CN102224155A - Protease inhibitors - Google Patents

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CN102224155A
CN102224155A CN2009801480408A CN200980148040A CN102224155A CN 102224155 A CN102224155 A CN 102224155A CN 2009801480408 A CN2009801480408 A CN 2009801480408A CN 200980148040 A CN200980148040 A CN 200980148040A CN 102224155 A CN102224155 A CN 102224155A
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methyl
alkyl
fluorine
disease
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V·伊瓦诺夫
B·塞缪尔森
P-O·约翰逊
P·坎伯格
H·维林
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Medivir AB
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Abstract

Compounds of the formula II, wherein R3 is C1-C3 alkyl or C3-C6 cycloalkyl, either of which is optionally substituted with one or two methyl and/or a fluoro, trifluoromethyl or methoxy, when R3 is C3-C6 cycloalkyl it may alternatively be gem substituted with fluoro; R4 is methyl or fluoro; m is 0, 1 or 2; E is a bond, or thiazolyl, optionally substituted with methyl or fluoro; A1 is CH or N, A2 is CR6R7 or NR6, provided at least one of A1 and A2 comprises N; n is 0 or 1 such that the ring containing A1 and A2 is a saturated, nitrogen-containing ring of 5 or 6 ring atoms; R6 is H, C1-C4 alkyl, C1-C4 haloalkyl, C1-C3 alkyl-O-C1-C3 alkyl, or when A2 is C, R6 can also be C1-C4 alkoxy or F; R7 is H, C1-C4 alkyl or F; or a pharmaceutically acceptable salt, N-oxide or hydrate thereof, have utility in the treatment of disorders characterized by inappropriate expression or activation of cathepsin K, such as osteoporosis, osteoarthritis, rheumatoid arthritis or bone metastases.

Description

Proteinase inhibitor
Invention field
The present invention relates to the inhibitor of L-Cysteine HCL Anhydrous, the particularly inhibitor of papoid superfamily.The invention provides and can be used for preventing or treat by the physiology proteolytic enzyme new compound of the disease that causes of cathepsin K imbalance for example.
Description of related art
The papoid superfamily of L-Cysteine HCL Anhydrous extensively distributes in the different plant species that comprises Mammals, invertebrates, protozoon, plant and bacterium.Some mammiferous kethepsins, comprise cathepsin B, F, H, K, L, O and S, be attributed to this superfamily, relate to its active improper adjusting in some metabolic disturbances, described metabolic disturbance comprises sacroiliitis, muscular dystrophy, inflammation, glomerulonephritis and tumour intrusion.Pathogenic kethepsin sample enzyme comprises bacillary gum mycoproteinase (gingipains), plasmodium falciparum L-Cysteine HCL Anhydrous (malarial falcipanins) I, II, III or the like and from Pneumocystis carinii (Pneumocystis carinii), the schizotrypanum cruzi (L-Cysteine HCL Anhydrous of Trypanosoma cruzei, trypanosoma bocagei (Trypanosoma brucei), (Crithidia fusiculata), schistosomicide (Schistosoma spp).
Comprise that at some diseases osteoporosis, gingival disease for example relate to the improper adjusting of cathepsin K in the hypercalcemia of gingivitis and periodontitis, osteitis deformans (Paget ' s disease), malignant tumour and the metabolic osteopathy.Consider that the level of cathepsin K in the chondroclast of osteoarthritic synovial membrane improves, cathepsin K relates to the disease that is characterized as excessive cartilage or substrate degradation, for example osteoarthritis and rheumatic arthritis.
Treatment osteopathy and richets for example osteoarthritis might need to use all the life cathepsin K inhibitor with osteoporosis, usually to the patient crowd's administration in the old age stage or the stage of approaching old age.This very high requirement places using on the easy degree of the medicine that is intended for use in these diseases.For example, attempt, the dosage that prolongs current diphosphate osteoid osteoporosis medicine to weekly or longer dosage regimen to help conformability.Yet even improve administration, other side effects of described diphosphate still keep.The conversion of diphosphate blocking-up bone, rather than as cathepsin K inhibitor, weaken the bone conversion.For the bone of health, it is very important keeping the remodeling process that diphosphate blocks fully.In addition, diphosphate has the very long transformation period in bone, and the osteonecrosis of jaw can prove itself if therefore for example act on, and it is impossible then removing diphosphate from bone.In contrast, cathepsin K inhibitor has usually and begins fast and finish the velocity function pattern, this means if problem has to be determinedly, and then can stop administration, inhibitor can not accumulated in ground substance of bone.
Therefore wish Gong the alternate osteoporosis and the osteoarthritis medicine that obtain having good pharmacokinetics and/or pharmacodynamic properties.
No. 2008/007107 open following formula: compound of International Patent Application WO
Figure BPA00001375069700021
Wherein Rd is the monocycle of replacement, and Rc is branched-chain alkyl or cycloalkyl, and Ra and Rb are various groups, comprise H, methyl, ethyl, ether, thioether, amine, sulfonate group or the like.Only compound through preparation contains H or methoxyl group on this position.
This area still needs effective inhibitor of cathepsin K.Demonstrate to cathepsin K but not to other kethepsins (for example selective) that optionally cathepsin K inhibitor is useful especially to cathepsin S and/or cathepsin L.Demonstrate effective inhibitor of the cathepsin K of hypertonicity for example and/or useful metabolism profile (profile), in clinical setting, can expect to have very big value.The relevant indication of cathepsin K for example osteoporosis or sacroiliitis expects to have administration cycle of prolongation, and therefore it is desirable to described compound has minimum toxicity or genotoxicity.
The invention summary
According to the present invention, provide formula II compound:
Figure BPA00001375069700031
Wherein
R 3Be C 1-C 3Alkyl or C 3-C 6Cycloalkyl, any is optional by one or two methyl and/or by fluorine, trifluoromethyl or methoxyl group replacement among both, works as R 3Be C 3-C 6During cycloalkyl, as selecting it can be by fluorine together with replacement;
R 4Be methyl or fluorine; M is 0,1 or 2;
E is a chemical bond or optional by the thiazolyl of methyl or fluorine replacement;
A 1Be CH or N,
A 2Be CR 6R 7Or NR 6, condition is A at least 1With A 2One of contain N;
R 6Be H, C 1-C 4Alkyl, C 1-C 4Haloalkyl, C 1-C 3Alkyl-O-C 1-C 3Alkyl, or work as A 2During for C, R 6Also can be C 1-C 4Alkoxyl group or F;
R 7Be H, C 1-C 4Alkyl or F
Or its pharmacy acceptable salt, N-oxide compound or hydrate.
Be to be understood that compound of the present invention can be used as hydrate and exists, for example the hydrate of following partial structural formula:
Figure BPA00001375069700032
And the present invention includes all so variable forms.
Be R aptly 3Be C 1-C 3Alkyl or C 3-C 6Cycloalkyl, any is optional by one or two methyl and/or by fluorine, trifluoromethyl or methoxyl group replacement among both.
For R 3, the representative group of cycloalkyl comprises cyclopropyl, cyclobutyl and particularly cyclopentyl or cyclohexyl, wherein arbitrary group replaces by fluorine or together with fluorine.Normally suitable at 2 of cyclopropyl, cyclobutyl or cyclopentyl 3 or cyclopropyl 4 together with fluorine.Also normally suitable at 4 of cyclohexyl together with fluorine.
In one embodiment of the present invention, R 3Represent leucic side chain.In second kind of embodiment of the present invention, R 3The side chain of expression Isoleucine.In the third embodiment of the present invention, R 3The side chain of expression Cyclohexylglycine.In the 4th kind of embodiment of the present invention, R 3The side chain of representative ring amyl group glycine.In the 5th kind of embodiment of the present invention, R 3The side chain of expression O-methylthreonine.In the 5th kind of embodiment of the present invention, R 3The leucic side chain of expression 4-fluorine.In the 6th kind of embodiment of the present invention, R 3The side chain of expression 3-methoxyl group Xie Ansuan.
Current R 3Preferred group comprise those groups that embody by following part-structure:
Figure BPA00001375069700041
In one embodiment of the present invention, m represents 2.Wherein m represents that 1 compound has special significance.Yet in the other embodiments of the present invention, m is 0, particularly when adjacent thiazolyl by Me or when preferably being replaced by F.
R 4Represent methyl or fluorine, particularly fluorine aptly.If m is 2, then current preferred each R 4Identical.
When m represents 1, R 4Suitably shown in following part-structure, located:
Figure BPA00001375069700042
E is chemical bond easily, and the ring that contains unsaturated nitrogen that promptly carries A1 and A2 is directly connected to the contraposition of phenyl ring.Yet current preferred E is a thiazolyl, and it is chosen wantonly by methyl or is more preferably replaced by fluorine.The preferred orientation of thiazole ring is:
Figure BPA00001375069700051
R wherein 5Be H, methyl or fluorine.
Contain A 1With A 2Ring be the saturated azo-cycle that contains of 5 or 6 annular atomses.Therefore, n is 0 or 1, and aptly, n is 1.Therefore representative ring comprises tetramethyleneimine-1-base, tetramethyleneimine-3-base, piperazine-1-base, piperidin-4-yl and piperidines-1-base.Described ring is easily by for example alkyl or haloalkyl, usually by methyl or propyl group or trifluoromethyl replacement.Perhaps described ring is by for example methoxymethyl or methoxy ethyl replacement of ether.Work as A 2During for carbon, but should encircle alkoxy for example methoxyl group or fluorine, particularly replace together with fluorine as alternatives.
The preferred embodiments of the invention have Formula Il a:
Figure BPA00001375069700052
Wherein
R 3C for side chain 2-C 6Alkyl or C 3-C 6Cycloalkyl, any is replaced by halogeno-group or trifluoromethyl among both;
R 4Be methyl or fluorine; M is 0 or 1 or 2;
R 5Be H, methyl or fluorine;
R 6Be C 1-C 4Alkyl;
Or its pharmacy acceptable salt, N-oxide compound or hydrate (this paper is referred to as compound of the present invention).
R 5Preferred fluorine is particularly when m is 0.Remaining optimum condition as mentioned among the formula II define.Should be understood to comprise the corresponding embodiment of formula IIa when hereinafter mentioning formula II.
The representative embodiment of formula II comprises:
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-3-methyl-butyl]-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclohexyl-2-oxo-ethyl]-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclopentyl-2-oxo-ethyl]-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-2-methyl-butyl]-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-3-methyl-butyl]-4-[5-methyl-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclohexyl-2-oxo-ethyl]-4-[5-methyl-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclohexyl-2-oxo-ethyl]-4-[5-methyl-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-2-methyl-butyl]-4-5-methyl-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-3-methyl-butyl]-4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclohexyl-2-oxo-ethyl]-4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclopentyl-2-oxo-ethyl]-4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-2-methyl-butyl]-4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-3-methyl-butyl]-3-fluoro-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclohexyl-2-oxo-ethyl]-3-fluoro-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-yl)-1-cyclopentyl-2-oxo-ethyl]-3-fluoro-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide;
N-[1-(6-nitrile-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-2-methyl-butyl]-3-fluoro-4-[2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzamide
And pharmacy acceptable salt, N-oxide compound and hydrate.
R 6Or R 7C 1-C nAlkyl definition is intended to comprise and contains side chain and unbranched both moieties of sum between one and n carbon atom.Such R 6Group or R 7Example be methyl, ethyl, propyl group (n-propyl and sec.-propyl), butyl (normal-butyl, isobutyl-, the tertiary butyl and sec-butyl).A R of particularly important 6Group is a methyl.Second R of particularly important 6Group is propyl group (a particularly n-propyl).Work as R 3Optional during by one or two methyl substituted, this part is the definable branched alkyl chain of 5 C atoms nearly also.
In certain embodiments of the invention, A 1Be N.
Other aspects of the present invention comprise the pharmaceutical composition that contains compound carrier pharmaceutically acceptable with it as hereinbefore defined or thinner.
Another aspect of the present invention is that compound is used for the treatment of purposes in the medicine disease mediated by cathepsin K in preparation as hereinbefore defined, described disease for example:
Osteoporosis,
Gingival disease (for example gingivitis and periodontitis),
Osteitis deformans,
The hypercalcemia of malignant tumour,
Metabolic osteopathy,
Be characterized as excessive cartilage or substrate degradation disease (for example osteoarthritis and rheumatic arthritis),
Osteocarcinoma, comprise tumorigenesis,
Pain (particularly chronic pain).
