-butyl, methoxyl group, oxyethyl group, positive propoxy, isopropoxy, n-butoxy, tert.-butoxy, isobutoxy, CF 3, CHF 2, CH 2f, CF 3cF 2, CH 2fCHCl, CCl 3, CHCl 2, CH 2cH 2cl, OCF 3, OCHF 2, OCH 2f, CF 3cF 2o, OCH 2cF 3or OCH 2cH 2cl;
R 3and R 5more preferably represent independently of one another H and be selected from following C 1-C 4alkyl, described C 1-C 4alkyl is optionally replaced by one or more halogens that are selected from fluorine and chlorine: methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl, sec-butyl, CF 3, CHF 2, CH 2f, CF 3cF 2, CH 2fCHCl, CCl 3, CHCl 2, CH 2cH 2cl; And
M represents 0;
N more preferably represents 0,1 or 2.
In a most preferred embodiment, R 1for H, R 2for H, R 3for CF 3, R 4for H, R 5for methyl, R 6for H, R 7for H, m be 0 and n be 0 or 1, the compound of described formula (I) is: 4-methyl-3-{[4-(3-pyridyl)-2-pyrimidyl] amino }-N-[5-[4-(3,3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl]-3-[(trifluoromethyl) phenyl]-benzamide (being hereinafter also called TEB-415), its structure is suc as formula shown in (I-a):
In the context of the present invention, each substituting group wide in range, preferred, preferred, most preferred definition can be bonded to each other on demand.This expression the present invention includes the compound of described formula (I), wherein, for example, substituent R 1there is preferred definition and substituent R 2and R 3there is general definition, or, for example, substituent R 2there is preferred definition, substituent R 3have preferred definition, all the other substituting groups have wide in range definition.For the sake of simplicity, the not concrete statement of these various combinations out, but should be thought and comprises within the scope of the invention.In addition, the present invention is also correspondingly applicable to prepare its various intermediates for the definition of formula (I) compound, also comprise the intermediate of preparing its intermediate.
In the context of the present invention, the compound itself that term " compound of the present invention " expression (I) is contained, with and pharmacologically acceptable salt, its N-oxide compound or its prodrug.
Formula of the present invention (I) compound can, by following method preparation, comprise:
(A) under the existence of thinner and optional under the existence of catalyzer and alkali, make the compound of formula (II) and the compound generation linked reaction of formula (III),
Wherein
R 1, R 2, R 3, R 4, m and n as above define, and
X represents leavings group, preferably halogen and triflate (OTf);
Wherein
R 5, R 6, R 7, m and n as above define.
In aforesaid method, the mol ratio of the compound reaction of the compound of formula (II) and formula (III) is generally 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1.5.
In aforesaid method, preferred catalyzer is can catalysis aromatic halide or the metal catalyst of trifluoromethanesulfonic acid compound and amine generation linked reaction formation C-N key, and preferable alloy palladium catalyst, as Pd (dppf) Cl 2, Pd (PPh 3) 2cl 2, Pd (PPh 3) 4, Pd (dba) 2or Pd (OAc) 2.
In aforesaid method, suitable alkali is alkali (soil) metal carbonate, alkali (soil) metal hydroxides or alkali (soil) metal alkoxide, preferred alkali metal carbonate, more preferably K 2cO 3, Cs 2cO 3.
In aforesaid method, suitable thinner is all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example sherwood oil, normal hexane, Skellysolve A, benzene, toluene and dimethylbenzene; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Alcohols, for example methyl alcohol, ethanol, propyl alcohol, Virahol, butanols, isopropylcarbinol and the trimethyl carbinol; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Also has sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably use the mixed solvent of Isosorbide-5-Nitrae dioxane and the trimethyl carbinol.
Or
(B) under the existence of thinner and optional under the existence of alkali, make compound and Buddhist nun sieve's ethyl ester or Buddhist nun sieve's methyl esters generation condensation reaction of formula (IV),
Wherein
R 1, R 2, R 3, m and n as above define;
(Buddhist nun sieve's ethyl ester or methyl esters)
In aforesaid method, formula (IV) compound is generally 1:1 to 1:3 with the mol ratio that Buddhist nun sieve's ethyl ester or Buddhist nun sieve's methyl esters react, preferably 1:1 to 1:2, more preferably 1:1.1;
Described Buddhist nun sieve's ethyl ester or Buddhist nun sieve's methyl esters are commercially available available, and for example Buddhist nun sieve's ethyl ester very can medication chemistry company limited purchased from Nanjing, and lot number is 111223, and commodity are called Buddhist nun sieve's ethyl ester.
In aforesaid method, suitable alkali is alkali (soil) metal alkoxide, preferred as alkali alkoxide, as potassium isopropoxide, sodium isopropylate, propyl carbinol potassium, propyl carbinol sodium, sec-butyl alcohol potassium, sec-butyl alcohol sodium, sodium tert-butoxide, potassium tert.-butoxide, preferably potassium tert.-butoxide or sodium tert-butoxide.
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example, normal hexane, Skellysolve A, benzene, toluene and dimethylbenzene; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Also has sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably use methylene dichloride or tetrahydrofuran (THF).
In addition, intermediate-formula (II) of using in the compound of preparation formula (I) and the compound of formula (IV) with and preparation method thereof also in protection scope of the present invention,
The compound of formula (II),
Wherein
R 1, R 2, R 3, R 4, m and n as above define, and
X represents leavings group, preferably halogen and triflate (OTf), more preferably iodine.
In a most preferred embodiment, the structure of formula (II) is shown in following formula (II-a):
The compound of formula (IV):
Wherein
R 1, R 2, R 3, m and n as above define;
In a most preferred embodiment, the structure of formula (IV) is shown in following formula (IV-a):
The compound of described formula (II) can be prepared by the following method:
Under the existence of thinner and optional under the existence of alkali, make the compound generation condensation reaction of compound and the formula V of formula (IV),
Wherein
R 1, R 2, R 3, m and n as above define;
Wherein
R 4, n and X as above define, and
R brepresent halogen or hydroxyl.
In aforesaid method, the mol ratio of formula (IV) compound and the reaction of formula V compound is generally 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1;
In aforesaid method, the proton acceptor that suitable alkali is all routines.Preferred tertiary amine, for example Trimethylamine 99, triethylamine, Tributylamine, N, accelerine, N, N-dimethyl benzylamine, pyridine, N, N-diisopropylethylamine, DMAP (DMAP), N-methyl piperidine, N-methylmorpholine, N, N-dimethyl aminopyridine, diazabicyclo octane (DABCO), diazabicyclo-nonene (DBN) or diazabicyclo undecylene (DBU), preferably, DIPEA and DMAP mixture;
In aforesaid method, work as R bduring for hydroxyl, this reaction is preferably carried out under the existence of condensing agent, and described condensing agent is selected from: DCC, EDCI, CDI, DIC, HATU, HBTU;
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example, normal hexane, Skellysolve A, benzene, toluene and dimethylbenzene; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Also has sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably use methylene dichloride, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane or dimethyl formamide.
Described formula (IV) compound can be prepared by the following method:
Under the existence of thinner and optionally, under the existence of catalyzer and alkali, make formula (VI) compound and formula (VII) compound generation linked reaction,
Wherein
R 1as above define with m;
Wherein
R 2, R 3, n and X as above define;
In aforesaid method, the mol ratio of formula (VI) compound and the reaction of formula (VII) compound is generally 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1.5.
