CN1198096A

CN1198096A - C-proteinase inhibitor for treatment of disorders related to overproduction of collagen

Info

Publication number: CN1198096A
Application number: CN96197271A
Authority: CN
Inventors: M·布伦内尔; W·B·何
Original assignee: Fibrogen Inc
Current assignee: Fibrogen Inc
Priority date: 1995-08-08
Filing date: 1996-08-08
Publication date: 1998-11-04
Also published as: KR19990036271A; MX9801093A; BR9609883A; JPH11511137A; EP0845987A1; WO1997005865A1; CA2229098A1; AU6951296A; EP0845987A4

Abstract

The present invention relates to the novel use of organic molecules capable of inhibiting C-proteinase activity in order to regulate, modulate and/or inhibit abnormal collagen formation.

Description

The C-protease inhibitor that is used for the treatment of the disease relevant with the excessive generation of collagen protein

The application relates to and is application on February 14th, 1996, name is called the part continuation application of the US application serial no 08/601203 of " the C-protease inhibitor that is used for the treatment of the disease relevant with the excessive generation of collagen protein ", and the latter is the part continuation application of the interim US application serial no 60/002038 of application on August 8 nineteen ninety-five.1. invention field

Collagen protein, together with other material, is essential for the suitable formation of connective tissue.Therefore, the generation superfluous or not enough generation or Abnormal collagen (comprising mismachined collagen protein) of collagen protein is relevant with many connective tissue diseases and symptom.Evidence suggests that C-protease is the enzyme of the main key of collagen protein Appropriate maturation, because of but ideal goal for suppressing, control and/or regulating collagen protein to form.

The present invention relates to suppress C-proteinase activity to regulate, to alleviate and/or to suppress the organic molecule of undesired collagen protein formation.More particularly, the present invention relates to the purposes that compound and its pharmaceutical composition are used for the treatment of the relevant various diseases of inappropriate with collagen protein or irregular generation.2. background of invention

Collagen structure.At present identified the collagen protein of 19 types, these collagen protein, comprise fibril collagen protein I, II, III type, synthetic as tropocollagen precursor molecule, the peptide expandable part that it contains amino and carboxyl terminal.These are referred to as peptide expandable part difference called after N-and the C-propetide of " forefoot area ".

Forefoot area is secreted cracking when producing ripe triple helix collagen molecules from cell at triple helix precursor molecule.When cracking, " maturation " collagen molecules can for example be incorporated in the collagen fabric of highly structural.Referring to for example Fessler and Fessler, 1978, Annu. Rev. Biochem. 47:129-162; Bornstein and Traub, 1979, protein (eds. Neurath, H. and Hill, R.H.), Academic Press, New York, 412-632 page; Kivirikko etc., 1984, Extracellular Matrix Biochemistry (eds.Piez, K. A. and Reddi, A. H.), Elsevier Science Publishing Co., New York, 83-118 page; Prockop and Kivirikko, 1984, N. Engl. J. Med. 311:376-383; Kuhn, 1987, the 26S Proteasome Structure and Function of collagen protein type (eds. Mayne, R. and Burgeson, R.E.) Academic Press, Inc. Orlando, Florida, 1-42 page.