Be provided in addition treating or prevent and comprise the The compounds of this invention of using safety and significant quantity, with treatment or prevention described disease by the cathepsin K mediation by the disease mediated method of cathepsin K.
Also be provided for treating or prevent The compounds of this invention by the disease of cathepsin K mediation.
In addition, provide new intermediate (as described herein) as one aspect of the present invention, it can be useful in the preparation of The compounds of this invention.
Following formula: compound is provided especially:
Figure BPA00001375069700081
Or the protected derivative of its N (for example by Boc, CBz or Fmoc protection).The present invention also provides it corresponding 1, the analogue and the protected derivative of N (for example Boc CBz or Fmoc are protected) of the protection of 3-dioxolane.Another new intermediate of the present invention has following formula:
Figure BPA00001375069700082
Or corresponding 1, the analogue of 3-dioxolane protection, N official can choose wantonly by for example Boc, CBz or the Fmoc protection of GPF (General Protection False base under each situation.
Compound of the present invention can form salt, and it constitutes another aspect of the present invention.The suitable pharmacy acceptable salt of formula II compound comprises: organic acid salt, the salt of carboxylic acid particularly, include but not limited to acetate, trifluoroacetate, lactic acid salt, gluconate, Citrate trianion, tartrate, maleate, malate, pantothenate, isethionate, adipate, alginate, aspartate, benzoate, butyrates, digluconate, chaulmoogric acid salt, the glucose enanthate, glycerophosphate, oxalate, enanthate, hexanoate, fumarate, nicotinate, pamoate, pectate, 3-phenylpropionic acid salt, picrate, pivalate, propionic salt, tartrate, lactobionate, Pivalate, camphorate, undecylate and succinate, organic sulfonate is mesylate for example, esilate, the 2-isethionate, camsilate, the 2-naphthalenesulfonate, benzene sulfonate, closilate and tosilate; And inorganic acid salt, for example hydrochloride, hydrobromate, hydriodate, vitriol, hydrosulfate, Hemisulphate, thiocyanate-, persulphate, phosphoric acid salt and sulfonate.
It is separated that compound of the present invention can be used as hydrate in some cases.Hydrate is usually by with an organic solvent for example two
Figure BPA00001375069700091
English, tetrahydrofuran (THF) or methyl alcohol recrystallization from moisture/ORGANIC SOLVENT MIXTURES prepares.Hydrate also can original position produce by use corresponding ketone to the patient.
The N-oxide compound of The compounds of this invention can be by the known method preparation of those of ordinary skills.For example, the N-oxide compound can prepare by the not oxidised form with oxygenant (for example trifluoro Peracetic Acid, peroxy maleic acid, benzoyl hydroperoxide, Peracetic Acid, metachloroperbenzoic acid or its analogue) processing The compounds of this invention in about 0 ℃ of inert organic solvents (for example halon, for example methylene dichloride) that is suiting.Perhaps, the N-oxide compound of The compounds of this invention can be from suitable raw-material N-oxide compound preparation.
The example of N-oxide compound of the present invention comprises the N-oxide compound with following part-structure:
Figure BPA00001375069700092
Not the The compounds of this invention of oxidised form can in 0 to 80 ℃ at suitable inert organic solvents (for example acetonitrile, ethanol, moisture two
Figure BPA00001375069700093
Alkane or its similar organic solvent) in by handling with reductive agent (for example sulphur, sulfurous gas, triphenylphosphine, lithium borohydride, sodium borohydride, phosphorus dichloride, phosphorus tribromide or its similar reductive agent), prepare from the N-oxide compound of respective compound of the present invention.
Should be noted that the group position on any molecular moiety of using in the definition as long as chemically stablize, can be any position on this part.
The group that uses in the variable-definition comprises all possible isomer, except as otherwise noted.For example butyl comprises the tertiary butyl, isobutyl-, normal-butyl or the like.
When any variable occurred surpassing one time in any composition, each definition was independently.
Unless mentions in addition or illustrate, otherwise the chemical name of compound comprises the mixture of isomeric forms on all possible stereochemistry that described compound may have.Described mixture can comprise all diastereomers and/or the enantiomorph of described compound basic molecular structure.All form of three-dimensional chemical isomer with pure form or mutual blended The compounds of this invention are all expected within the scope of the present invention.
The pure stereoisomers form of compound that this paper is mentioned and intermediate is defined as other enantiomorphs that do not have described compound or the identical basic molecular structure of intermediate basically or the isomer of diastereomer form.Specifically, term " stereochemistry is pure " relates to and has stereoisomerism excessive at least 80% (being a kind of isomer minimum 90% and other possible isomer maximums 10%) the nearly compound or the intermediate of stereoisomerism excessive 100% (i.e. 100% a kind of isomer and do not have other isomer), more particularly, have excessive 90% to 100% compound of stereoisomerism or intermediate, even more particularly have excessive 94% to 100% compound of stereoisomerism or intermediate, most particularly have stereoisomerism excessive 97% to 100% compound or intermediate.Term " enantiomer-pure " should be understood by similar mode to " diastereomer is pure ", just with regard to separately enantiomeric excess and diastereomeric excess in the mixture in discussing.
Compound of the present invention can be used as its independent steric isomer preparation with the optical resolution agent reaction to form a pair of diastereomer compound, separate diastereomer and to reclaim optically active enantiomorph by the racemic mixture with described compound.When although the fractionation of enantiomorph can use the covalency non-enantiomer derivative of formula II compound to carry out, preferably can dissociated mixture (salt of for example crystallization, diastereomer).Diastereomer has different physical properties (for example fusing point, boiling point, solubility, reactivity or the like), and can be by utilizing the very fast separation of these differences.Diastereomer can pass through for example HPLC separation of chromatography, or preferably separates by the separation/disassemble technique based on solubility difference.Then by any practical approach that can not cause racemization with resolving agent, reclaim optically pure enantiomorph.Being applicable to can be at Jean Jacques Andre Collet, Samuel H.Wilen, Enantiomers, Racemates and Resolutions, John Wiley ﹠amp by the more detailed description of the technology of racemic mixture compound steric isomer; Sons, Inc. finds in (1981).
Any subgroup compound of formula II compound defined herein or formula II comprises radio isotope or radiolabeled compound, one of them or more a plurality of atom are replaced by the isotropic substance of this atom, and isotropic substance is promptly identical with the ordination number of the atom of finding at occurring in nature usually but atom that atomic mass is different.Can be introduced into the isotropic substance example of any subgroup compound of formula I compound or formula I, include but not limited to: the isotropic substance of hydrogen, for example 2H and 3H (also being expressed as D respectively is that deuterium and T are tritium); The isotropic substance of carbon, for example 11C, 13C and 14C; The isotropic substance of nitrogen, for example 13N and 15N; The isotropic substance of oxygen, for example 15O, 17O and 18O; The isotropic substance of phosphorus, for example 31P and 32P; The isotropic substance of sulphur, for example 35S; The isotropic substance of fluorine, for example 18F; The isotropic substance of chlorine, for example 36Cl; The isotropic substance of bromine, for example 75Br, 76Br, 77Br and 82Br; And the isotropic substance of iodine, for example 123I, 124I, 125I and 131I.
Included isotopic selection will be depended on the concrete application of this compound in the compound isotopically labelled.For example for medicine or the analysis of substrate tissue distribution, for example wherein introduced radio isotope 3H or 14The compound of C can be the most useful usually.For radiophotography application examples such as positron emission tomography instrument (PET), the positron radiation isotropic substance for example 11C, 18F, 13N or 15O can be useful.Higher isotope for example deuterium is 2The introducing of H, any subgroup compound that can be formula I compound or formula I provides bigger metabolic stability, and it can cause that for example the interior transformation period of body of compound prolongs or dosage requires to reduce.
Synthetic for convenience, preferred formula II compound is in the isotropic substance state of nature usually.
Any subgroup compound of isotope-labeled formula I compound or formula II can be through using suitable isotope-labeled reagent or starting material rather than corresponding nonisotopically labelled reagent or starting material, by with hereinafter flow process and/or embodiment in the similar method preparation of those methods described, or prepare by routine techniques well known by persons skilled in the art.
Will be appreciated that the present invention extends to prodrug, solvate, mixture and discharges other forms of The compounds of this invention in vivo.
Though promoting agent is individually dosed is possible, and preferred its exists as the part of pharmaceutical preparation.Such preparation will comprise above defined promoting agent and one or more plant acceptable carrier/vehicle and optional other treatment composition.Described carrier is from saying it must is acceptable with the compatible meaning of other compositions of preparation, and harmless to the recipient.
Described preparation comprises the preparation that is suitable for rectum, nose, part (comprising oral cavity and hypogloeeis), vagina or parenteral (comprising subcutaneous, intramuscular, intravenously and intracutaneous) administration, but preferred formulation is an oral medicinal preparation.Described preparation can exist with unit dosage easily, for example tablet and slow releasing capsule, and can be by any method preparation of knowing in the pharmaceutical field.
These methods comprise the step that above defined promoting agent and carrier are combined.Generally speaking, described preparation makes formed product prepare by evenly and closely combining with promoting agent and liquid carrier or solid carrier in small, broken bits or with the two then if necessary.The present invention extends to the method that is used for pharmaceutical compositions, and it comprises formula II compound or its pharmacy acceptable salt and pharmaceutically acceptable carrier or solvent are combined.If the preparation of pharmaceutical preparation relates to drug excipient and mixes with the tight of salt form activeconstituents, then preferred usually the use is the vehicle of non-alkalescence in nature, i.e. acidity or neutral vehicle.
For example every kind of discrete unit existence that comprises capsule, cachet or the tablet of predetermined amount promoting agent of oral medicinal preparation among the present invention; Exist with powder or granule; Exist with the solution or the suspensoid of promoting agent in liquid, aqueous or on-aqueous liquid; Or exist and exist with bolus (bolus) with oil-in-water liquid emulsion or water-in-oil liquid emulsion etc.
About oral administration composition (for example tablet and capsule), the suitable carrier of term comprises solvent, such as ordinary excipients, for example tackiness agent, for example syrup, gum arabic, gelatin, sorbyl alcohol, tragacanth gum, polyvinylpyrrolidone (polyvidone), methylcellulose gum, ethyl cellulose, Xylo-Mucine, hydroxypropyl-methylcellulose gum, sucrose and starch; Weighting agent and carrier, for example W-Gum, gelatin, lactose, sucrose, Microcrystalline Cellulose, kaolin, N.F,USP MANNITOL, secondary calcium phosphate, sodium-chlor and Lalgine; Reach lubricant for example Magnesium Stearate, sodium stearate and other metallic stearates, stearin, stearic acid, silicone oil, talcum, wax, oil and colloided silica.Also can use correctives for example spearmint oil, wintergreen oil, cherry seasonings or its analogue.The adding tinting material is discerned formulation easily may be satisfactory.Tablet also can carry out dressing by the method for knowing in this area.
Tablet can pass through compressing tablet or mold pressing, chooses wantonly with one or more kind auxiliary agents to prepare.The tablet of compacting can be by preparing described promoting agent optional and tackiness agent, lubricant, inert diluent, sanitas, tensio-active agent or dispersant with the promoting agent compressing tablet of free-flowing form (for example powder or particle) in suitable machine.The tablet of mold pressing can prepare by in suitable machine the mixture with the moistening powdered compounds of inert liquid diluent being carried out mold pressing.Described tablet can be chosen in addition dressing or cut wantonly and can prepare so that the promoting agent of slowly-releasing or controlled release is provided.
Other preparations that are suitable for oral administration comprise lozenge, and it contains promoting agent in flavoured base (being generally sucrose and gum arabic or tragakanta); Lozenge, it contains promoting agent in inert base (for example gelatin and glycerine) or sucrose and gum arabic; And mouth wash shua, it contains promoting agent in suitable liquid carrier.
The suitable dose of The compounds of this invention or preparation will depend on indication and patient, and determine by the conventional animal test easily.Provide be about 0.01-100uM, more preferably the dosage of (being used to suppress the physiology proteolytic enzyme of papoid superfamily) concentration is satisfactory usually and can realize in the cell of 0.01-10uM, for example 0.1-25uM.
Compound of the present invention prepares by various liquid phases and mechanochemical method.