In aforesaid method, suitable catalyzer can be can catalysis aryl family's halogenide or trifluoromethanesulfonic acid compound and amido compound between the transition-metal catalyst of linked reaction, for example Cu (I) or Cu (II) salt, as CuI, Cu 2o or Cu (OAc) 2.。
In aforesaid method, suitable alkali can be mineral alkali or organic bases, and wherein mineral alkali is selected from alkali (soil) metal carbonate, alkali (soil) metal hydroxides or alkali (soil) metal alkoxide, preferred alkali metal carbonate, more preferably K 2cO 3, Cs 2cO 3.Organic bases is selected from Trimethylamine 99, triethylamine, Tributylamine, N, accelerine, N, N-dimethyl benzylamine, pyridine, N, N-diisopropylethylamine, DMAP (DMAP), N-methyl piperidine, N-methylmorpholine, N, N-dimethyl aminopyridine, diazabicyclo octane (DABCO), diazabicyclo-nonene (DBN) or diazabicyclo undecylene (DBU), preferably, triethylamine, pyridine;
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example sherwood oil, normal hexane, Skellysolve A, benzene,toluene,xylene or oil of mirbane; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Alcohols, for example methyl alcohol, ethanol, propyl alcohol, Virahol, butanols, isopropylcarbinol, the trimethyl carbinol and ethylene glycol; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably use dimethyl sulfoxide (DMSO), dimethyl formamide or Isosorbide-5-Nitrae dioxane.
In addition, the method for the compound of intermediate-formula (VI) of using in the compound of preparation formula (IV) and the compound of preparation formula (VI) also in protection scope of the present invention,
Wherein
R 1as above define with m;
In a most preferred embodiment, the structure of formula (VI) is shown in following formula (VI-a):
Described formula (VI) compound can be prepared by the following method:
Under the existence of thinner, make formula (VII) compound and carbonamidine or its reactant salt,
Wherein
R 1as above define with m, Y is the halogen that is selected from chlorine, bromine or iodine, preferably chlorine or bromine;
In aforesaid method, the salt of carbonamidine is selected from amitraz hydrochloride, carbonamidine acetate, carbonamidine formate, carbonamidine vitriol;
In aforesaid method, the compound of formula (VII) and the mol ratio of carbonamidine or its reactant salt are generally 1:1 to 1:10, preferably 1:3 to 1:6, more preferably 1:5.
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example sherwood oil, normal hexane, Skellysolve A, benzene,toluene,xylene or oil of mirbane; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Alcohols, for example methyl alcohol, ethanol, propyl alcohol, Virahol, butanols, isopropylcarbinol, the trimethyl carbinol and ethylene glycol; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably make spent glycol, dimethyl sulfoxide (DMSO) or Isosorbide-5-Nitrae dioxane.
The compound of described formula (VII) can be prepared by the following method:
(A) under the existence of thinner and under a kind of existence of alkali, the compound of formula (VIII) is reacted with halogenated acetic acids,
Wherein
R 1as above define with m, and R crepresent C 1-C 6alkyl, preferably C 1-C 3alkyl, more preferably methyl, ethyl, n-propyl, sec.-propyl;
In aforesaid method, the mol ratio of the compound of formula (VIII) and halogenated acetic acids reaction is generally 1:1 to 1:6, preferably 1:2 to 1:4, more preferably 1:3;
In aforesaid method, suitable alkali can be C 1-6lithium alkylide or two (C 1-6alkyl) amido lithium, preferably n-Butyl Lithium, tert-butyl lithium or lithium diisopropyl amido, more preferably lithium diisopropyl amido;
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example sherwood oil, normal hexane, Skellysolve A, benzene,toluene,xylene or oil of mirbane; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Also have sulfoxide type, for example dimethyl sulfoxide (DMSO).More preferably use ether, tetrahydrofuran (THF); Or
(B) under the existence of thinner and under a kind of existence of alkali, make compound and the diazomethane reaction of formula (IX),
Wherein
R 1as above define with Y;
In aforesaid method, the compound of formula (IX) and the mol ratio of diazomethane reaction are generally 1:3 to 1:10, preferably 1:3 to 1:8, more preferably 1:5;
In aforesaid method, suitable thinner can be all organic solvents that are inertia to participating in the material of this reaction.Preferably use hydro carbons, for example sherwood oil, normal hexane, Skellysolve A, benzene,toluene,xylene or oil of mirbane; Halohydrocarbon, for example methylene dichloride, trichloromethane, tetracol phenixin, chlorobenzene and orthodichlorobenzene; Ketone, for example acetone and methyl isopropyl Ketone; Ethers, for example ether, tetrahydrofuran (THF), Isosorbide-5-Nitrae dioxane, glycol dimethyl ether and diglyme; Ester class, for example ethyl acetate; Nitrile, for example acetonitrile; Alcohols, for example methyl alcohol, ethanol, propyl alcohol, Virahol, butanols, isopropylcarbinol, the trimethyl carbinol and ethylene glycol; Amides, for example dimethyl formamide; And sulfone class, for example tetramethylene sulfone; Also has sulfoxide type, the mixture of for example dimethyl sulfoxide (DMSO) and above-mentioned solvent.More preferably use ether, methylene dichloride.
In aforesaid method, the compound of formula (IX) is commercially available, maybe can carry out halogenating reaction by its corresponding acid by the ordinary method of this area and make.
The present invention also provides the compound of formula (I) and N-oxide compound or its pharmacologically acceptable salt thereof for suppressing the active purposes of receptor type tyrosine kinase or non-receptor type Tyrosylprotein kinase, be preferred for suppressing receptor type TYR kinase activity, more preferably for suppressing ABL kinase activity.
The present invention also provides the compound of formula (I) and N-oxide compound or its pharmacologically acceptable salt thereof for the preparation of the activity that suppresses receptor type tyrosine kinase or non-receptor type Tyrosylprotein kinase, preferably suppress receptor type TYR kinase activity, more preferably suppress the purposes of the medicine of ABL kinase activity.
In one embodiment, the compound of described formula (I), comprises that the mensuration that its N-oxide compound or its pharmacologically acceptable salt suppress the activity of receptor type tyrosine kinase is to be undertaken by measuring its IC50 that kinase activity is suppressed.
In one embodiment, described for the concentration of compound of described formula (I) that suppresses described tyrosine kinase activity from 10 μ M to 80 μ M, for example 20 μ M, 30 μ M, 40 μ M, 50 μ M or 60 μ M, and use DMSO as cosolvent, described DMSO concentration is 1% to 10%.In a preferred embodiment, described DMSO concentration is 2.5%.
In one embodiment, the compound of certain density formula (I) being contacted with ATP with described kinases and TK/STK3--vitamin H substrate is to contain 50mM Hepes/NaOH pH7.0,0.02%NaN 3, 0.01%BSA, 0.1mM ortho-vanadate (orthovanadate), 5mM MgCl 2, 1mM MnCl 2with in the solution of 1mM DTT, carry out, and described reaction is carried out in room temperature.
In one embodiment, described detection liquid is to contain Streptavidin--the mixing of XL665 and TK/STK3 antibody europium kryptofix 222 detects liquid; In some embodiments, described detection liquid is the Streptavidin of the solution dilution that contains 50mM Hepes/NaOH pH7.0,0.1%BSA, 0.8M KF and 20mM EDTA--the mixing of XL665 and TK/STK3 antibody europium kryptofix 222 (1:100/1:50) detects liquid, and described reaction is at room temperature reaction 1h.
In one embodiment, the detection of described reaction is with ENVISIO(Perkinelmer) (320nm excites instrument detection fluorescent signal; 665nm, 615nm transmitting) realize.Calculating the inhibiting rate in each hole by complete active hole and background signal hole, averages in multiple hole, with professional picture analysis software PRISM5.0, each testing compound is carried out to the matching that half suppresses active (IC50) simultaneously.