The disease relevant with the abnormal generation of collagen protein.One group of critical disease is relevant with the inappropriate or irregular generation of collagen protein, it comprises pathology fibre modification or cicatrix, comprise endocardial sclerosis, idiopathic interstitial fibre modification, interstitial pnemnofibrosis, epimysium fibre modification, Symmer ' s fibre modification, the all fibre modifications of maincenter, hepatitis, dermatofibroma, biliary cirrhosis, alcoholic cirrhosis, acute pnemnofibrosis, idiopathic pulmonary fibrosis, acute respiratory distress syndrome, kidney fibre modification/glomerulonephritis, kidney fibre modification/diabetic nephropathy, scleroderma/systemic scleroderma, scleroderma/locality scleroderma, keloid, cicatrix is loose, serious joint accretion/arthritis, myelofibrosis, corneal scarring, vesical fibrosis, duchenne muscular dystrophy, cardiac fibers degeneration, myofibrosis/retina is separated, esophagostenosis, Pai Er (payronles) disease.Other fibrotic conditions can cause or be brought out by operation, comprises that cicatrix correction/plastic operation, glaucoma, cataract fibre modification, corneal scarring, joint accretion, graft versus host disease, tendon operation, nerve are carried secretly, dupuytren contracture, OB/GYN adhesion/fibre modification, pelvis adhesion, epidural fibre modification, restenosis.A kind of strategy for the treatment of these diseases is the excessive generations of pathologic that suppress collagen protein.Therefore the molecule that, evaluation and separation energy are controlled, suppressed and/or adjusting collagen protein produces is main medical science interest.

Mutual relation between collagen protein formation and C-protease.In recent years data thinks, C-protease is main crucial enzyme, and its catalysis is fibril collagen protein for example, comprises the cracking of the C-propetide of I type, II type and III collagen type.Referring to the US application serial no 60/002038 of the application on August 8 nineteen ninety-five as provisional application, and list of references wherein.

C-protease is first at people and Mus fibrocyte (Goldberg etc., 1975, cell 4:45-50; Kessler and Goldberg, 1978, Anal. Biochem. 86:463-469) and chicken tendon fiber cell (Duskin etc., 1978, Arch. Biochem. Biophys. 185:326-332; Leung etc., 1979, J. Biol. Chem. 254:224-232) in tissue culture medium (TCM), find.The acid protease that can remove C-end propetide from I type tropocollagen is also identified.Davidson etc., 1979, Eur.J.Biochem. 100:551.

The partially purified protease with C-proteinase activity obtained from chicken braincap in nineteen eighty-two.Njieha etc., 1982, Biochemistry 23:757-764.In 1985, separated from the conditioned medium of Embryo Gallus domesticus tendon, purification and characterization chicken C-protease.Hojima etc., 1985, J. Biol. Chem. 260:15996-16003.The Mus C-protease of having purified in the fibrocellular tissue culture medium (TCM) of Mus subsequently.Kessler etc., 1986, Collagen Relat.Res. 6:249-266; Kessler and Adar, 1989, Eur. J. Biochem. 186:115-121.Finally, as the related application of above-mentioned reference and disclosed list of references wherein, identified the human body C-protease of cDNA coding.

With the experiment that these chickens and Mus C-protease purification form are carried out, show, enzyme can work in the process that forms function collagen fabric.Fertala etc., 1994, J. Biol. Chem.269:11584.

C-protease inhibitor.Due to the importance that collagen protein is produced that enzyme demonstrates, scientist has identified many protease inhibitor.Referring to such as Hojima etc., document is the same.For example some metal-chelator has been proved the activity with C-protease inhibitor.Equally, chymotrypsin inhibitor and pepsin A are found to be stronger C-protease inhibitor.In addition α, ₂-macroglobulin, ovostatin and calf serum show at least part of inhibition C-proteinase activity.

Equally, dithiothreitol, DTT, SDS, concanavalin A, Zn ²⁺, Cu ²⁺and Cd ²⁺be reported in and under low concentration, have inhibition ability.Equally, some reducing agent, some aminoacid, phosphate and ammonium sulfate have inhibition ability under 1-10mM concentration.In addition, enzyme demonstrates by basic amino acid lysine and arginine and suppresses.Leung etc., document is the same; Ryhanen etc., 1982, Arch. Biochem. Biophys.215:230-236.Finally, it is found that high concentration chlorination sodium or Tris-HCl buffer suppress C-proteinase activity.For example, according to reports, use 0.2,0.3 and 0.5M sodium chloride, C-proteinase activity be reduced to respectively under 0.15M code test concentration, observe 66,38 and 25%.Tris-HCl buffer under 0.2-0.5M concentration has significantly suppressed activity, Hojima etc., and document is the same.In contrast, microbial inhibitor is considered to weak or there is no C-proteinase activity as leupeptin, phosphoric acid methadone (phosphoramidon), antipain, bestatin elastin laminin and amandin (amastatin).