Described compound prepares with P1, the P2 of reflection end product inhibitor and the structural unit of P3 part usually.Do not wish to accept by any way opinion or the specifically constraint of the tentative binding pattern attribution of variable, the employed abstract concept P1 of this paper, P2 and P3 only provide for convenient, and have its conventional Schlecter ﹠amp basically; Berger implication and representing is considered to occupy respectively those parts of S1, S2 and the S3 sublocus of enzyme in the inhibitor, wherein S1 closes on cleavage site, and S3 is away from cleavage site.Which kind of binding pattern the compound plan of formula II definition is regardless of it within the scope of the invention.
In the broadest sense, described P1 structural unit will have following formula:
Figure BPA00001375069700141
Wherein
R 1And R 2As hereinbefore defined, two Rb group definition ketals, dimethyl ketal for example, or define cyclic ketal for example 1,3-dioxolane together;
And Rc is a hydroxy-protective group.Insight more seldom, in situation about being prolonged by P2 and P3 as the P1 structural unit of ketone, Rc is H or the ketone official energy of representing the end product inhibitor.
WO05/066180 has described the preparation about the intermediate of above-mentioned P1 structural unit, and described intermediate comprises:
Figure BPA00001375069700142
In The compounds of this invention synthetic, the fs is the functionalized P1 structural unit of preparation in solution normally.The route of the 6-aldehyde intermediate that flow process 1 explanation is suitable.
Figure BPA00001375069700151
I) Dai Si-Martin's oxygenant, DCM; Ii) trimethyl orthoformate, pTs, MeOH:iii) Pd (OH) 2, H 2, MeOH; Iv) Boc 2O, 10%Na 2CO 3, v) Dai Si-Martin's oxygenant, DCM:vi) 1) and CH 3PPh 3Br, KOtBu, THF; Vii) 1) 9-BBN-H, THF, 2) NaBO 3, H 2O, THF; Viii) Dai Si-Martin's oxygenant, DCM.
Flow process 1
Initial bicyclol (1a) can be by preparing of describing among the WO05/066180.For example use Dai Si-Martin's oxygenant (Dess-Martin Periodinane) oxidation hydroxyl-functional, then handle with trimethyl orthoformate as passing through in the presence of the tosic acid, realize that the ketone group official that will produce can be converted into dimethyl ketal ketal (1b) is provided in acid.Use catalyzer such as Pd (OH) 2Or its analogue realizes the removal of Cbz and benzyl protecting group for example by hydrogenolytic cleavage, then the unhindered amina that produces carried out the boc protection, and alcohol (1c) is provided.Use the free alcohol that oxidation produces in solvent such as methylene dichloride of Dai Si-Martin's oxygenant for example, then in the presence of KOt.Bu or its analogue, use the Diethylaminoethyl triphenylphosphine
Figure BPA00001375069700152
Carry out wittig reaction (Wittig reaction) so that alkene (1d) to be provided.For example realize two key hydroxylations by handling with 9-BBN-H, primary alconol (1e) is provided, it can use any suitable oxidation style for example to handle with Dai Si-Martin's oxygenant or its analogue subsequently, is oxidized to corresponding aldehyde (1f).
Flow process 2 explanations are from the typical method of the initial 6-nitrile P1 structural unit of the 6-aldehyde intermediate of flow process 1.
Figure BPA00001375069700161
Aldehyde 1f (flow process 2) is converted into the conversion of corresponding required nitrile 1h (flow process 2) to be undertaken by making oxime 1g (flow process 2) dehydration.Therefore the aldehyde that is dissolved in ethanol/water mixture can be used NH 2OH and sodium acetate are handled, for example in ambient temperature overnight.Available TLC monitoring starting material consume.Conventional aftertreatment (work-up) provides thick oxime 1g (E and Z isomer), and it can not need to be further purified and use.Thick oxime 1g can dissolve in methylene dichloride and add triethylamine in-78 ℃.Can add and be dissolved in for example trifluoromethane sulfonic acid acid anhydrides in the methylene dichloride of organic solvent, allow reaction be warmed to room temperature usually simultaneously.In case as if reaction finished, just resistates is extracted and chromatographic separation, for example separate in the enterprising circumstances in which people get things ready for a trip spectrum of silica gel, produce typical nitrile structural unit 1h, the normal yield height.Usually unique a kind of isomer of nitrile 1h for example has the stereochemical nitrile 1h of C-6S and is separated, because reaction is carried out can not losing the stereochemistry integrity usually.Numerous examples show in the document, are keeping similar stereochemistry to carry out from aldehyde to this conversion of corresponding nitrile, and described document is people such as Hutt for example, Journal of Organic Chemistry, 72 (26), 10130,2007.
Be generally and obtained final compound, P1 structural unit for example above 1h carries out the N deprotection in a usual manner, for example handles to remove the N-Boc protecting group with the Acetyl Chloride 98Min. that is dissolved in methyl alcohol.For unhindered amina subsequently, use for example DIPEA among HATU, the DMF of standard coupling condition, for example introduce the P2 residue by BocP2-OH.Remove terminal Boc protection with the Acetyl Chloride 98Min. that is dissolved in methyl alcohol once more, and the standard of use coupling condition for example the DIPEA among HATU, the DMF introduce the P3 residue by P3-OH.Remove the dimethyl ketal protection with TFA at last, produce required final compound.
Prolong usually at the suitable coupling agent basic oxygen base of phosphofluoric acid benzotriazole-1-tripyrrole alkyl phosphorus for example
Figure BPA00001375069700171
(PyBOP), phosphofluoric acid O-benzotriazole-1-base-N, N, N ', N '-tetramethyl--urea (HBTU), phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-1,1,3,3-tetramethyl--urea
Figure BPA00001375069700173
(HATU), 1-(3-dimethylaminopropyl)-3-ethyl-carbodiimide hydrochloride (EDC) or 1,3-dicyclohexylcarbodiimide (DCC) exists down, choose wantonly at I-hydroxybenzotriazole (HOBT) and alkali N for example, carry out under the existence of N-diisopropylethylamine, triethylamine, N-methylmorpholine etc.Reaction is carried out at 20 to 30 ℃ usually, preferably carries out at about 25 ℃, needs finish in 2 to 24 hours.Suitable reaction solvent is an inert organic solvents, for example halogenated organic solvent (for example methylene dichloride, chloroform etc.), acetonitrile, N, dinethylformamide, like the solvent of ether tetrahydrofuran (THF), two for example
Figure BPA00001375069700174
Alkane etc.
Perhaps, prolongation coupling step above can be by at first being converted into the P3/P2 structural unit for example succinimide ester of active acid derivant, itself and P1 amine reacted carry out then.Reaction needed finish in 2 to 3 hours usually.The condition of using in this reaction depends on the character of active acid derivant.If for example it is a chloride derivative, is reflected at so under the existence of suitable alkali (for example triethylamine, diisopropylethylamine, pyridine etc.) and carries out.Suitable reaction solvent is a polar organic solvent, for example acetonitrile, N, dinethylformamide, methylene dichloride or its any suitable mixture.
The P2 structural unit is for example L-leucine, L-Isoleucine, O-methyl-L-Threonine, L-3-hydroxyvaline, 4-fluorine leucine, L-cyclopentyl glycine or L-Cyclohexylglycine of the protected amino acid of N normally; and P3 comprises capping group usually; for example have for example phenylformic acid of N-alkyl-piperazinyl-E part in contraposition, this N-alkyl-piperazinyl-E part has been introduced into or provides with its synthon.
Suitably protected one structural unit can at first prepare and coupling together subsequently, preferably coupling in the following order: P2+P1 → P2-P1, then N-alkylpiperazine base-E-phenylformic acid *+ P2-P1 → N-alkylpiperazine base-E-benzoic ether-P2-P1, wherein *The expression activated form is so that reduce the racemization of P2.
Between two seed amino acids, between one seed amino acid and a kind of peptide, or the coupling between two kinds of peptide fragment can use for example following method of standard coupling method to carry out: the trinitride method, mixed carbonic acid-carboxylic acid anhydride (carbonic-carboxylic acid anhydride) (isobutyl chlorocarbonate) method, carbodiimide (dicyclohexylcarbodiimide, DIC, or water-soluble carbodiimide) method, active ester (p-nitrophenyl ester, N-hydroxy succinic acid imide ester) method, Woodward reagent K-method, the carbonyl dimidazoles method, phosphorus reagent or oxidation-reduction method.Some method in these methods (particularly carbodlimide method) can be strengthened by adding I-hydroxybenzotriazole or 4-DMAP.These linked reactions can be carried out in solution (liquid phase) or solid phase.
More particularly, the free carboxy that relates in the presence of coupling agent a kind of reactant of coupling step is connected amido linkage with the free amine group dehydration coupling of other reactants to form.Generally speaking the description of such coupling agent can be found on the textbook of chemistry of peptides, M.Bodanszky for example, " Peptide Chemistry ", the second edition revised edition, Springer-Verlag, Berlin, Germany, (1993), it abbreviates Bodanszky hereinafter as, and its content is hereby incorporated by.The example of suitable coupling agent is N, N '-dicyclohexylcarbodiimide, at N, I-hydroxybenzotriazole or N-ethyl-N '-[(3-dimethylamino) propyl group] carbodiimide under N '-dicyclohexylcarbodiimide exists.Practical, useful coupling agent is commercially available phosphofluoric acid (benzotriazole-1-base oxygen base) three-(dimethylamino) phosphines Itself, or the phosphofluoric acid in the presence of I-hydroxybenzotriazole or 4-DMAP (benzotriazole-1-base oxygen base) three-(dimethylamino) phosphines
Figure BPA00001375069700182
Another kind of practicality, useful coupling agent are commercially available Tetrafluoroboric acid 2-(1H-benzotriazole-1-yl)-N, N, N ', N '-tetramethyl-urea
Figure BPA00001375069700183
Another practicality, useful coupling agent are commercially available phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-N, N, N ', N '-tetramethyl-urea
Figure BPA00001375069700184
Described linked reaction is for example carried out in methylene dichloride, acetonitrile or the dimethyl formamide at inert solvent.Add excessive tertiary amine such as diisopropylethylamine, N-methylmorpholine, N-crassitude or 4-DMAP and be approximately 8 with the pH that keeps reaction mixture.Temperature of reaction is usually in the scope between 0 ℃ and 50 ℃, and the reaction times is usually in the scope between 15 minutes and 24 hours.
Functional group as the alpha-non-natural amino acid of component usually must be protected to avoid formation not need chemical bond during linked reaction.Spendable protection is based on Greene, " Protective Groups in Organic Chemistry ", John Wiley; Sons, New York (1981) and " The Peptides:Analysis, Synthesis, Biology ", the 3rd volume, Academic Press, New York lists in (1981), and hereinafter referred is Greene, and its prospectus is hereby incorporated by.
The common protected one-tenth ester of the α-carboxyl of C-terminal residue, this ester can be cleaved and be obtained carboxylic acid.Spendable protecting group comprises: 1) alkyl ester; for example methyl ester, trimethyl silyl ester and tertiary butyl ester, 2) aralkyl ester, for example benzyl ester of benzyl ester and replacement; or 3) through weak base or gentle reduction method can be cleaved ester, for example trichloro ethyl ester and phenacyl ester.
Normally N is protected to remain each amino acid whose alpha-amino group of link coupled.Can use any protecting group known in the art.These examples of groups comprise: 1) acyl group, for example formyl radical, trifluoroacetyl group, phthaloyl base and p-toluenesulfonyl; 2) aromatic urethanes base, for example benzyloxycarbonyl and the 9-fluorenyl methoxy carbonyl (Fmoc) of benzyloxycarbonyl (Cbz or Z) and replacement; 3) aliphatic carbamate base, for example tertbutyloxycarbonyl (Boc), ethoxy carbonyl, di-isopropyl methoxyl group-carbonyl and allyloxy carbonyl; 4) cycloalkyl amino formic acid ester group, for example cyclopentyloxy carbonyl and Buddha's warrior attendant alkoxy carbonyl; 5) alkyl, for example trityl group and benzyl; 6) trialkylsilkl, for example trimethyl silyl; And 7) contain thiol group, for example thiophenyl carbonyl and dithia succinyl.Preferred alpha-amino group protecting group is Boc or Fmoc.It is commercially available that many due cares are used for peptide synthetic amino acid derivative.