The present invention also provides the compound of formula (I) and N-oxide compound or its pharmacologically acceptable salt thereof the purposes for inhibition tumor cell propagation.
In one embodiment, the compound of described formula (I) is that TEB-415 and described tumor cell line are L1210, Jurkat E6-1, HL-60, A549, K562, RPMI8226, THP-1 and/or Ba/F3.
In one embodiment, the final concentration of described compound be 0 μ M to 100 μ M, for example 10 μ M, 20 μ M, 30 μ M, 40 μ M, 50 μ M, 60 μ M or 80 μ M.In one embodiment, the culture condition of described tumour cell is to hatch 72 hours in 37 DEG C, 100% relative humidity, 5%CO2 incubator.
In one embodiment, described detection is to use SpectraMax M5MicroplateReader to measure the absorbancy at 450nm wavelength place, and using 650nm place absorbancy as reference, calculate inhibiting rate, then adopt mapping analysis software GraphPad Prism5.0 matching IC50 curve and calculate that IC50 value carries out.
The present invention also provides the compound of formula (I) and N-oxide compound or its pharmacologically acceptable salt thereof to breed relevant disease-particularly purposes of the medicine of cancer for the preparation for the treatment of to inhibition tumor cell, and described cancer comprises leukemia, myelomatosis, stomach knurl and lung cancer.
In one embodiment, described detection method compound used is that TEB-415, test materials used are mouse (kinds: Mus Musculus; Strain: NODSCID) transplantation model.In one embodiment, described mouse transplantation model is the heteroplastic mouse of K562 people source chronic myelocytic leukemia.
In some embodiments, the concentration of TEB-415 used be 1mg/ml to 20mg/ml, for example 3mg/ml, 5mg/ml, 7.5mg/ml, 10mg/ml or 15mg/ml.
In some embodiments, using imatinib as positive control, using 10%NMP/90%PEG200 as solvent control.
In one embodiment, tumor cell inoculation, to mouse, is treated to tumor growth is to 400-1000mm 3for example 800mm 3described mouse is put to death; Select well-grown tumor tissues, under aseptic condition, be seeded to mouse; Treat that tumour grows to 100-200mm 3for example 160mm 3grouping administration.Use vernier callipers to measure gross tumor volume, measure major diameter and the minor axis of tumour, calculate tumor growth according to gross tumor volume, and calculate the inhibiting rate after administration.Application SPSS17.0 statistics software carries out One-Way ANOVA inspection, statistical analysis between gross tumor volume is organized.P<0.05 thinks significant difference.
The present invention also provide the compound of formula (I) and N-oxide compound thereof or its pharmacologically acceptable salt and optionally at least one pharmaceutically acceptable vehicle for the preparation for the treatment of the disease with tyrosine kinase pathway imbalance, the purposes of the medicine of the disease that special and ABL kinase pathways are lacked of proper care.
The present invention also provide the compound of formula (I) and N-oxide compound thereof or its pharmacologically acceptable salt and optionally at least one pharmaceutically acceptable vehicle for the preparation of the treatment disease relevant to tumor cell proliferation, the particularly purposes of the medicine of cancer, described cancer comprises leukemia, myelomatosis, stomach knurl and lung cancer.
The present invention also provides the compound of contained (I) and N-oxide compound thereof or its pharmacologically acceptable salt and the optional pharmaceutical composition of at least one pharmaceutically acceptable vehicle.
The optional employing of the wherein said pharmaceutically acceptable vehicle pharmaceutically acceptable vehicle adapting with pharmaceutical dosage form as known in the art, for example be recorded in Remington ' s Pharmaceutical Sciencesby E.W.Martin.See also Wang, Y.J.and Hanson, M.A., Journal of Parenteral Science and Technology, Technical Report No.10, Supp.42:2S, in 1988.
Pharmaceutical composition of the present invention can adopt the form of pharmaceutical preparation as mentioned below:
Pharmaceutical preparation of the present invention comprises applicable oral administration, administered parenterally (comprising subcutaneous, intracutaneous, intramuscular, vein) and rectal administration, but most suitable approach can depend on for example experimenter's illness and obstacle.
Described preparation can be presented easily by the form with unit dosage, also can be by the known either method preparation of pharmaceutical field technician.All methods all comprise the carrier-bound step with one or more ancillary components of composition by activeconstituents.In general, described preparation is prepared in the following way: make described activeconstituents and liquid vehicle or segmentation solid carrier or aforementioned both evenly and be nearly combined, the preparation of then as required moulding product one-tenth being wished.
Be applicable to for example each capsule, cachet agent or tablet that comprises activeconstituents described in predetermined amount of unit that the preparation of the present invention of oral administration can be rendered as separation; For pulvis or granule; For the solution in waterborne liquid or non-aqueous liquid or suspensoid; For oil-in-water liq emulsion or water-in-oil-type liquid emulsion.Described activeconstituents can also be rendered as bolus, electuary or paste.
Tablet can be by compression or moulding making, one of optional and one or more ancillary component.The tablet of compression can by compression in applicable machine can for example pulvis of free-flowing form or the activeconstituents of granule form prepare, optional and tackiness agent, lubricant, inert diluent, lubricant, tensio-active agent or dispersant.Moulding tablet can be prepared with the moistening powder compounds mixture of inert liquid diluent by moulding in applicable machine.Described tablet optionally can be by dressing or by indentation, or can be formulated slowly to discharge or controlled release described activeconstituents wherein.Compound of the present invention can be for example to be suitable for directly discharging or delaying the form administration discharging.Directly discharge or delay to discharge the applicable pharmaceutical composition that can contain the compounds of this invention by use and realize, or, particularly, in the situation that delaying to discharge, realize by the device that uses for example hypodermic implant or osmotic pump.Compound of the present invention can also pass through liposome administration.
Preferably, composition of the present invention is suitable for subcutaneous administration, for example, by injection subcutaneous administration.
The preparation of administered parenterally comprises water-based and nonaqueous aseptic injectable solution, and it can comprise antioxidant, buffer reagent, fungistat and can make described preparation and solute that target receptor's blood etc. oozes; And water-based and nonaqueous sterile suspension, it can comprise suspension agent and thickening material.In ampoule and phial that described preparation can for example seal at the container of single dose or multiple doses, and can preserve with freeze-drying (lyophilization) state, only need to before making with it, add for example salt solution of aseptic liquid vehicle or water for injection.Can be from the sterilized powder of mentioned kind, granule and tablet preparation injection solution and the suspension of preparation then and there.If needed, described pharmaceutical composition can also comprise for example sodium acetate or the sorbitol anhydride laurates such as a small amount of nontoxic such as wetting agent of auxiliary substance or emulsifying agent, sanitas, pH damping fluid.
Preparation for rectal administration can be rendered as enema,retention or the suppository containing for example theobroma oil, synthetic glyceride or polyoxyethylene glycol of common carrier.This carrier is generally solid at normal temperatures, but in rectal cavity, liquefies and/or dissolve to discharge described medicine.
Preferred unit dose formulations is the preparation of the described activeconstituents that contains significant quantity as described above or its suitable part.
Should be appreciated that, except the composition of above specifically mentioning, preparation of the present invention can also comprise other reagent conventional in this area relevant with the preparation type of just touching upon, and the preparation that is for example suitable for oral administration can comprise seasonings.
Following examples are provided, and to facilitate those skilled in the art to understand better the present invention, described embodiment, only for exemplary purpose, is not intended to limit the scope of the present disclosure.