Various determination of test method have been used in C-proteinase activity and inhibition thereof.Referring to for example Kesslerand Goldberg, 1978, Anal. Biochem.86:463; Njieha etc., 1982, biochemistry 21:757-764.Although had these tests, due to the limited availability of human body C-protease, at least carried out not yet research and the test of large-scale potential C-protease inhibitor.Described in many publications, enzyme is difficult to by conventional biochemical method separated, and until the report in above-mentioned reference and relevant patent application, the cDNA sequence of this enzyme of encoding is unknown.

Exploitation suppresses the compound of C-proteinase activity.Consider the important function in collagen protein formation and maturation, it is the inappropriate or irregular generation for the treatment of and collagen protein and the ideal goal of ripe relevant disease that C-protease demonstrates.Yet it is effective or the inhibitor of C-proteinase activity that the inhibitor of up to the present identifying is not all proved for the treatment disease relevant with collagen protein.

Can be special inhibition C-proteinase activity with the evaluation of the active compound that regulates and adjust abnormal or unsuitable collagen protein and produce because of but desirable, and be object of the present invention.3. summary of the invention

The present invention relates to can regulate, adjust and/or suppress collagen protein generation and/or ripe organic compound by affecting C-proteinase activity.Compound of the present invention especially has following formula: a. inhibitor A

or

Wherein

R ₁be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, halo aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₂be selected from H, low alkyl group;

R ₃be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, halo aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₄be selected from aryl, heteroaryl, alkyl, aralkyl, heteroarylalkyl, alkyl amino, aryl alkyl amino:

X is selected from SO ₂, C=O;

Y is selected from OH, HOHN (azanol), H ₂n, alkyl amino;

Z is direct bond, methylene, oxygen, sulfur, amino;

N is 0 or 1;

Or b. inhibitor B

Wherein

R ₁be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₂be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₃be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₄be selected from H, low alkyl group;

R ₅be selected from H, low alkyl group, carboxyalkyl, (list or dialkyl amido) alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl, alkoy alkyl acyl alkyl;

Or c. inhibitor C

Wherein

R ₁be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₂be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₃select white H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₄be selected from H, low alkyl group;

Or d. inhibitor D

Wherein

R ₄be selected from H, low alkyl group;

Or e. inhibitor E

Wherein

R ₁be selected from OH, alkoxyl, low alkyl group, alkyl amino, peptide;

X is selected from N, C;

R ₃to be selected from H, low alkyl group, list or multi-haloalkyl, carboxyalkyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, biaryl, biaryl alkyl, hydroxy alkyl, alkoxyalkyl, acyloxy alkyl, mercaptoalkyl, (amino, list or dialkyl amido) alkyl, amidoalkyl, cycloalkyl, Heterocyclylalkyl, cycloalkyl-alkyl, Heterocyclylalkyl alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl;

R ₄be selected from H, low alkyl group;

R ₅be selected from H, low alkyl group, carboxyalkyl, (list or dialkyl amido) alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl, alkoy alkyl acyl alkyl.

The invention still further relates to pharmaceutical composition, it contains above-claimed cpd and pharmaceutically useful carrier or the excipient for the treatment of effective dose.Said composition is by suppressing generation and/or the maturation of C-proteinase activity scalable collagen protein.

The invention still further relates to disclosed compound regulates, suppresses and/or adjust C-proteinase activity with the purposes of the treatment disease relevant with the inappropriate or irregular generation of collagen protein for passing through with compositions.

More particularly, compositions of the present invention can be included in the method for the treatment of and the inappropriate or disease that irregular generation is relevant of collagen protein, and described disease comprises, but is not restricted to rheumatoid arthritis, scleroderma, pathology fibre modification or cicatrix.4. definition

" C-protease " should refer to by-Ala ↓ Asp-Asp-and/or Gly ↓ Asp-Glu-cracking can processed collagen protein molecular, the enzyme of derivant or fragment or their precursor.What this term should comprise human body C-protease and theirs has C-protease like active derivant, analog, fragment and variant.