The alpha-amino group protecting group is cut before next coupling step usually.When using the Boc base, the method for selection is pure trifluoroacetic acid or the trifluoroacetic acid that is dissolved in methylene dichloride, or is dissolved in two
Figure BPA00001375069700191
The HCl of alkane or ethyl acetate.Then before coupling or original position with basic solution water-containing buffering liquid for example, or the tertiary amine that is dissolved in methylene dichloride or acetonitrile or dimethyl formamide is with the ammonium salt neutralization that is produced.When using the Fmoc base, selective reagent is to be dissolved in the piperidines of dimethyl formamide or the piperidines of replacement, but can use any secondary amine.Deprotection carries out between 0 ℃ of temperature and the common 20-22 of room temperature ℃.
In case the inhibitor sequence is finished, then with according to the selection of protecting group fixed any form remove the protecting group of any remnants.These methods are well known to those skilled in the art.
The P2 structural unit of the protected L-amino acid form of N is easily in commercial acquisition; for example L-leucine, L-Isoleucine, L-Cyclohexylglycine, O-methyl-L Threonine and other amino acid obtain commercial; many protecting group varients are arranged, for example CBz, Boc or Fmoc.R 3Other varients prepare from commercially available starting material easily.R wherein for example 3For-C (CH 3) 2OCH 3Compound can be by (S)-(+)-2-amino-3-hydroxy-3-methyl butyric acid and 3 that will be protected by CBz, (3,2b) pyrroles's reaction prepares to form required P2-P1 unit 3-dimethoxy-six hydrogen-furo.P2 side chain alcohol can use methyl iodide to methylate to obtain required P2 under conventional sodium cyanide, imidazoles, THF condition now, and the α center is not substantially by racemization.This P2-P1 part now can be by described herein synthetic, i.e. CBz removal and coupling are carried out.
WO05/565299 has described the preparation of γ-fluorine leucine P2 structural unit.The another kind of Fmoc and N-Boc-γ fluorine leucine structural unit is synthetic as shown in people Syn.Lett.2005 no 8 1278-1280 such as Truong.
The preparation of P3 structural unit is described among WO05/066180, the WO08/007114 or prepares by similar method easily.For example flow process E hereinafter shows the preparation of P3 structural unit, wherein the thiazolyl of E for being replaced by fluorine:
I.HOAc, Br 2, RT, 2h, productive rate 55%; Ii.KF, 18-hat-6, CH 3CN, 90 ℃, 16h, productive rate 31%; Iii.HOAc, Br 2, 45C, 4h, productive rate 100%; Iv.4-methylpiperazine-1-thioformamide, ethanol, 70 ℃, 2h, productive rate 74%, v.LiOH, THF, H 2O, RT, 16h, productive rate 79%.
Flow process E
4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-benzoic synthetic raw material 4-acetylbenzoic acid methyl esters obtains commercial.Realize bromination with the bromine that is dissolved in the acetate at alpha-position, required 4-(2-bromo-ethanoyl)-methyl benzoate is provided with respect to ketone.In the presence of 18-hat-6, handle 4-(2-bromo-ethanoyl)-methyl benzoate in 90 ℃ subsequently, 4-(2-fluoro-ethanoyl)-methyl benzoate is provided after column chromatography with Potassium monofluoride.Repeat bromination with the bromine that is dissolved in the acetate at alpha-position, required 4-(2-bromo-2-fluoro-ethanoyl)-methyl benzoate is provided with respect to ketone.The formation of thiazole usually by carrying out in 70 ℃ of heating 4-(2-bromo-2-fluoro-ethanoyl)-methyl benzoate and 4-methylpiperazine-1-thioformamide in 2 hours.In the process of cooling, required 4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-methyl benzoate is settled out.The deprotection of methyl esters uses lithium hydroxide solution and required acid to carry out, and when carrying out aftertreatment with hydrochloric acid, obtains 4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl with good output usually]-phenylformic acid is a dihydrochloride.
Term used herein " N-protected base " or " N is protected " are meant and are intended for use in protecting the N-end of amino acid or peptide or amino those groups of avoiding undesired reaction of protection in building-up process.N-protected base commonly used is disclosed in Greene, " Protective Groups in Organic Synthesis " (John Wiley ﹠amp; Sons, New York, 1981), it is hereby incorporated by.The N-protected base comprises: acyl group, for example formyl radical, ethanoyl, propionyl, valeryl, tertiary butyl ethanoyl, 2-chloracetyl, 2-acetyl bromide, trifluoroacetyl group, tribromo-acetyl base, phthaloyl base, ortho-nitrophenyl oxygen base ethanoyl, α-chlorobutyryl, benzoyl, 4-chlorobenzene formacyl, 4-benzoyl bromide, 4-nitro benzoyl etc.; Alkylsulfonyl, for example benzenesulfonyl, p-toluenesulfonyl etc.; Form the group of carbamate, benzyloxycarbonyl for example, to the chlorine benzyloxycarbonyl, to methoxyl group benzyloxy base carbonyl, to the nitro benzyloxycarbonyl, 2-nitro benzyloxycarbonyl, to the bromo-benzyloxy-carbonyl, 3,4-dimethoxy benzyloxycarbonyl, 4-methoxyl group benzyloxy base carbonyl, 2-nitro-4,5-dimethoxy benzyloxycarbonyl, 3,4,5-trimethoxy benzyloxycarbonyl, 1-(to xenyl)-1-methyl ethoxy carbonyl, α, alpha-alpha-dimethyl-3,5-dimethoxy benzyloxycarbonyl, the hexichol methoxycarbonyl, tert-butoxycarbonyl, the di-isopropyl methoxycarbonyl, isopropoxy carbonyl, ethoxy carbonyl, methoxycarbonyl, allyloxy carbonyl, 2,2,2-trichlorine ethoxy carbonyl, phenyloxycarbonyl, the 4-nitrophenoxy carbonyl, fluorenyl-9-methoxycarbonyl, the cyclopentyloxy carbonyl, the Buddha's warrior attendant alkoxy carbonyl, cyclohexyloxy carbonyl, phenyl thiocarbonyl group etc.; Alkyl, for example benzyl, trityl group, benzyloxymethyl etc.; And silyl, for example trimethyl silyl etc.Favourable N-protected base comprises formyl radical, ethanoyl, benzoyl, valeryl, tertiary butyl ethanoyl, benzenesulfonyl, benzyl (bz), tert-butoxycarbonyl (BOC) and benzyloxycarbonyl (Cbz).
The protecting group of hydroxyl and/or carboxyl also has summary widely in Greene (ditto), comprise: ether, the for example methyl ether of methyl ether, replacement, for example methoxymethyl, methylthiomethyl, benzyloxymethyl, tert.-butoxy methyl, 2-methoxy ethoxy methyl etc.; Silyl ether, for example trimethyl silyl (TMS) ether, t-butyldimethylsilyl (TBDMS) ether, tribenzyl silyl ether, triphenyl silyl ether, t-butyldiphenylsilyl ether, triisopropyl silyl ether etc.; The ether that replaces, for example 1-ethoxyl methyl, 1-methyl isophthalic acid-methoxy ethyl, tertbutyl ether, allyl ethers, benzylic ether, to methoxy-benzyl ether, diphenyl methyl ether, trityl group ether etc.; Aralkyl, for example trityl and phenyl xanthenyl (pixyl) (9-hydroxyl-9-phenyl xanthene derivative, particularly muriate).The ester hydroxyl protecting group comprises ester, for example manthanoate, benzyl manthanoate, chloracetate, methoxyacetic acid ester, phenoxy acetic acid ester, pivalate, Buddha's warrior attendant acid esters,
Figure BPA00001375069700221
Acid esters, benzoic ether etc.The carbonic ether hydroxyl protecting group comprises methyl ethylene, allyl group, cinnamyl, benzyl etc.
Embodiment describes in detail
Only by explanation various embodiments of the present invention are described now according to following examples.
Reference example 1
The P3 structural unit
Step a) 4-cyano group Propiophenone
Figure BPA00001375069700222
According to relevant preparation 4-cyano-acetophenone described (Synth.Commun 1994,887-890), with the 4-brom-acetophenone (5.65g, 26.4mmol), Zn (CN) 2(1.80g, 15.3mmol) and Pd (PPh 3) 4(2.95g, (35mL uses mixture 2.6mmol) at deoxidation DMF in 80 ℃
Figure BPA00001375069700223
Molecular sieve stores, and feeds Ar before using) the middle backflow 18 hours.Mixture is at toluene (100mL) and 2N NH 4Distribute between the OH (100mL).Organic phase 2N NH 4OH (100mL) extraction, with the saturated NaCl aqueous solution (2x100mL) washing, dry and evaporation.Carry out the 10mmol scale reaction similarly and crude product is merged.Flash chromatography separates (330g silicon-dioxide, 6/1 sherwood oil-EtOAc), obtain white solid (5.17g, 89%).
1H?NMR(CDCl 3)δppm：1.22(t，3H，J＝7.2Hz)，3.00(q，2H，J＝7.3Hz)，7.75(d，2H，J＝8.8Hz)，8.03(d，2H，J＝8.4Hz)
13C?NMR(CDCl 3)δppm：7.8，32.1，116.1，117.9，128.3，132.4，139.7，199.2
Step b) 4-propionyl phenylformic acid
Figure BPA00001375069700231
(4.67g, 29.3mmol) (90mL is 180mmol) with two with 2N NaOH with 4-cyano group Propiophenone
Figure BPA00001375069700232
Alkane (90mL) spends the night in 95 ℃ of backflows.Mixture water (150mL) dilution, with ether (75mL) washing, dense HCl is acidified to pH 2 and uses ether (3x 75mL) extraction.Organic phase is also evaporated with the saturated NaCl aqueous solution (3x75mL) washing, drying, obtains yellow solid (5.12g, 98%).
1H?NMR(CDCl 3+CD 3OD)δppm：1.18(t，3H，J＝7.2Hz)，2.99，(q，2H，J＝7.1Hz)，7.95(d，2H，J＝8.4Hz)，8.08(d，2H，J＝8.8Hz)
13C?NMR(CDCl 3)δppm：7.9，32.1，127.7，130.0，134.0，140.0，168.0，200.8
Step c) 4-propionyl methyl benzoate
With above phenylformic acid (890mg, 5mmol), NaHCO 3(1.26g, 15mmol) and methyl iodide (935L, DMF 15mmol) (10mL) solution is in stirred overnight at room temperature.Mixture extracts with the saturated NaCl aqueous solution (50mL) dilution and with ether (3x 50mL).Organic phase water (50mL) washing, dry and evaporation.Flash chromatography separates that (90g silicon-dioxide, 2/1 sherwood oil-EtOAc) obtain white solid (744mg, 77%).
1H NMR (CDCl 3) δ ppm:1.24 (t, 3H, J=7Hz), 3.03 (q, 2H, J=7Hz), 3.95 (s, 3H), 8.0 and 8.12 (ABq, 4H)
Step d) 4-(2-bromine propionyl) methyl benzoate
Figure BPA00001375069700241
Under nitrogen in 50 ℃ with 4-propionyl phenylformic acid Methyl esters(744mg, 3.87mmol), pyrrolidone tribromide hydrogen salt (hydrotribromide) (1.98g) and 2-Pyrrolidone (380mg, THF 4.5mmol) (38mL) solution heating 3 hours.With mixture cooling, filtration, concentrated, be dissolved in again then in the ether (50mL).With diethyl ether solution water successively (20mL), saturated Na 2S 2O 5The aqueous solution (20mL), the saturated NaCl aqueous solution (20mL) and water (20mL) washing, dry and evaporation obtains yellow oil (1.025g), and the latter is directly used in the Hantzsch coupling.Measure according to 1H NMR, this raw material contains 91% required bromoketone, 5% starting ketone and 4%4-bromo-1-butanols.
1H NMR (CDCl 3) δ ppm:1.92 (d, 3H, J=7Hz), 3.96 (s, 3H), 5.28 (q, 1H, J=7Hz), 8.07 and 8.14 (ABq, 4H)
Step e) 4-[2-(4-tert-butoxycarbonyl piperazine-1-yl)-5-methylthiazol-4-yl] the benzene first The acid methyl esters
Figure BPA00001375069700242
At N 2Down in 70 ℃ will be above α-bromoketone and 4-thiocarbonyl piperazine-1-carboxylic acid tert-butyl ester (J.Med.Chem., 1998,5037-5054,917mg 3.73mmol) all refluxed 2 hours in 36mLTHF.Throw out is filtered, and the filtrate evaporation is obtained yellow solid.Flash column chromatography separates that (silicon-dioxide, 5/1 sherwood oil-EtOAc) obtain the 624mg faint yellow solid.(silicon-dioxide, 2/1 sherwood oil-EtOAc) obtain more polyvoltine compound of 32mg to throw out through chromatographic separation.Overall yield is 44%.