Preparation method
The preparation of embodiment 1:2-chloro-1-(3,3-, bis-fluoro cyclobutyl) ethyl ketone
Under-20 ° of C, by N, N-lithium diisopropylamine (1.2M, 6.0eq.) slowly drops to Mono Chloro Acetic Acid (3.0eq.) in the solution of tetrahydrofuran (THF) and stirs half an hour.Then slowly drip 3,3-bis-fluoro cyclobutyl formate methyl esters (1.0eq.), dropwise the rear room temperature that is slowly warming up to, then drip dilute hydrochloric acid cancellation reaction, water dichloromethane extraction, anhydrous sodium sulfate drying, filters and obtains crude product, and it can be directly used in next step reaction without purifying further. 1H?NMR(CDCl 3,300MHz):δ4.12(s,2H),3.46-3.34(m,1H),2.90-2.71(m,4H)。
The preparation of embodiment 2:2-bromo-1-(3,3-, bis-fluoro cyclobutyl) ethyl ketone
3,3-, bis-fluoro cyclobutyl formates (13.6g, 10mmol) and thionyl chloride (50mL) mixture reflux are stirred 2 hours, after finishing, remove excessive thionyl chloride and obtain 3,3-, bis-fluoro cyclobutylmethyl acyl chlorides.By 3,3-bis-fluoro cyclobutylmethyl acyl chlorides (2.1g, 13.6mmol) be dissolved in methylene dichloride, Leng Que, Dao 20 ° of C of –, then drips diazomethane (136mL, 1.0M, 10.0eq.) solution, react after about half an hour, under-30 ° of C, drip saturated hydrobromic acid solution (20mL, 10.0eq).Reaction mixture is slowly warming up to stirring at room temperature 1 hour, then successively with saturated sodium bicarbonate and salt solution washing, organic phase dried over sodium sulfate, obtains 2-bromo-1-(3 except after desolventizing, 3-bis-fluoro cyclobutyl) ethyl ketone crude product 2.5g, can directly be used as next step reaction. 1H?NMR(CDCl 3,300MHz):δ3.92(s,2H),3.50-3.38(m,1H),2.93-2.71(m,4H).
The preparation of embodiment 3:4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles
The ethylene glycol solution of 2-bromo-1-(3,3-, bis-fluoro cyclobutyl) ethyl ketone or 2-chloro-1-(3,3-, bis-fluoro cyclobutyl) ethyl ketone and carbonamidine acetate (5.0eq.) is stirred and spent the night at 80-135 ° of C.After reaction finishes, reaction solution is cooling, then join in saturated aqueous common salt.By extracted with diethyl ether, organic phase is water successively, and then saturated common salt water washing adds anhydrous sodium sulfate drying.Filter, be spin-dried for solvent and obtain crude product, can obtain pure 4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles through silica gel column chromatography, yield is 15%-60%. 1H?NMR(CDCl 3,300MHz):δ8.34(br,1H),7.64(d,J=0.9Hz,1H),6.88-6.86(m,1H),3.43-3.31(m,1H),3.01-2.65(m,4H);MS-ESI(M+H+):159.0;HR-MS(ESI):cald.for?C 7H 9F 2N 2 +(M+H +):159.0728,found159.0732.
The preparation of embodiment 4:3-(4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-5-trifluoromethylaniline
Under protection of inert gas; by 4-(3; 3-bis-fluoro cyclobutyl)-1H-imidazoles (380mg; 2.4mmol); the bromo-5-5-trifluoromethylaniline of 3-(480mg; 2.0mmol); salt of wormwood (350mg; 2.5mmol); cuprous iodide (57mg, 0.15eq.), oxine (44mg; 0.15eq.) and anhydrous dimethyl sulphoxide (2mL) add in pressure bottle, then heating (about 115-120 ° C) stirring reaction 48 hours.After reaction finishes, reaction solution is cooled to 40-50 ° of C, adds the saturated ammoniacal liquor of 15mL to stir after 2 hours and continue to be cooled to room temperature, use extracted with diethyl ether.Organic phase water, saturated common salt water washing, anhydrous sodium sulfate drying, filter, except desolventizing obtains crude product, obtain pure compound 3-(4-(3 through silica gel column chromatography, 3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-5-trifluoromethylaniline 400mg, yield is 63%. 1H?NMR(CDCl 3,300MHz):δ7.79(d,J=0.9Hz,1H),7.07(s,1H),6.94(s,1H),6.86(s,1H),6.79(dd,J=0.9,2.1Hz,1H),4.10(br,2H),3.43-3.31(m,1H),3.02-2.73(m,4H);MS-ESI(M+H +):318.1;HR-MS(ESI):cald.For?C 14H 13F 5N 3+(M+H+):318.1024,found318.1023.
The preparation of embodiment 5:N-(3-(4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-trifluoromethyl) iodo-4 methyl benzamides of-3-
By iodo-3-4-methyl benzoyl chloride (308mg, 1.1eq.), 3-(4-(3,3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-5-trifluoromethylaniline (317mg, 1.0mmol), N, N-diisopropylethylamine (0.25mL, 1.5eq.) and DMAP (30mg, 0.25eq.) be dissolved in organic solvent, stirring at room temperature reaction (8 hours).After reaction finishes, cancellation adds water, by extracted with diethyl ether, dry, filter, obtain crude product after removing organic solvent, obtain pure compound N-(3-(4-(3 through silica gel column chromatography, 3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-trifluoromethyl) the iodo-4 methyl benzamide 504mg of-3-, yield is 90%. 1H?NMR(DMSO-d 6,300MHz):δ8.41(d,J=1.8Hz,1H),8.27(d,J=1.0Hz,1H),8.25(t,J=1.8Hz,1H),8.12(s,1H),7.91(dd,J=1.8,7.9Hz,1H),7.74(s,1H),7.69(s,1H),7.49(d,J=8.2Hz,1H),3.37-3.26(m,1H),2.97-2.65(m,4H),2.42(s,3H);MS-ESI(M+H +):562.0;HR-MS(ESI):cald.for?C 22H 18F 5IN 3O+(M+H +):562.0409,found562.0416.
Embodiment 6:4-methyl-5-{[4-(3-pyridyl)-2-pyrimidyl] amino }-N-{3-[4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl]-3-[(trifluoromethyl) phenyl] preparation (according to the preparation method A in specification sheets) of-benzamide
Under protection of inert gas; by N-(3-(4-(3; 3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-trifluoromethyl) the iodo-4-methyl benzamide of-3-(247mg; 0.44mmol; 1.1eq.); 4-(3-pyridyl)-2-amine pyrimidine (69mg; 1.0eq.); Pd (0) catalyzer (10-15%); cesium carbonate (182mg, 1.4eq.), 1; the mixture of 4-dioxane (1.6mL) and the trimethyl carbinol (0.8mL) adds in pressure bottle, and gained mixture was 110-115 ° of C stirring reaction 10 hours.After reaction finishes, reaction solution is cooled to room temperature, add methylene dichloride dilution, add water and wash away inorganic salt, organic phase dried over sodium sulfate, filters, except desolventizing obtains crude product, obtain pure compound 4-methyl-N-[3-(4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-trifluoromethyl through column chromatography] 3-[[4-(3-pyridyl) 2-pyrimidyl] aminobenzamide 170mg, yield is 70%. 1H?NMR(DMSO-d 6,300MHz):δ9.25(d,J=1.2Hz,1H),8.64(dd,J=1.0,4.6Hz,1H),8.52(d,J=5.2Hz,1H),8.42(dt,J=8.0,1.8Hz,1H),8.32(d,J=1.6Hz,1H),8.28(t,J=1.8Hz,1H),8.27(d,J=0.9Hz,1H),8.14(s,1H),7.74-7.71(m,2H),7.68(s,1H),7.50-7.41(m,3H),3.38-3.26(m,1H),2.97-2.65(m,4H),2.33(s,3H);MS-ESI(M+H +):606.2;HR-MS(ESI):cald.for?C 31H 25F 5N 7O+(M+H +):606.2035,found606.2033.