1H NMR (CDCl 3) δ ppm:1.46 (s, 9H), 2.43 (s, 3H), 3.42, (m, 4H), 3.54 (m, 4H), 3.90 (s, 3H), 7.68 and 8.04 (ABq, 4H).
Step f) 4-[2-(4-tert-butoxycarbonyl piperazine-1-yl)-5-methylthiazol-4-yl] phenylformic acid
Figure BPA00001375069700251
With above methyl esters (564mg, 1.35mmol) with 1.35mL 2N NaOH, 5mL THF and 3.65mL water in 60 ℃ of heating 4 hours.Reaction mixture is evaporated, pours into the saturated NaCl aqueous solution of 20mL and 20mL CH 2Cl 2In, then in ice bath with 5% citric acid acidifying to pH 3.2x10mL CH is also further used in each layer separation 2Cl 2Extracted organic phase.Organic phase is merged, water (10mL) washing, dry and evaporation obtains faint yellow solid (537mg, 98%).
1H NMR (CDCl 3) δ ppm:1.48 (s, 9H), 2.47 (s, 3H), 3.47 (m, 4H), 3.57 (m, 4H), 7.74 and 8.12 (ABq, 4H).
13C?NMR(CDCl 3)δppm：12.6，28.3，42.8，48.1，80.3，119.1，127.8，128.2，130.1，140.5，145.6，154.6，167.2，171.4.
LCMS：(M+H) +404，(M-H) -402.
Step g) 4-[5-methyl-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl] phenylformic acid
Figure BPA00001375069700252
With 4-[4-(4-carboxyl-phenyl)-5-methyl-thiazol-2-yl]-piperazine-1-carboxylic acid tert-butyl ester (0.421mmol) is dissolved in 1 of 4M HCl, and 4-two
Figure BPA00001375069700253
In the alkane solution, and in stirring at room 1 hour.Under vacuum, remove then and desolvate, resistates 4-(5-methyl-2-piperazine-1-base-thiazole-4-yl)-phenylformic acid is suspended in the methyl alcohol (10ml), and with the AcOH/AcONa damping fluid (pH～5.5,5ml) and formaldehyde (0.547mmol) processing.Reaction mixture in stirring at room 1 hour, is used NaCNBH then 3(0.547mmol) handle and in stirred overnight at room temperature.Then under vacuum, remove and desolvate, and resistates is carried out purifying, obtain title compound (0.403mmol, 95%) through column chromatography.
MS(ES)m/z?318(100％，[M+H] +).
Reference example 2
Another kind of P3 structural unit
3-fluoro-4-[2-(4-methylpiperazine-1-yl)-thiazole-4-yl] the benzoate hydrochlorate
Figure BPA00001375069700261
Step a) 4-bromo-3-fluorophenyl carbamate
(2.46g 11.2mmol) is dissolved in MeOH (9mL) and the toluene (4mL) and cools off in ice bath with 4-bromo-3-fluorobenzoic acid.(11mL, 2.0M are dissolved in the hexane, 22mmol) continue to exist up to yellow dropwise to add (trimethyl silyl) diazomethane.Solution in stirring at room 40 minutes, is concentrated then in a vacuum.Second batch of carboxylic acid (2.43g) handled equally.Merging is from two batches crude product and carry out flash chromatography and separate that (silicon-dioxide, 5/1 pentane-EtOAc) obtain methyl esters is white solid (4.92g, productive rate 95%).
1H?NMR(400MHz，CDCl 3)δppm 7.77(m，1H)，7.71(m，1H)，7.64(m，1H)，3.93(s，3H).
Step b) 4-acetoxy-3-fluorophenyl carbamate
Under rare gas element, to zinc powder (480mg, 7.34mmol) and anhydrous cobaltous bromide (II) (96.6mg, 0.44mmol) be dissolved in add in the suspension of MeCN (4mL) chlorallylene (105 μ L, 1.28mmol) and TFA (20 μ L, 0.26mmol).After stirring at room 10 minutes, add aryl bromide (1.003g, 4.30mmol are dissolved among the 5mL MeCN) from (a), (0.45mL 4.79mmol) also adds MeCN (1mL) more then to add diacetyl oxide.The mixture stirring is spent the night,, use EtOAc (3x20mL) extraction then with 1M HCl (20mL) cancellation.Organic phase is used saturated NaHCO successively 3The aqueous solution (20mL) and saturated NaCl (2x20mL) washing, dry (Na 2SO 4) and concentrate.Flash chromatography separates (silicon-dioxide, 6/1 to 4/1 sherwood oil-EtOAc bromide that obtains reclaiming (161.1mg, 16%) and required ketone (white solid, 305.5mg, 36%).
NMR(CDCl 3)δppm： 1H(400MHz)7.94-7.86(m，2H)，7.80(dd，1H，J＝11.2，1.6Hz)，3.95(s，3H)，2.67(d，3H，J＝4.4Hz)； 19F(376MHz)-109.2(m)； 13C(100MHz)195.4(d，J＝3.7Hz)，165.1(d，J＝2.2Hz)，161.6(d，J＝255Hz)，135.8(d，J＝8.1Hz)，130.7(d，J＝2.9Hz)，129.0(d，J＝14Hz)，125.2(d，J＝3.6Hz)，117.9(d，J＝26Hz)，52.7(s)，31.4(d，J＝7.3Hz).
Step c) 4-(2-acetobrom oxygen base)-3-fluorophenyl carbamate
To from b) ketone (198mg, 1.01mmol) with pyrrolidone tribromide hydrogen salt (532mg, add in mixture 1.07mmol) THF (10mL) and 2-Pyrrolidone (91 μ L, 1.20mmol).After 60-65 ℃ of heating 2 hours, mixture is concentrated under vacuum then at EtOAc (20mL) and saturated Na 2S 2O 3Distribute (10mL).Water extracts with EtOAc (10mL).Merge organic phase, with saturated NaCl (2x10mL) washing, dry (Na 2SO 4) and concentrate.Flash chromatography separate (silicon-dioxide, 7/1 sherwood oil-EtOAc), obtain white solid (0.2634g), its contain 84% required bromide (according to 19The integration at FNMR peak is determined).
NMR(CDCl 3)δppm： 1H(400MHz)7.93(m，1H)，7.88(m，1H)，7.79(dd，1H，J＝11.2，1.6Hz)，4.50(d，2H，J＝2.4Hz)，3.94(s，3H)； 19F(376MHz)-108.4(m).
Step d) 3-fluoro-4-[2-(4-methylpiperazine-1-yl)-thiazole-4-yl] methyl benzoate
To above bromoketone (193mg, 0.70mmol) and 4-methyl-piperazine-1-thioformamide (113mg, add EtOH (5.0mL) in 0.71mmol) and in 70 ℃ with mixture heating up 2 hours 15 minutes.Throw out is filtered, wash and dry and sign under vacuum with cold EtOH.This process is repeated with the bigger scale of 1.75g bromoketone (6.36mmol).
NMR(1/1?CDCl 3-CD 3OD)δppm： 1H(400MHz)8.20(m，1H)，7.86(dd，1H，J＝8.4，1.6Hz)，7.76(dd，1H，J＝11.4，1.8Hz)，7.38(d，1H，J＝2.4Hz)，4.23(br，2H)，3.95，(s，3H)，3.65(br，4H)，3.32(br，2H)，2.98(s，3H)； 19F(376MHz)-114.0(m).LCMS[M+H] +＝336.
Merging is from the throw out of twice preparation and be suspended in saturated NaHCO 3(50mL).Mixture is extracted with EtOAc.Wash organic phase with water, dry (Na 2SO 4), and the evaporation obtain title compound, be paste solid (1.76g).
Step e) 3-fluoro-4-[2-(4-methylpiperazine-1-yl)-thiazole-4-yl] the benzoate hydrochlorate
Will (1.76g be 5.25mmol) with 6M HCl (40mL) heating 5.5 hours from the methyl esters of (d) in 80 ℃.Add 6M HCl (10mL) again and in 90 ℃ of heated mixt 1 hour 15 minutes.After the cooling,, obtain end product, be the paste solid of quantitative yield then with mixture vaporising under vacuum and lyophilize from water.
NMR(DMSO-d6)δppm： 1H(400MHz)11.60(br，1H)，8.18(t，1H，J＝8.0Hz)，7.82(dd，1H，J＝8.4，1.6Hz)，7.72(dd，1H，J＝12.0，1.6Hz)，7.48(d，1H，J＝2.8Hz)，4.11(m，2H)，3.58(m，2H)，3.49(m，2H)，3.19(m，2H)，2.80(d，3H，J＝4.4Hz)； 19F(376MHz)-113.5(m)； 13C(100MHz)168.9，166.0，159.0(d，J＝250Hz)，143.4，131.4(d，J＝8Hz)，129.8，125.8(d，J＝11Hz)，125.6，116.6(d，J＝24Hz)，111.1(J＝15Hz)，51.1，45.0，41.9.LCMS[M+H] +＝322.
Reference example 3
6-aldehyde-intermediate
Figure BPA00001375069700291
6-formyl radical-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester
Step a
(3as, 6aS)-6R-benzyloxy-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid benzyl ester
In 45 minutes, (12.5g 30mmol) is dissolved among the DCM (250mL) with Dai Si-Martin's reagent.Under nitrogen environment, in the oxidizing agent solution that stirs, add 6-benzyloxy-3-hydroxyl-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid benzyl ester (preparing) (7.4g, DCM 20mmol) (50mL) solution by describing among the WO05/066180 in room temperature.After the restir 90 minutes, think according to TLC and to finish reaction.Add 10%Na 2S 2O 3The aqueous solution (200mL) and in room temperature with mixture restir 15 minutes.Biphasic system is transferred to separating funnel also with EtOAc extracting twice (being respectively 200mL and 100mL).The saturated NaHCO of organic phase that merges 3The aqueous solution (100mL) and salt solution (100mL) washing are once used Na 2SO 4Drying is filtered and solvent is removed in a vacuum, produces the crude product title compound, is transparent oily matter (7.69g); ESI+, m/z:368 (M++1).
Step b
(3aS, 6aS)-6R-benzyloxy-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-benzyl carboxylate The base ester
The ketone group derivative (7.6g) of step a) is dissolved in the dry methyl alcohol (100mL).Under nitrogen environment, add trimethyl orthoformate (30mL) and pTsOH (0.2g) in room temperature.Mixture was heated 8 hours in 60 ℃.When thinking that according to TLC reaction has been finished, it is cooled to room temperature and concentrated in a vacuum.Crude product through the silica gel column chromatography purifying, is obtained title compound with ethyl acetate-heptane (1: 4) wash-out, be transparent oily matter (5.9g is through 2 steps, 71%); ESI+, and m/z:382 (M+-OMe).
Step c
Figure BPA00001375069700302
(3aS, 6aS)-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-6R-alcohol
Compound (2.5g, methyl alcohol 6.4mmol) (60mL) and Pd (OH) with step b) 2(0.7g) solution in room temperature at H 2Environment stirred 48 hours down.When thinking that according to TLC reaction is finished, mixture is filtered and concentrates in a vacuum, obtain thick title compound, be brown oil (1.15g); ESI+, m/z:190 (M++1).
Steps d
(3aS, 6aS)-6R-hydroxyl-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid uncle fourth Ester
Will be from 3 of step c), (2.80g 14.8mmol) is dissolved in 75mL two to 3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-6-alcohol
Figure BPA00001375069700312
In alkane/water (2: 1) mixture.Dropwise add 10%Na 2CO 3Solution (25mL) is to pH 9-9.5.Mixture is cooled to 0 ℃ and add the Boc acid anhydrides with portion in ice-water-bath.Reactant is in stirred overnight at room temperature, if necessary by adding 10%Na again 2CO 3Solution makes the pH of mixture remain on 9-9.5.According to TLC (50: 50 ethyl acetate: isohexane) come monitoring reaction.In case finish, with regard to the salt of filtering mixt eliminate to form, and evaporating solvent in a vacuum.Aqueous mixture extracts with 3x100mL EtOAc, and the organic phase of merging is used Na with 100mL water and the water washing of 100mL salt 2SO 4Drying is filtered also evaporating solvent in a vacuum, obtains 3.79g title carbamate, is transparent oily matter (89%) purity 94% (HPLC), ESI+, m/z:312 (M++Na).
Step e
Figure BPA00001375069700313
3,3-dimethoxy-6-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester
To the alcohol that is dissolved in DCM (80mL) from step e) (3.674g, add in 12.70mmol) Dai Si-Martin's oxygenant (7.00g, 16.5mmol) and with solution in stirring at room 3 hours.Then by adding 10%Na 2S 2O 3(aq) (150mL) make reaction cancellation and the slurries that generate were stirred 15 minutes.Mixture is transferred in the separating funnel, and will respectively be separated.Water is used saturated NaHCO subsequently with DCM extraction three times 3Solution is with the organic phase washed twice that merges, and is dry then, filters and concentrate.Thick material obtains title compound (2.882g, 79%) through flash column chromatography (toluene/ethyl acetate 3: 1) purifying.
Step f
Figure BPA00001375069700321
3,3-dimethoxy-6-methylene radical-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester
Will (1.10g 3.83mmol) be dissolved in the dry THF (30mL) and with solution and be cooled to 0 ℃ from the keto compounds of step e.The Diethylaminoethyl triphenylphosphine that added the branch that classifies in three categories every 2 hours
Figure BPA00001375069700322
(4.0g is 11.2mmol) with KOtBu (1.17g, dry THF 10.5mmol) (40mL) solution.After 6 hours, pour into solution and diethyl ether (70mL) in the separating funnel and (2*40mL) extract with 10% citric acid (aq).The saturated NaHCO of organic phase 3Na is used in the aqueous solution (40mL) washing 2SO 4Drying is filtered and solvent is evaporated in a vacuum.(heptane: ethyl acetate 4: 1) purifying obtains title compound (524mg, 48%) to crude product through the flash chromatography separation
1H?NMR(CDCl 3，400MHz)δ1.48(s，9H)，3.27(s，3H)，3.40(d，3H，J＝16.6)，3.57-3.64(m，1H)，3.84(d，1H，J＝9.5)，3.92(d，1H，J＝16.3)，4.07-4.25(m，1H)，4.35-4.49(m，1H)，4.98(bs，1H)，5.22(d，1H，J＝16.4)，5.34(s，1H).
Step g
Figure BPA00001375069700323
6-methylol-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester
Will (524mg 1.84mmol) be dissolved in the dry THF (70mL) from the alkene of step f).(7.34mL 3.67mmol) and with solution stirring spends the night to add 9-BBN-H (0.5M is in THF).Solvent is removed by rotary evaporation and is dissolved in again among the THF (20mL).In solution, slowly add MeOH (10mL), when gas evolution stops, in solution, adding H 2O (20mL) then adds NaBO 3With solution stirring filtration afterwards in 18 hours and with EtOAc (70mL) dilution filtrate, use salt solution (2*50mL) wash filtrate again.Organic phase Na 2SO 4Drying is filtered also evaporating solvent in a vacuum.(heptane: ethyl acetate 2: 1) purifying obtains title compound (477mg, 86%) to crude product by flash chromatography.
1H?NMR(CDCl 3，400MHz)δ1.47(s，9H)，2.09-2.25(m，2H)，3.02-3.20(m，1H)，3.29(s，3H)，3.39(s，3H)，3.65-3.93(m，4H)，4.44(d，1H，J＝5.7)，4.70-4.84(m，1H).
Step h
Figure BPA00001375069700331
6-formyl radical-3,3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester
To the 1g alcohol of the step g) that is dissolved in dry DCM (10mL) (370mg, add in solution 1.22mmol) Dai Si-Martin's oxygenant (673mg, 1.59mmol).Reactant was stirred 40 minutes, then by adding 10mL 10%Na 2S 2O 3: NaHCO 3(saturated) carries out cancellation at 1: 1.Described solution diluted with DCM (50mL) and use 10%Na 2S 2O 3: NaHCO 3(saturated) is 1: 1 mixture (50mL) extraction.Organic phase Na 2SO 4Drying is filtered and evaporation.(heptane: ethyl acetate (2: 1) purifying obtains title compound (290mg, 79%) through flash chromatography with crude product.
1H NMR (CDCl 3, 400MHz) δ 1.47 (s, 9H), 2.90-3.06 (m, 1H), 3.29 (s, 3H), 3.38 (s, 3H), 3.67-3.85 (m, 2H), 3.88-4.55 (m, 3H), 4.93-5.19 (m, 1H), 9.64 *With 9.80 *(s, 1H). *Two peaks are by due to the rotational isomer.
Reference example 4
6-nitrile P1 structural unit
Figure BPA00001375069700341
Step a)
Figure BPA00001375069700342
515mg (1.71mmol) 6-formyl radical-3 from reference example 3; 3-dimethoxy-six hydrogen-furo [3,2-b] pyrroles-4-carboxylic acid tert-butyl ester, 130.6mg (1.88mmol) oxammonium hydrochloride and 168mg (2.05mmol) sodium acetate in ethanol-water mixture (10/15ml) in stirred overnight at room temperature.Distribute with the mixture evaporation and between water and ethyl acetate phase.Dried over sodium sulfate is used in organic phase salt water washing, and evaporation is also gone up purifying at silicon-dioxide (EtOAc-hexane 1: 1), obtains the mixture of above-described cis-isomeride and trans-isomer(ide).R f0.43 and 0.48 (EtOAc-hexane 1: 1).Output 395mg (73%)
Figure BPA00001375069700343
With the oxime of step a) (114mg, 0.36mmol) with triethylamine (105 μ L 0.757mmol) are dissolved in 1, in the 5ml methylene dichloride and cooling up to-78 ℃.In 7 minutes, dropwise add the trifluoromethanesulfanhydride anhydride be dissolved in the 600 μ L methylene dichloride (61. μ L, 0.36mmol).Make reaction mixture warm up to room temperature and stirred 2 hours.Mixture with refrigerative 5% citric acid, sodium bicarbonate, salt water washing in advance, use dried over sodium sulfate with 15ml DCM dilution, evaporation and at silicon-dioxide (gradient EtOAc-hexane 1: 3-1: 1) last purifying.R f(0.65 EtOAc-hexane 1: 1).Output 65mg (61%)
Reference example 5
Typical P1/P2 deprotection and coupling
Figure BPA00001375069700351
(95mg 0.032mmol) is dissolved in the 5ml methyl alcohol that is cooled to 0 ℃ and dropwise adds the 0.5ml Acetyl Chloride 98Min. with the nitrile structural unit of reference example 4.With the mixture that generates in stirring at room 4 hours, evaporation then.Resistates is dissolved among the 2.5ml DMF, adds 80mg (0.32mmol) Boc-Leu-OH, then add the 0.5ml diisopropylethylamine.With the mixture cooling that generates up to 0 ℃ and add 160mg (0.42mmol) HATU (phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-N, N, N ', N '-tetramethyl-urea
Figure BPA00001375069700352
).Make reaction mixture warm up to room temperature and stirred 1.5 hours.With mixture evaporation, distribute between mutually in water and ethyl acetate, water, salt water washing organic phase are used dried over sodium sulfate, evaporate and on silicon-dioxide (EtOAc Rf 0.35) purifying, obtain the 82mg title compound.For two steps, output is 82mg (62%).
Reference example 6
Another kind of P3 structural unit
Figure BPA00001375069700361
I.AcOH, bromine, RT, 2h, productive rate 55%; Ii.KF, acetonitrile ,-6,90 ℃ of 18-hats, 16h; Productive rate 31%; Iii.AcOH, bromine, 45 ℃, 4h, productive rate 100%; Iv.4-methyl-piperazine-1-thioformamide, heating, 2h, productive rate 74%; LiOH, RT, 16h, productive rate 100%.
The availability of raw material-
4-acetylbenzoic acid methyl esters can obtain from Aldrich; 11 suppliers (perhaps Chem Pur ProductsLtd is at German most convenient) of 4-methyl-piperazine-1-thioformamide-in SciFinder, find.
Step a) 4-(2-bromo-ethanoyl)-methyl benzoate
Figure BPA00001375069700362
In acetate (20mL) solution of 4-ethanoyl-methyl benzoate (8.4mmol), add bromine (8.4mmol).Stirring at room 2 hours, during this period of time red the disappearance also formed pale precipitation with reactant.Product is collected after filtration and with cold methanol/water (200mL 1: 1) washing, is obtained white powder (55%).1H?NMR(400MHz，CDCl 3)3.98(3H，s)，4.20(2H，s)，8.02(2H，d，J＝8Hz)，8.18(2H，d，J＝8Hz)。
Step b) 4-(2-fluoro-ethanoyl)-methyl benzoate
Adding 18-is preced with-6 (0.1mmol) and reactant was heated 30 minutes in 90 ℃ in acetonitrile (1mL) suspension of Potassium monofluoride (3.11mmol).Add 4-(2-bromo-ethanoyl)-phenylformic acid (1.56mmol) and with reactant in 90 ℃ of heating 16 hours.Reactant water (10mL) dilutes and (3 * 20mL) extract with ethyl acetate.Product is purifying on silicon-dioxide, with the 5-15% eluent ethyl acetate that is dissolved in isohexane, concentrates required component in a vacuum, obtains title compound, is white solid (31%).1H?NMR(400MHz，CDCl 3)3.98(3H，s)，5.55(2H，d，J＝50Hz)，7.95(2H，d，J＝8Hz)，8.18(2H，d，J＝8Hz).
Step c) 4-(2-bromo-2-fluoro-ethanoyl)-methyl benzoate
In acetate (5mL) suspension of 4-(2-fluoro-ethanoyl)-phenylformic acid (1.19mmol), add bromine (1.19mmol).Reactant in 45 ℃ of heating 4 hours, is during this period of time formed green solution.Reactant is concentrated and twice and methylbenzene azeotropic generation title compound in a vacuum, be green solid (100%).In next step, use this crude product.1H?NMR(400MHz，CDCl 3)3.98(3H，s)，7.04(1H，s)，8.05-8.10(4H，m).
Step d) 4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-methyl benzoate
Figure BPA00001375069700373
4-(2-bromo-2-fluoro-ethanoyl)-methyl benzoate (1.18mmol) and 4-methyl-piperazine-1-thioformamide (1.18mmol) are dissolved in the ethanol (10mL).Reactant was heated 2 hours under refluxing.Reactant is cooled to room temperature, causes the product precipitation.Collect product after filtration and use cold washing with alcohol.Give the aftertreatment of product aqueous carbonic acid hydrogen sodium, produce title compound, be colorless oil (74%) .MS (ES+) 337 (M+H, 100%).
Step f)
4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-the phenylformic acid dihydrochloride
Figure BPA00001375069700381
To 4-[5-fluoro-2-(4-methyl-piperazine-1-yl)-thiazole-4-yl]-add lithium hydroxide (0.5mmol) in tetrahydrofuran (THF)/water (2.5mL, 4: 1) solution of methyl benzoate (0.43mmol).With reactant stirring at room 16 hours.Reactant concentrates in a vacuum and adds hydrochloric acid (2N 3mL) causes the product precipitation, is white solid.Collect product after filtration and obtain title compound, be white solid (79%).MS(ES+)322(M+H，100％).