Embodiment 7:4-methyl-5-{[4-(3-pyridyl)-2-pyrimidyl] amino }-N-{3-[4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl]-3-[(trifluoromethyl) phenyl] preparation (according to the preparation method B in specification sheets) of-benzamide
Under protection of inert gas, to 3-(4-(3, 3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-5-trifluoromethylaniline (1.58g, 5.0mmol) with 4-methyl-3-[[4-(3-pyridyl)-2-pyrimidyl] amino] ethyl benzoate (1.84g, 5.5mmol, 1.1eq.) anhydrous THF suspension liquid in slowly drip the THF solution of 15% potassium tert.-butoxide (3.0eq.), maintain interior temperature 0-10 ° C, dropwise rear 45 ° of C reactions 3 hours that are slowly warming up to, after reaction finishes, cooling, water cancellation reaction, extraction, dry, filter, except desolventizing obtains crude product, obtain TEB-415 sterling 1.8g through column chromatography purification, yield 60%.Qualification is as embodiment 6.
Embodiment 8:4-methyl-5-{[4-(3-pyridyl)-2-pyrimidyl] amino }-N-{3-[4-(3,3-, bis-fluoro cyclobutyl)-1H-imidazoles-1-yl]-3-[(trifluoromethyl) phenyl] preparation (according to the preparation method B in specification sheets) of-benzamide
Under protection of inert gas, to 3-(4-(3, 3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl)-5-5-trifluoromethylaniline (1.58g, 5.0mmol) with 4-methyl-3-[[4-(3-pyridyl)-2-pyrimidyl] amino] methyl benzoate (1.76g, 5.5mmol, 1.1eq.) anhydrous THF suspension liquid in slowly drip the THF solution of 15% potassium tert.-butoxide (3.0eq.), maintain interior temperature 0-10 ° C, dropwise rear 45 ° of C reactions 3 hours that are slowly warming up to, after reaction finishes, cooling, water cancellation reaction, extraction, dry, filter, except desolventizing obtains crude product, obtain TEB-415 sterling 1.8g through column chromatography purification, yield 60%.Qualification is as embodiment 6.
Bioassay
Embodiment 1: compound TEB-415 suppresses IC50 to the activity of kinases Flt3, PDGFR alpha, PDGFR beta, AKT1, EGFR, Kit, JAK2 or Abl and measures
Test objective
Test compounds TEB-415 suppresses active IC50 value to the half of kinases Flt3, PDGFR alpha, PDGFR beta, AKT1, EGFR, KIT, JAK2 or Abl.
Material and instrument
2104 multilabel Reader (PerkinElmer); OptiPlate-384, White Opaque384-well MicroPlate (Cat.6007290, PerkinElmer); HTRF kinEASE TK (Cat.62TKOPEC, Cisbio); HTRF kinEASE STK3 (Cat.62ST3PEB, Cisbio); Abl enzyme (Cat:14-529, Millipore); PDGFR α (Cat:PV3811, Invitrogen); EGFR (Cat:PV3872, Invitrogen); Kit enzyme (Cat:BML-SE447-0010, Biomol); JAK2 enzyme (Cat:BML-SE435-0010, Biomol); PDGFR β (Cat:P3082, Invitrogen); AKT1 (Cat:P2999, Invitrogen); FLT3 (Cat:14-500, Millipore); 10mM ATP (Cat.PV3227, Invitrogen); 1M DTT (Cat.D5545, Sigma); 1M MgCl 2(Cat.M8266, Sigma); 1M MnCl 2(Cat.244589, Sigma); Testing compound TEB-415.
Testing sequence
1) reagent preparation
Table 1 illustrates the preparation of Flt3, PDGFR α, PDGFR β, AKT1, EGFR, Kit, JAK2 or eight kinds of kinases reagent of Abl.
Eight kinds of each components of kinase whose reaction system of table 1. and concentration table
1 × FLT3 enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M MnCl 2, 1 μ L1M DTT, 793 μ L ddH 2o;
1 × PDGFR β enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 20 μ L SEB, 1 μ L1M MnCl 2, 773 μ L ddH 2o;
1 × Abl enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl2,1 μ L1M DTT, 20 μ L SEB, 774 μ L ddH in 1mL1 × kinase buffer liquid 2o;
1 × EGFR enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 1 μ L1M MnCl 2, 793 μ L ddH 2o;
1 × JAK2 enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 794 μ L ddH 2o;
1 × AKT1 enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 794 μ L ddH 2o;
1 × Kit enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 10 μ L SEB, 784 μ L ddH 2o;
1 × PDGFR α enzyme buffer liquid: contain 200 μ L5 × enzyme buffer liquid, 5 μ L1M MgCl in 1mL1 × kinase buffer liquid 2, 1 μ L1M DTT, 8 μ L SEB, 1 μ L1M MnCl 2, 785 μ L ddH 2o;
The concrete concentration of 5 × substrate-TK/STK3 and ATP working fluid: substrate-TK/STK3 and ATP is in table 1.With 1 × kinase buffer liquid dilution substrate-TK/STK3 and ATP to reaction density 5 times;
5 × enzyme working fluid: the concentration optimization of eight kinds of kinases (Flt3, PDGFR alpha, PDGFR beta, AKT1, EGFR, KIT, JAK2 or Abl) completes in work before, and screening concentration is in table 1.With 1 × kinase buffer liquid preparation, 5 × enzyme working fluid;
The concentration of 4 × Sa-XL665 working fluid: Sa-XL665 in reaction is referring to table 1.With detecting damping fluid preparation 4 × Sa-XL665 working fluid;
4 × TK/STK3-Ab-cryptate working fluid: TK-Ab-Cryptate is diluted to 100 times as working fluid with detecting damping fluid; With detecting damping fluid, STK3-Ab-Cryptate is diluted to 50 times as working fluid.
2) experiment process
After all reagent prepares according to the method described above, outside dezymotizing, after equilibrating to room temperature, start to carry out application of sample, referring to table 2.
TK/STK3--vitamin H substrate, ATP, enzyme and certain density compound are containing 50mM Hepes/NaOH pH7.0,0.02%NaN3,0.01%BSA, 0.1mM ortho-vanadate (orthovanadate), 5mM MgCl 2, 1mM MnCl 2with room temperature reaction in the solution of 1mM DTT.The inhibition concentration of testing compound TEB-415 is since 50 μ M, and 5 times of gradient dilutions use 2.5% DMSO as cosolvent.To the Streptavidin that adds the solution dilution of 10 μ l through containing 50mM Hepes/NaOH pH7.0,0.1%BSA, 0.8M KF and 20mM EDTA in all reacting holes--XL665 and TK/STK3 antibody europium kryptofix 222 (1:100/1:50) mix and detect liquid, after room temperature reaction 1h, (320nm excites to detect fluorescent signal with ENVISION (Perkinelmer) instrument; 665nm, 615nm transmitting).Calculating the inhibiting rate in each hole by complete active hole and background signal hole, averages in multiple hole, with professional picture analysis software PRISM5.0, each testing compound is carried out to the matching that half suppresses active (IC50) simultaneously.
It is as follows that table 2. is tested application of sample schema:
3) data analysis
Transmit/615nm of transmitting ratio (ER)=665nm transmits;
Inhibiting rate=(ER positiveeR negative control-ER sampleeR sample)/(ER positiveeR positive right according to-ER negativeeR negative control) * 100%;
Test-results
Application HTRF kinEASE TK/STK3kit detection compound TEB-415 verifies mensuration to the half-inhibition concentration (IC50) of Flt3, PDGFR α, PDGFR β, AKT1, EGFR, KIT, JAK2 and Abl enzyme respectively, compound TEB-415 is the doubling dilution with 5 times from 50 μ M respectively, controlling DMSO concentration is 1%, measure concentration for 10, Simultaneous Determination positive compound staurosporine (Staurosporine), each concentration is multiple hole.Test-results is referring to Fig. 1-Fig. 8
Interpretation of result
The IC50 value test result that table 3. kinase activity suppresses gathers
Can find out that by upper table TEB-415 has fabulous selectivity to ABL kinases, other similar kinases is not possessed to inhibition.Thereby make TEB-415 there is less toxicity.
Embodiment 2: the IC50 of compound TEB-415 extracorporeal anti-tumor cell proliferation measures
Test objective
Application CCK-8 detection kit detection compound TEB-415 suppresses IC50 value to the In Vitro Anti propagation of 8 tumor cell lines (L1210, Jurkat E6-1, HL-60, A549, K562, RPMI8226, THP-1 and Ba/F3).
Materials and methods
Cell strain:
L1210 mouse lymph leukemia cell line (purchased from Shanghai cell biological institute of the Chinese Academy of Sciences); Jurkat E6-1 human T lymphocyte leukemia cell line (purchased from Shanghai cell biological institute of the Chinese Academy of Sciences); A549 Non-small cell lung carcinoma cell strain (purchased from Shanghai cell biological institute of the Chinese Academy of Sciences); HL-60 people's Acute Myeloid Leukemia Cells Contributing strain (purchased from Shanghai cell biological institute of the Chinese Academy of Sciences); K562 people's chronic myelocytic leukemia cell strain (purchased from U.S. ATCC); RPMI8226 Multiple myeloma cell lines (purchased from U.S. ATCC); THP-1 people's monokaryon leukemia cell line (purchased from U.S. ATCC); Ba/F3 mouse pro-B lymphocyte strain (purchased from U.S. ATCC).
Reagent and consumptive material:
Cell Counting Kit-8(Cat#CK04-13, Dojindo); 96 well culture plates (Cat#3599, Corning Costar); Foetal calf serum (Cat#10099-141, GIBCO); Substratum (Invitrogen); Desk-top microplate reader SpectraMax M5Microplate Reader(Molecular Devices); Compound TEB-415 (MW605.56,10mg).
Testing sequence
1) reagent preparation
The preparation of table 4. substratum
Cell strain Substratum
L1210 DMEM+10％FBS
JurkatE6-1 RPMI1610+10％FBS
HL-60 RPMI1640+10％FBS+1％L-Glutamine
A549 F12K+10％FBS
K562 IMDM+10％FBS
RPMI8226 RPMI1640+10％FBS
THP-1 RPMI1640+10％FBS+0.05mM2-mercaptoethanol
Ba/F3 RPMI1640+10％FBS+10ng/mL?murine?IL-3
2) preparation of compound: making final concentration by DMSO diluted compounds is 10mM.
3) cell cultures
A) collect logarithmic phase cell, counting, with perfect medium Eddy diffusion cell;
B) adjust cell concn to suitable concn, inoculation 96 orifice plates, 100 μ l cell suspensions are inoculated in every hole;
C) cell is at 37 DEG C, 100% relative humidity, 5%CO 2in incubator, hatch 24 hours.
4) IC50 test
A) collect logarithmic phase cell, counting, with perfect medium Eddy diffusion cell, adjusts cell concn to suitable concn (determining according to cell density optimization Test result), inoculation 96 orifice plates, and every hole adds 100 μ l cell suspensions.Cell is at 37 DEG C, 100% relative humidity, 5%CO 2in incubator, hatch 24 hours;
B) with substratum, testing compound is diluted to after 500 μ M to gradient dilution 8 times.Add cell by 25 μ l/ holes.Compound final concentration is from 100 μ M to 0 μ M, 4 times of gradient dilutions, totally 10 concentration point;
C) cell is placed in 37 DEG C, 100% relative humidity, 5%CO 2in incubator, hatch 72 hours;
D) inhale and abandon substratum (except suspension cell), add perfect medium (for suspension cell strain, directly adding 10 μ l CCK-8) containing 10%CCK-8 to be placed in 37 DEG C of incubators and hatch 2-4 hour;
E) after light shaking, on SpectraMax M5Microplate Reader, measure the absorbancy at 450nm wavelength place, using 650nm place absorbancy as reference, calculate inhibiting rate.
5) data processing
Be calculated as follows the inhibiting rate of medicine to growth of tumour cell:
Growth of tumour cell inhibiting rate %=[(A c-A s)/(A c-A b)] × 100%
A s: the OA (cell+CCK-8+ testing compound) of sample; A c: the OA (cell+CCK-8+DMSO) of negative control; A b: the OA (substratum+CCK-8+DMSO) of positive control.
Adopt professional mapping analysis software GraphPad Prism5.0 matching IC50 curve and calculate IC50 value.
Test-results
This test has detected the In Vitro Anti proliferation function of compound TEB-415 to 8 tumor cell lines (L1210, Jurkat E6-1, HL-60, A549, K562, RPMI8226, THP-1 and Ba/F3).Compound effects final concentration is from 100 μ M to 0 μ M, 4 times of gradient dilutions, totally 10 points. in test process, after observing one of them compound TEB-415 (100 μ M and 25 μ M) being added to cell culture medium under higher activity, occur muddy and separate out phenomenon, having caused the data of these two activity points can not reflect exactly the restraining effect of this compound on cell proliferation.Concrete test-results is referring to Fig. 9-Figure 16 and table 5.
The IC50 of table 5. extracorporeal anti-tumor cell proliferation measures and gathers
Embodiment 3: the pharmacodynamic study of compound TEB-415 treatment K562 people source chronic myelocytic leukemia heteroplastic transplantation model
Supply reagent product compound method (each matching while using):
(1) 3mg/ml, 7.5mg/ml, 15mg/ml TEB-415: take respectively 9 milligrams, 22.5 milligrams, 45 milligrams TEB-415 powder and be dissolved in 0.3 milliliter of NMP, after it dissolves completely, add 2.7 milliliters of PEG200 to mix;
(2) 7.5mg/ml imatinib: take 22.5 milligrams of imatinib powder and be dissolved in 0.15 milliliter of DMSO, after it dissolves completely, add the physiological saline (0.9% sodium chloride injection) (Kelun Pharm Ind Co., Ltd., Sichuan, lot number: M090714) of 2.85 milliliters to mix.
(3) solvent control adopts 10%NMP/90%PEG200(Tianjin recovery fine chemistry industry institute).
Experimental animal and feeding and management
Test mouse used (kind: Mus Musculus; Strain: NODSCID), age in 6-8 week, male, body weight 20-26 gram, 75, provide (production licence number: SCXK(capital) 2009-0004 by Beijing HFK Bio-Technology Co., Ltd.; The certification of fitness number: 0245145).
Experimental animal is raised in the laminar flow clean room of the SPF level fixed temperature and humidity of Pharmaron's animal center (AAALAC authenticated unit), uses independent ventilation cage tool IVC, temperature for (22 ± 3) DEG C, humidity be 40-80%.Feed is a clean level mouse material, purchased from the feed corporation,Ltd that pulls together of Beijing Austria of section; Tap water is super-filtration purifying water, and food and water all pass through autoclaving processing.Animal can freely absorb high pressure sterile food and drinking-water.Be number of animals by the punching of ear edge.