Embodiment 1
N-[1-(6-cyano group-3-oxo-six hydrogen-furo [3,2-b] pyrroles-4-carbonyl)-3-methyl-Ding Base]-4-[2-(4-methyl-piperazine-1-yl) thiazole-4-yl]-benzamide
Step a)
Figure BPA00001375069700391
(60mg 0.147mmol) is dissolved in the 3ml methyl alcohol, is cooled to 0 ℃ and dropwise add the 0.4mL Acetyl Chloride 98Min. with the P1/P2 structural unit of reference example 5.The mixture that produces in stirring at room 4 hours, evaporation then.Resistates is dissolved among the 2.5mL DMF, adds the acid of 52mg (0.147mmol) P3 structural unit (by preparing among the WO0566180), then add the 0.5mL diisopropylethylamine.With the mixture cooling that produces up to 0 ℃ and add 70mg (0.184mmol) HATU (phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-N, N, N ', N '-tetramethyl-urea ).Make reaction mixture warm up to room temperature and stirred 1.5 hours.With mixture evaporation, distribute between mutually in water and ethyl acetate, and water, salt water washing organic phase, use dried over sodium sulfate, evaporation and at the last purifying of silicon-dioxide (the EtOAc solution of 5%MeOH).Merge required component and concentrate in a vacuum and obtain material requested, output is 56mg (64%), LC/MS 597 (M+1)
Step b)
Figure BPA00001375069700393
With 3mL TFA-water mixture (2.5% is water-soluble in TFA) treatment step ketal (56mg, 0.094mmol) 4 hours a).Reaction is monitored by LC/MS.With the reaction mixture evaporation, be dissolved in the acetonitrile (5mL), stirred 1 hour with solid sodium carbonate, then solid filtering is gone out, mother liquor concentrates in a vacuum, and through preparation HPLC (NH 4OAc damping fluid, 30-80 system (MeCN-water)) purifying obtains the required product of 25mg (productive rate 45%).LC/MS M+1 551, M+19 569(hydrate forms)
Embodiment 2
Figure BPA00001375069700401
(33mg 0.08mmol) is dissolved in and is cooled to 0 ℃ in the 3ml methyl alcohol, and dropwise adds the 0.4ml Acetyl Chloride 98Min. and add acid with the P1-P2 structural unit (1) of protected reference example 5.In stirring at room gained mixture 4 hours, evaporation then.Resistates is dissolved among the 2.5mlDMF, adds the P3 acid (being hydrochloride) of 29mg (0.08mmol) reference example 2, then add the 0.5ml diisopropylethylamine.With the mixture cooling that produces up to 0 ℃ and add 39mg (0.101mmol) HATU (phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-N, N, N ', N '-tetramethyl-urea
Figure BPA00001375069700402
).Make reaction mixture warm up to room temperature and stirred 1.5 hours.Make mixture evaporation, distribute between mutually in water and ethyl acetate, dried over sodium sulfate use in organic phase water, salt water washing, evaporation and at the last purifying of silicon-dioxide (5%MeOH is dissolved among the EtOAc).(3) output is 32mg (67%), LC/MS 615 (M+1)
Handle ketal (3) (32mg, 0.054mmol) 4 hours with 3ml TFA-water mixture (2.5% is water-soluble in TFA).Reaction is monitored by LC/MS.With reaction mixture evaporation, be dissolved in the acetonitrile (5ml), stirred 1 hour with solid sodium carbonate, then solid filtering is gone out, mother liquor concentrates in a vacuum, and on preparation type LC/MS purifying, by preparation HPLC (NH 4The OAc damping fluid, 30_80 system (MeCN-water)) purifying, obtain 13mg product (productive rate 42%).LC/MS M+1 569, M+19 587(hydrate forms)
Embodiment 3
Figure BPA00001375069700411
(33mg 0.08mmol) is dissolved in and is cooled to 0 ℃ and dropwise add the 0.4ml Acetyl Chloride 98Min. in the 3ml methyl alcohol, adds acid with the P1P2 structural unit (1) of protected reference example 5.The mixture that produces in stirring at room 4 hours, evaporation then.Resistates is dissolved among the 2.5mlDMF, adds the P3 acid (being hydrochloride) of 29mg (0.08mmol) reference example 6, then add the 0.5ml diisopropylethylamine.With the mixture cooling that produces up to 0 ℃ and add 39mg (0.101mmol) HATU (phosphofluoric acid O-(7-azepine benzo triazol-1-yl)-N, N, N ', N '-tetramethyl-urea
Figure BPA00001375069700412
).Make reaction mixture warm up to room temperature and stirred 1.5 hours.Mixture evaporation distributes between mutually in water and ethyl acetate, and dried over sodium sulfate is used in organic phase water, salt water washing, evaporates and purifying on silicon-dioxide (5%MeOH is dissolved in EtOAc).(5) output is 30mg (63%), LC/MS 615 (M+1)
Handle ketal (5) (30mg, 0.051mmol) 4 hours with 3ml TFA-water mixture (2.5% is water-soluble in TFA).Reaction is monitored by LC/MS.With the reaction mixture evaporation, be dissolved in the acetonitrile (5ml), stirred 1 hour with solid sodium carbonate, then solid filtering is gone out, mother liquor concentrates in a vacuum, and purifying on preparation type LC/MS is by preparation HPLC (NH 4The OAc damping fluid, 30_80 system (MeCN-water)) purifying, obtain 11mg product (productive rate 38%).LC/MS M+1 569, M+19 587(hydrate forms)
Biological Examples
The mensuration of cathepsin K proteolysis catalytic activity
The suitable analysis of cathepsin K uses the human recombinant enzyme of describing among the PDB for example to carry out.
ID BC016058 standard; MRNA; HUM; 1699BP.
DE human tissue Proteinase K (pycnodysostosis), (cDNA clones MGC:23107 to mRNA
RX?MEDLINE；.RX PUBMED； 12477932.
DR?RZPD； IRALp962G1234.
DR?SWISS-PROT； P43235；
The recombinant tissue Proteinase K can comprise in intestinal bacteria, pichia spp (Pichia) and the rhabdovirus system at various commercially available expression systems expresses.Purifying enzyme by ordinary method through removal and leader sequence activates.
Measure the standard analysis condition of kinetic constant and use fluorogenic substrate, be generally H-D-Ala-Leu-Lys-AMC, and at the 100mM Mes/Tris that contains 1mM EDTA and 10mM 2 mercapto ethanol, pH 7.0, or 100mM sodium phosphate, 1mM EDTA, 0.1%PEG4000 pH 6.5, or contain the 100mM sodium acetate of 5mM EDTA and 20mM halfcystine, measure among the pH 5.5, optional in all cases with 1M DTT as stablizer.Employed enzyme concn is 5nM.Substrate storage liquid prepares with 10mM in DMSO.Screening is carried out under immobilized substrate concentration 60 μ M, and dynamics research is used from the substrate doubling dilution of 250 μ M and carried out in detail.Total DMSO concentration in the analysis keeps below 3%.All analyses are carried out all at ambient temperature.Product fluorescence (exciting at 390nm, in the 460nm emission) is read the plate instrument with Labsystems Fluoroskan Ascent fluorescence and is monitored.After producing the AMC product, in 15 minutes, produce the product curve of progress.
Cathepsin S Ki measures
This analyzes the human tissue proteolytic enzyme S of use baculovirus expression and the boc-Val-Leu-Lys-AMC fluorogenic substrate from the Bachem acquisition in 384 orifice plates, can carry out parallel test to 7 kinds of test compounds and the positive control that comprises known tissue proteolytic enzyme S inhibitor contrast in orifice plate.
The substrate dilution
The 12.5%DMSO that adds 280 μ l/ holes to the two capable B-H that are listed as of 96 deep hole polypropylene boards.The substrate that adds 70 μ l/ holes to row A.Analysis buffer (pH 6.5 for 100mM sodium phosphate, 100mM NaCl) to row A adding 2x250 μ l/ hole is mixed, and along the downward double capable H that is diluted to of plate.
The inhibitor dilution
Row 2-5 in 4 row of polypropylene board at the bottom of the 96 hole V and row 7-12 add the analysis buffer in 100 μ l/ holes.The analysis buffer that adds 200 μ l/ holes to row 1 and row 6.
Row 1 to first trip are incorporated in first test compound for preparing among the DMSO, and its capacity provides the rough K of initial mensuration usually i10 to 30 times between.Rough K iCalculate according to the prerun operameter, wherein the 1mM boc-VLK-AMC in 10 μ l/ holes (1/10 diluent that the DMSO storage liquid of 10mM dilutes in analysis buffer) is dispensed to 96 hole Microfluor TMThe capable B to H of plate is dispensed to capable A with 20 μ l/ holes.Add every 10mM test compound 2 μ l in the independent hole on the row 1-10 of row A.Adding 90 μ l to each hole of row B-H contains the analysis buffer of 1mM DTT and 2nM cathepsin S and adds 180 μ l to row A.Use the multiple tracks transfer pipet to mix row A and doublely be diluted to capable G.Mix row H and reading in spectrophotofluorometer.Reading is the Prism data, its match the competitive inhibition equation, set S=100 μ M and K M=100 μ M are to obtain K iEstimated value, maximum nearly 100 μ M.
Row 6 to first row add second kind of test compound, add the third test compound etc. to second row of going 1.Row 6 to row of the end add 1 μ l contrast.Mix row 1 and the double row 5 that are diluted to.Mix row 6 and the double row 10 that are diluted to.
Use is set to the multistage transfer pipet in 8 roads of 5x10 μ l, and the substrate in 10 μ l/ holes is dispensed on the 384 hole analysis plates.First row that substrate is diluted plate are dispensed to all that originate in capable A on the analysis plates and list.The suction nozzle of multiple tracks transfer pipet (tip) spacing will correctly be skipped staggered rows.Secondary series is dispensed to all row that start from row B.
Use is set to the multistage transfer pipet in 12 roads of 4x10 μ l, and the inhibitor in 10 μ l/ holes is dispensed on the 384 hole analysis plates.First row that inhibitor is diluted plate is dispensed to the staggered rows that starts from A1 on the analysis plates.The suction nozzle spacing of multiple tracks transfer pipet will correctly be skipped staggered row.Similarly, second row, the third line and fourth line are dispensed to staggered rows and the row that start from A2, B1 and B2 respectively.
Mix 20ml analysis buffer and 20 μ l 1M DTT.The cathepsin S that adds capacity is to obtain the ultimate density of 2nM.
Use for example Multidrop 384 of divider, to the institute porose adding 30 μ l/ holes of analysis plates and with spectrophotofluorometer Ascent reading for example.
The enzyme of fluorescence reading (excitation wavelength and emission wavelength are respectively 390nm and 460nm, use the bandpass filter setting) reflection fluorogenic substrate is cut degree, although inhibitor is the linear velocity in each hole after the match.
Use SigmaPlot 2000,, measure V, Km and Ki value every kind of inhibitor institute foraminous match speed match competitive inhibition equation.
The Ki of cathepsin L
Use program above and make following revision, measured the Ki of cathepsin L.
Described enzyme is commercially available human tissue proteolytic enzyme L (for example Calbiochem).Substrate is the H-D-Val-Leu-Lys-AMC that obtains from Bahcem.Analysis buffer is 100mM sodium acetate 1mM EDTA, pH5.5).DMSO storage liquid (10mM is in 100%DMSO) is diluted to 10% in analysis buffer.Before facing usefulness, enzyme is prepared as 5nM concentration and adds the 1mM dithiothreitol (DTT) in analysis buffer.The 2ul 10mM inhibitor for preparing among the 100%DMSO is dispensed to capable A.10 μ l, 50 μ M substrates (1/200 diluent that=10mM DMSO storage liquid dilutes in analysis buffer)
Suppress research
Use above analysis and the test compound of variable concentrations, the screening potential inhibitor.Reaction adds enzyme by the buffered soln to substrate and inhibitor and starts.K iValue is calculated according to equation 1.
v 0 = VS K M ( 1 + I K i ) + S - - - ( 1 )
V wherein 0Be speed of response, V is a top speed, and S has Michaelis-Menton constant K MConcentration of substrate, and I is the concentration of inhibitor.
The following expression of result
A is lower than 50nM
B?50-500nM
C?501-1000nM
D?1001-5000nM
E?5001-10000nM
F surpasses 10000nM
Table 1
Embodiment Test number The Ki cathepsin K The Ki cathepsin S Ki cathepsin L
1 Test 1 A D B
1 Test 2 3.0nM 3300nM 530nM
Therefore formula II compound is effective inhibitor of cathepsin K and still selective to closely-related cathepsin S and L.
Metabolic stability
In the cytosol analysis to The compounds of this invention and shown in Comparative Examples carried out the metabolic stability test, wherein said compound is hatched with commercially available human liver cytosol part, and the disappearance of compound is by HPLC or LC/MS monitoring.Compare with the blood from independent part, the human liver cytosol part of merging is unlikely represented the individuality that peels off, and can freezingly preserve, unlike whole blood.Therefore the cytosol analysis provides consistent analyzing test bench as to compound environment in vivo, the guide of stability in the time of for example in being exposed to whole blood.
In brief, in 37 ℃ with test compound (2 μ M) the human liver cytosol that merges (Xenotech LLC Lenexa US, 1mg/mL protein, in the 0.1M phosphoric acid buffer in, pH 7.4) in hatched 1 hour.Hatch by adding the startup of 1mM NADPH cofactor.In the time of 0,20,40 and 60 minute, take out periodic inferior sample and it " is collided precipitation " by the ice-cold acetonitrile that adds 3 times of capacity.Sample is centrifugal under the temperature that reduces, and analyzes with the supernatant liquor separation and with LC-MS-MS.
Perhaps, in people or monkey whole blood and/or commercially available hepatomicrosome, carry out similar stability analysis.
Table 2
Figure BPA00001375069700461
Comparative Examples is represented in the scope of WO2008/007107 mentioned above at 6 compounds that C-C is arranged.It prepares from compound 1d (flow process 1) in an easy manner.Therefore use the outer alkene 1d of available ring, alkene in ethyl acetate, carries out the stereoselectivity hydrogenation of alkene with Adams catalyzer (platinum dioxide) by the cis hydrogenation under the hydrogen environment.This hydrogenation produces a kind of product basically, promptly has the stereochemical C-6 methyl of R-isomer (LCMS[M+H]=288 as seen), its productive rate height.For step of hydrogenation, herein before visible face selectivity and the document selectivity of the twin nuclei that is closely related of report similar (people such as Srinivas, Synlett, 1999,555-556).The structural unit of preparation like this is carried out deprotection, prolongs and is oxidized to about the active keto-acid of illustrative The compounds of this invention above.
The body internal stability improves tolerable compound whole day better distribution in vivo, no matter be QD or BID administration.This is for the illness that wherein is changed significantly round the clock osteoporosis particularly important for example.
Perviousness
This measuring inhibitor is by the transhipment of human gi-tract cell.Analyze and use the Caco-2 cell of passage number between 40 and 60 of knowing.
The top is to basic side transhipment
Common every kind of compound is tested with the 2-4 hole.Base side and vertical hole will be contained 1.5mL and 0.4mL transport buffer (TB) respectively, and the normal concentration of substances is 10 μ M.In addition, all testing liquid and damping fluids will contain 1%DMSO.Before experiment, with transport plates with the pre-bag of the substratum that contains 10% serum by 30 minutes to avoid the non-specific binding with plastic material.After cultivating 21 to 28 days on the filter membrane upholder, cell prepares to be used for the perviousness experiment.
Transport plates contains 3 row that 4 holes are respectively arranged for No. 1.Row 1 expression washing, row 2 expressions " 30 minutes ", and row 3 expressions " 60 minutes ".Transport plates contains 3 row that 4 holes are respectively arranged for No. 2, row 4 expressions " 90 minutes ", and row 5 expressions " 120 minutes ", and all the other delegation do not specify.
Remove from the substratum of topped hole and inset is transferred to washing row (No. 1) among 2 blocks of plates of no inset at transport plates (No. 1 plate), described 2 blocks of plates are expert at and have been used 1.5mL transport buffer (HBSS in 1 to 5,25mM HEPES, pH 7.4) preparation.In A → B screening, the TB in the basic side opening also contains 1% bovine serum albumin.
0.5mL transport buffer (pH 6.5 for HBSS, 25mM MES) is joined in the inset, and make cell monolayer in the polymix vibrator in 37 ℃ of balances 30 minutes in the transhipment buffering system.In balance to buffering system, by the every hole of EVOM chop stick Instrument measuring through the epithelium resistance value (Transepithelial electrical resistance, TEER).The TEER value is usually between every hole 400 to 1000 Ω (passage number that depends on use).
Remove transport buffer (TB, pH 6.5) from tip side, inset is transferred to 30 minutes row (No. 2), will comprise that the fresh 425 μ L TB (pH 6.5) of substances are added in the hole, top (donor).Plate is hatched in the polymix vibrator in 37 ℃, and its low vibration rate is approximately 150 to 300rpm.
Be expert at 2 hatched 30 minutes after, inset moved to new pre-hot radical side (recipient) hole in per 30 minutes; Row 3 (60 minutes), 4 (90 minutes) and 5 (120 minutes).
After about 2 minutes and experiment when finishing, from vertical solution, take out 25 μ L samples.These sample representatives are from experiment beginning and the donor sample that finishes.
Take out 300 μ L at each preset time point from basic side (recipient) hole, and when experiment finishes TEER value (post value) behind the determination experiment.Add acetonitrile to all samples of collecting, ultimate density is 50% in the sample.The sample of collecting is analyzed up to HPLC or LC-MS in-20 ℃ of storages.
The base side is to vertical transhipment
Common every kind of compound is used the test of 2-4 hole.Base side opening and topped hole will contain 1.55mL and 0.4mL TB respectively, and the normal concentration of substances is 10 μ M.In addition, all testing liquid and damping fluids will contain 1%DMSO.Before experiment with the pre-bag of the substratum that contains 10% serum by transport plates 30 minutes, to avoid the non-specific binding with plastic material.
After cultivating 21 to 28 days on the filter membrane upholder, cell prepares to be used for the perviousness experiment.Remove substratum and inset is transferred to washing row (No. 1) on the new plate (transport plates) of no inset from topped hole.
Transport plates comprises 3 row that 4 holes are respectively arranged.Row 1 expression " washing ", and row 3 is " an experiment row ".Before the transport plates No. 1, washing row with 1.5mL TB (pH 7.4) preparation, at experiment row No. 3 (donor sides) with 1.55mL TB (pH 7.4) preparation that comprises substances.
0.5mL transport buffer (pH 6.5 for HBSS, 25mM MES) is added in the inset of No. 1 row, and with cell monolayer in the polymix vibrator in 37 ℃ of balances 30 minutes in the transhipment buffering system.Balance is to buffering system, by the TEER value in each hole of EVOM chop stick Instrument measuring.
Transport buffer (TB, pH 6.5) is removed and inset is transferred to row 3 from tip side, and with the fresh TB of 400 μ L, pH 6.5 joins in the inset.Taking out 250 μ L after 30 minutes from vertical (recipient) hole also replaces with fresh transport buffer.After this took out 250 μ L samples in per 30 minutes and replace with fresh transport buffer that experiment finishes TEER value behind the determination experiment when the experiment end up to 120 minutes the time.Take out 25 μ L samples from basic side (donor) chamber after about 2 minutes and when experiment finishes.These sample representatives are from experiment beginning and the donor sample that finishes.
Add acetonitrile in the sample of all collections, ultimate density is 50% in the sample.The sample of collecting is analyzed up to HPLC or LC-MS-20 ℃ of storages.
Calculate
Accumulation absorbent components FA with respect to the time CumMensuration.FA CumCalculate according to following formula
FA cum = Σ C RI C DI
C wherein RiBe the recipient's concentration when i finishes at interval, and C DiBe the donor concentration when i begins at interval.Should obtain linear relationship.
Permeability coefficient (P App, measurement result cm/s) is calculated according to following formula:
P app = ( k · V R ) ( A · 60 )
Wherein k is transport velocity (min -1), as passing through accumulation absorbent components (FA Cum) slope that obtains with the linear regression of the function of time (min) defined V RBe the volume in the receiving chamber (mL), and A is the area (cm of filter membrane 2).
Reference compound
Figure BPA00001375069700493
Figure BPA00001375069700501
Perviousness by gastrointestinal tissue is stronger to be favourable, because its allows to use smaller dose to reach and the weaken similar level of compound of perviousness with more high dosage administration.Low dosage is favourable, because reduced the per daily dose cost of commodity, is vital parameter in the medicine that the latter absorbed in the time cycle that prolongs.
Mutagenicity
The test in Ames (Ames) test easily of the mutagenesis potentiality of compound, usually various bacterial isolateses for example among Salmonella typhimurium (Salmonella typhimurium) TA100, TA102, TA 1535, the TA 1537, having and do not having and for example under concentration 30,300 and 3000ug/ plate, carrying out under the liver S9 activation of component.
Ames test can be easily in many CRO place's acquisitions all over the world.
Abbreviation
Figure BPA00001375069700502
All references that this application relates to comprise patent and patent application, are incorporated herein by reference to greatest extent at this.
Run through in whole specification sheets and the appending claims, unless context need refer else, otherwise word ' comprises (comprise) ' and its variation for example ' comprises (comprises) ' and ' containing ', be interpreted as hint and comprise described integer, step, integer group or step group, but do not get rid of any other integer, step, integer group or step group.

Claims (15)

1. formula II compound:
Figure FPA00001375069600011
Wherein
R 3Be C 1-C 3Alkyl or C 3-C 6Cycloalkyl, any is optional by one or two methyl and/or by fluorine, trifluoromethyl or methoxyl group replacement among both, works as R 3Be C 3-C 6During cycloalkyl, as selecting it can be by fluorine together with replacement;
R 4Be methyl or fluorine; M is 0,1 or 2;
E is a chemical bond or optional by the thiazolyl of methyl or fluorine replacement;
A 1Be CH or N,
A 2Be CR 6R 7Or NR 6, condition is A at least 1With A 2One of contain N;
N is 0 or 1, makes to contain A 1And A 2Ring be the saturated azo-cycle that contains of 5 or 6 annular atomses;
R 6Be H, C 1-C 4Alkyl, C 1-C 4Haloalkyl, C 1-C 3Alkyl-O-C 1-C 3Alkyl, or work as A 2During for C, R 6Can also be C 1-C 4Alkoxyl group or F;
R 7Be H, C 1-C 4Alkyl or F;
Or its pharmacy acceptable salt, N-oxide compound or hydrate.
2. according to the compound of claim 1, have Formula Il a:
Figure FPA00001375069600021
Wherein
R 3C for side chain 2-C 6Alkyl or C 3-C 6Cycloalkyl, any is chosen wantonly and replaces by one or two fluorine or by trifluoromethyl among both;
R 4Be methyl or fluorine, m is 0,1 or 2;
R 5Be H, methyl or fluorine;
R 6Be C 1-C 6Alkyl; Or its pharmacy acceptable salt, N-oxide compound or hydrate.
3. according to the compound of arbitrary aforementioned claim, R wherein 3Be leucic side chain.
4. according to the compound of arbitrary aforementioned claim, wherein m represents 0, and R 5Expression F.
5. according to each compound in the claim 1 to 4, wherein n represents 1, R 4Be F, and R 5Be H.
6. according to the compound of claim 5, R wherein 4The position shown in following part-structure:
Figure FPA00001375069600022
7. according to each compound among the claim 2-6, wherein R 6Be CH 3
8. according to the compound of claim 1, it is selected from:
Figure FPA00001375069600031
Or its pharmacy acceptable salt, hydrate or N-oxide compound.
9. pharmaceutical composition, it comprises the defined compound of arbitrary aforementioned claim and its pharmaceutically acceptable carrier or thinner.
10. each defined compound is used for preventing or treating the application of the medicine that is selected from following disease in the claim 1 to 8 in preparation:
Osteoporosis,
Gingival disease (for example gingivitis and periodontitis),
Osteitis deformans,
The hypercalcemia of malignant tumour,
Metabolic osteopathy,
Be characterized as excessive cartilage or substrate degradation disease (for example osteoarthritis and rheumatic arthritis),
Osteocarcinoma, comprise tumorigenesis,
Pain (particularly chronic pain).
11. according to each compound in the claim 1 to 8, it is used for the treatment of or prevents following disease:
Osteoporosis,
Gingival disease (for example gingivitis and periodontitis),
Osteitis deformans,
The hypercalcemia of malignant tumour,
Metabolic osteopathy,
Be characterized as excessive cartilage or substrate degradation disease (for example osteoarthritis and rheumatic arthritis),
Osteocarcinoma, comprise tumorigenesis,
Pain (particularly chronic pain).
12. be used for the treatment of by the disease mediated method of cathepsin K, comprise curee to needs treatments use safety and significant quantity according to each compound in the claim 1 to 8.
13. the method for claim 12, wherein said disease is selected from:
Osteoporosis,
Gingival disease (for example gingivitis and periodontitis),
Osteitis deformans,
The hypercalcemia of malignant tumour,
Metabolic osteopathy,
Be characterized as excessive cartilage or substrate degradation disease (for example osteoarthritis and rheumatic arthritis),
Osteocarcinoma, comprise tumorigenesis,
Pain (particularly chronic pain).
14. following formula: compound:
Figure FPA00001375069600051
Rb group definition ketal wherein, for example dimethyl ketal, or described Rb group define cyclic ketal for example 1,3-dioxolane or the protected derivative of its N together.
15. according to the compound of claim 14, it is:
Figure FPA00001375069600052
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