Every 5 mouse one cages, cage has polycarbonate to be made, and volume is 300mm × 180mm × 150mm.Soft corn cob autoclave sterilization processed cleans bedding and padding, changes weekly twice.Every cage tool has label, indicates size of animal, sex, strain, time of reception, group and time on-test.
Cell cultures
With 10% foetal calf serum that contains deactivation, the IMDM substratum of 100U/ml penicillin and 100 μ g/ml Streptomycin sulphates and 2mM glutamine is at 37 DEG C, 5%CO 2incubator in cultivate K562 cell.Cell cultures initial concentration is 5 × 10 5individual/milliliter, every 3 to 4 days after cell covers with sub-bottle go down to posterity.Inoculation by the tumour cell in logarithmic phase for in-vivo tumour.
Tumor cell inoculation and grouping
The K562 tumour cell 1.0 × 10 that PBS is resuspended 7it is subcutaneous that/0.1ml is inoculated in SCID right side of mice fore flank flank, inoculates altogether 5.Treat that tumor growth is to 800mm 3time put to death animal, under aseptic condition, peel off tumour, select well-grown tumor tissues, tumor tissues is cut into 2 × 2 × 2mm with knife blade 3knurl piece, is seeded to SCID right side of mice side of body flank subcutaneous, and corotation connects 75 animals.Treat that tumour grows to 160mm 3the administration of dividing into groups when the left and right, totally 5 groups, 10 every group.
The measurement of tumour and test index
Use weekly vernier callipers to carry out twice measurement to gross tumor volume, measure major diameter and the minor axis of tumour, its volume calculation formula is: volume=0.536 × major diameter × minor axis 2.Calculate tumor growth according to gross tumor volume
Inhibiting rate (TGI)=(1-T/C) × 100%, wherein T is the mean value of each tested material group relative tumour volume, C is the mean value of solvent control group relative tumour volume.
When treatment finishes, by the peaceful and comfortable execution of mice with tumor of control group and imatinib group, all the other TEB-415 treatment groups continue to observe to 100 days.
Table 6 illustrates the pharmacodynamic study dosage regimen of application compound TEB-415 treatment K562 people source chronic myelocytic leukemia heteroplastic transplantation model.
The pharmacodynamic study dosage regimen of table 6.TEB-415 treatment K562 people source chronic myelocytic leukemia heteroplastic transplantation model
Group Number of cases Treatment Dosage * (mg/kg) Route of administration The course for the treatment of
1 10 Solvent control -- p.o qd×14days
2 10 TEB-415 30 p.o qd×14days
3 10 TEB-415 75 p.o qd×14days
4 10 TEB-415 150 p.o qd×14days
5 10 Imatinib 75 p.o qd×14days
Note: administration volume is according to experimental animal body weight by 10 μ l/g, and while there is weight loss 15-20%, drug withdrawal is to row administration again after weight recovery; * drug dose shows the amount of medicine sterling.
Statistical analysis
Application SPSS17.0 statistics software carries out One-Way ANOVA inspection, statistical analysis between gross tumor volume is organized.P<0.05 thinks significant difference.
Test-results
During treatment, there is small size fluctuation in the experimental animal body weight of each treatment group.The experimental animal body weight that continues observation TEB-415 treatment group after drug withdrawal all returns to initial level, and body weight change is shown in Fig. 1,2.
Tumor growth suppresses result referring to table 7 and Figure 17,18,19.
The tumor-inhibiting action of table 7.TEB-415 to K562 people source chronic myelocytic leukemia heteroplastic transplantation model
Note: a.mean ± standard error: b.observe to 100 days; c.with control group comparison.
P value: group 2vs. group 3=0.010; Group 2vs. group 4=0.010; Group 2vs. organizes 5 < 0.001;
Group 3vs. organizes 5 < 0.001; Group 4vs. organizes 5 < 0.001.
In this test, by test compounds TEB-415 three dosage levels to the body of K562 people source chronic myelocytic leukemia heteroplastic transplantation model tumor growth in restraining effect comparing with the result for the treatment of of positive control medicine imatinib.
The inoculation of K562 knurl piece is after 22 days, and the gross tumor volume average of solvent control group is 3014mm 3.Imatinib 75mg/kg group gross tumor volume average is 2096mm 3(TGI=30.1%, P=0.103).TEB-41530mg/kg group gross tumor volume average is 70mm 3(TGI=97.7%, with relatively P<0.001 of control group), administration finishes 4 days, and (inoculating 26 days) has the tumor regressions of 4 mouse, has no afterwards recurrence.The tumour of TEB-41575mg/kg and 150mg/kg treatment group all disappears, and continues to observe 100 days without tumor recurrence phenomenon.
TEB-415 has extremely strong antitumous effect to K562 tumor model.The tumour of middle high dosage (75mg/kg and 150mg/kg) group disappears completely, observes and does not have recurrence sign in 100 days, means that disease reaches healing.Low dosage TEB-415(30mg/kg) tumor control rate for the treatment of group is also significantly higher than imatinib 75mg/kg group (P<0.001).During treatment, respectively organize experimental animal to test-compound well-tolerated, the security of medicine is fine.
Describing the present invention with specific preferred embodiment is herein for making reader implement time of the present invention there is no inappropriate operation, should not be construed the scope of the present invention that limits.It will be understood by those skilled in the art that the present invention is also applicable to variation and the adjustment outside specific description, and the present invention includes all this variation and adjustment.
Contriver requires to fall into amendment in claim scope and spirit below and the right of change.

Claims (13)

1. the fluoro tetramethylene base glyoxaline compound of formula (I) and N-oxide compound, its pharmacologically acceptable salt or its prodrug,
Wherein
R 1, R 2, R 4, R 6or R 7represent independently of one another H, halogen, C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 1-C 8alkoxyl group, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many;
R 3and R 5represent independently of one another H, C 1-C 8alkyl, C 2-C 8thiazolinyl or C 2-C 8alkynyl, wherein said C 1-C 8alkyl, C 2-C 8thiazolinyl and C 2-C 8alkynyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many; And
R arepresent independently H or C 1-C 8alkyl;
M represents 0,1 or 2;
N represents 0,1,2,3 or 4;
Preferably, R 1, R 2, R 4, R 6or R 7represent independently of one another H, halogen, C 1-C 6alkyl, C 1-C 6alkoxyl group, wherein said C 1-C 6alkyl and C 1-C 6alkoxyl group is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many;
R 3and R 5represent independently of one another H or C 1-C 6alkyl, wherein said C 1-C 6alkyl is optionally selected from halogen ,-CN ,-OR independently a,-C (O) R a,-C (O) OR a,-OC (O) Ra and-PhR asubstituting group replace one or many; And
R arepresent independently H or C 1-C 6alkyl;
M represents 0 or 1;
N represents 0,1,2 or 3;
More preferably, R 1, R 2, R 4, R 6or R 7represent independently of one another H, fluorine, chlorine, bromine and be selected from following C 1-C 4alkyl and C 1-C 4alkoxyl group, described C 1-C 4alkyl and C 1-C 4alkoxyl group is optionally replaced by one or more halogens that are selected from fluorine and chlorine: methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl, sec-butyl, methoxyl group, oxyethyl group, positive propoxy, isopropoxy, n-butoxy, tert.-butoxy, isobutoxy, CF 3, CHF 2, CH 2f, CF 3cF 2, CH 2fCHCl, CCl 3, CHCl 2, CH 2cH 2cl, OCF 3, OCHF 2, OCH 2f, CF 3cF 2o, OCH 2cF 3or OCH 2cH 2cl;
R 3and R 5represent independently of one another H and be selected from following C 1-C 4alkyl and C 1-C 4alkoxyl group, described C 1-C 4alkyl and C 1-C 4alkoxyl group is optionally replaced by one or more halogens that are selected from fluorine and chlorine: methyl, ethyl, n-propyl, sec.-propyl, normal-butyl, isobutyl-, the tertiary butyl, sec-butyl, CF 3, CHF 2, CH 2f, CF 3cF 2, CH 2fCHCl, CCl 3, CHCl 2, CH 2cH 2cl; And
M represents 0;
N represents 0 or 1;
Most preferably, R 1for H, R 2for H, R 3for CF 3, R 4for H, R 5for methyl, R 6for H, R 7for H, m be 0 and n be 0 or 1, the compound of described formula (I) is 4-methyl-3-{[4-(3-pyridyl)-2-pyrimidyl] amino }-N-[5-[4-(3,3-bis-fluoro cyclobutyl)-1H-imidazoles-1-yl]-3-[(trifluoromethyl) phenyl]-benzamide, its structure is suc as formula shown in (I-a):
2. the method for the compound of the formula (I) of preparation claim 1, comprising:
(A) under the existence of thinner and optional under the existence of catalyzer and alkali, make the compound of formula (II) and the compound generation linked reaction of formula (III),
Wherein
R 1, R 2, R 3, R 4, m and n be as definition in claim 1, and
X represents to be selected from the leavings group of halogen and triflate, preferably iodine;
Wherein
R 5, R 6, R 7, m and n be as definition in claim 1;
Or
(B) under the existence of thinner and optional under the existence of alkali, make compound and Buddhist nun sieve's ethyl ester or Buddhist nun sieve's methyl esters generation condensation reaction of formula (IV),
Wherein
R 1, R 2, R 3, m and n be as definition in claim 1,
(Buddhist nun sieve's ethyl ester or methyl esters).
3. according to the preparation method of claim 2, wherein, the mol ratio of the compound reaction of the compound of formula (II) and formula (III) is generally 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1.5; Catalyzer is can catalysis aromatic halide or the metal catalyst of trifluoromethanesulfonic acid compound and amine generation metal linked reaction, and preferable alloy palladium catalyst, as Pd (dppf) Cl 2, Pd (PPh 3) 2cl 2, Pd (PPh 3) 4, Pd (dba) 2or Pd (OAc) 2; Alkali is alkali (soil) metal carbonate, alkali (soil) metal hydroxides or alkali (soil) metal alkoxide, preferred alkali metal carbonate, more preferably K 2cO 3, Cs 2cO 3; The mol ratio that the compound of formula (IV) reacts with Buddhist nun sieve's methyl esters or Buddhist nun sieve's ethyl ester is 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1.1; Alkali is alkali (soil) metal alkoxide, preferred as alkali alkoxide, more preferably potassium tert.-butoxide or sodium tert-butoxide.
4. the compound of formula (II):
Wherein
R 1, R 2, R 3, R 4, m and n be as definition in claim 1, and
X represents leavings group, preferably halogen and triflate, and more preferably iodine,
Most preferably, the compound of formula (II) is the compound suc as formula (II-a) structure:
5. the method for the compound of the formula (II) of preparation claim 4, comprising:
Under the existence of thinner and optional under the existence of alkali, make the compound generation condensation reaction of compound and the formula V of formula (IV),
Wherein
R 1, R 2, R 3, m and n be as definition in claim 1;
Wherein
R 4, n is as definition in claim 1, and X represents leavings group, preferably halogen and triflate, more preferably iodine,
R brepresent halogen or hydroxyl;
In described method, the mol ratio of formula (IV) compound and the reaction of formula V compound is 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1; Suitable alkali is tertiary amine, preferably the mixture of DIPEA and DMAP.
6. the compound of formula (IV)
Wherein
R 1, R 2, R 3, m and n be as definition in claim 1;
Most preferably, the compound of formula (IV) is the compound suc as formula the structure of (IV-a):
7. the method for the compound of the formula (IV) of preparation claim 6, comprising:
Under the existence of thinner and optionally, under the existence of catalyzer and alkali, make formula (VI) compound and formula (VII) compound generation linked reaction,
Wherein
R 1with m as in claim 1 definition;
Wherein
R 2, R 3, n represents leavings group, preferably halogen and triflate, more preferably iodine as definition and X in claim 1;
In described method, the mol ratio of formula (VI) compound and the reaction of formula (VII) compound is 1:1 to 1:3, preferably 1:1 to 1:2, more preferably 1:1.5; Suitable catalyzer is Cu (I) or Cu (II) salt, as CuI, and Cu 2o or Cu (OAc) 2; Suitable alkali is mineral alkali or organic bases, wherein mineral alkali preferred alkali metal carbonate, more preferably K 2cO 3, Cs 2cO 3, organic bases is triethylamine or pyridine.
8. according to the compound of the formula in the preparation method of claim 7 (VI):
Wherein
R 1with m as in claim 1 definition;
Most preferably, the compound of formula (VI) is the compound suc as formula the structure of (VI-a):
9. the compound of the formula of claim 1 (I) and N-oxide compound thereof or its pharmacologically acceptable salt are for the preparation of the activity that suppresses receptor type tyrosine kinase or non-receptor type Tyrosylprotein kinase, preferably suppress receptor type TYR kinase activity, more preferably suppress the purposes of the medicine of ABL kinase activity.
10. the compound of the formula of claim 1 (I) and N-oxide compound thereof or its pharmacologically acceptable salt are for the preparation of the purposes of the medicine of inhibition tumor cell propagation, and described tumour cell comprises L1210, Jurkat E6-1, HL-60, A549, K562, RPMI8226, THP-1 and/or Ba/F3.
The compound of 11. formulas that comprise claim 1 (I) and N-oxide compound thereof or its pharmacologically acceptable salt and the optionally pharmaceutical composition of at least one pharmaceutically acceptable vehicle, described pharmaceutical composition preferably adopts the medicine type of subcutaneous administration.
The pharmaceutical composition of 12. claims 11 is for the preparation for the treatment of and the tyrosine kinase pathway relevant disease of lacking of proper care, the lack of proper care purposes of medicine of relevant disease of special and ABL kinase pathways.
The pharmaceutical composition of 13. claims 11 is for the preparation of the purposes of the medicine of the treatment disease, particularly cancer relevant to tumor cell proliferation, described cancer comprise leukemia, myelomatosis,
Stomach knurl and lung cancer.
CN201310006964.2A 2013-01-09 2013-01-09 Fluoro cyclobutyl imidazole compound Pending CN103910714A (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1938282A (en) * 2004-03-29 2007-03-28 默克专利有限公司 Imidazol derivatives as tyrosine kinase inhibitors
CN101045727A (en) * 2002-07-05 2007-10-03 诺瓦提斯公司 Inhibitors of tyrosine kinases
CN102316738A (en) * 2009-02-18 2012-01-11 盛泰萨路申有限公司 Amides as kinase inhibitors

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101045727A (en) * 2002-07-05 2007-10-03 诺瓦提斯公司 Inhibitors of tyrosine kinases
CN1938282A (en) * 2004-03-29 2007-03-28 默克专利有限公司 Imidazol derivatives as tyrosine kinase inhibitors
CN102316738A (en) * 2009-02-18 2012-01-11 盛泰萨路申有限公司 Amides as kinase inhibitors

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