" pharmaceutically useful salt " refers to and retains the biological effectiveness of free acid and the salt of character, they can by with inorganic or organic base, such as sodium hydroxide, magnesium hydroxide, ammonia, trialkylamine, dialkylamine, monoalkylamine, binary amino acid, sodium acetate, Potassium Benzoate, triethanol ammonium etc.

" alkyl " refers to saturated aliphatic hydrocarbon, comprises straight chain, side chain and cycloalkyl.Alkyl preferably contains 1-12 carbon atom.It is most preferably 1-7 carbon atom, the more preferably low alkyl group of 1-4 carbon atom.Typical alkyl group comprises methyl, ethyl, propyl group, isopropyl, butyl, isobutyl group, the tert-butyl group, amyl group, hexyl etc.Alkyl can be substituted or unsubstituted.When being substituted, substituent group is carboxyl, hydroxyl, sulfydryl, cycloalkyl, Heterocyclylalkyl, halogen, alkoxyl, alkyl amino preferably.

" aryl " refers to aromatic group, and it contains at least one ring with conjugated pi electron system, and it comprises isocyclic aryl, heteroaryl and biaryl, and they can optionally be substituted.Preferred aryl is to replace or unsubstituted phenyl or pyridine radicals.Preferred aryl substituent, preferably phenyl or pyridine radicals, be halogen, trihalomethyl, hydroxyl, SH, NO ₂, amine, thioether, cyano group, alkoxyl.5. detailed Description Of The Invention

The present invention relates to the compound that can adjust and/or regulate collagen protein to form by suppressing C-proteinase activity.

More particularly, the present invention relates to such compound, it suppresses C-proteinase activity as Therapeutic Method, to treat or to control various connective tissue diseases, comprises fibrotic conditions, joint disease or the disease that is caused or caused by the infringement of performing the operation or happen suddenly.5.1. compound

The present invention relates generally to have compound and/or the compositions of following general formula; A. inhibitor A

Or

Wherein

R ₂be selected from H, low alkyl group;

X is selected from SO ₂, C=O;

Y is selected from OH, HOHN (azanol is amino), H ₂n, alkyl amino;

Z is direct bond, methylene, oxygen, sulfur, amino;

N is 0 or 1;

Or b. inhibitor B

wherein

R ₄be selected from H, low alkyl group;

Or c. inhibitor C

Wherein

R ₄be selected from the alkyl of H, low alkyl group;

Or d. inhibitor D

Wherein

R ₄be selected from H, low alkyl group;

R ₅to be selected from H, low alkyl group, carboxyalkyl, (list or dialkyl amido) alkyl, alkyl (sulfenyl, sulfinyl or sulfonyl) alkyl, alkoy alkyl acyl alkyl;

Or e. inhibitor E

Wherein

R ₁be selected from OH, alkoxyl, low alkyl group, alkyl amino, peptide;

X is selected from N, C;

R ₄be selected from H, low alkyl group;

In specific embodiment of the invention scheme, compound of the present invention can have following formula: with its pharmaceutically useful salt;

Or: with its pharmaceutically useful salt;

Or:

with its pharmaceutically useful salt;

Or:

with its pharmaceutically useful salt;

Or:

with its pharmaceutically useful salt;

Or: with its pharmaceutically useful salt;

Or:

with its pharmaceutically useful salt;

Or: with its pharmaceutically useful salt;

Or: with its pharmaceutically useful salt.

Can there is enantiotropy in chemical formula of the present invention.A kind of in the change form that only can express possibility due to the structural formula drawing in description, should understand the present invention and comprise to have by suppressing C-proteinase activity and adjust and/or regulate the generation of collagen protein and/or any tautomeride form of ripe ability.In addition, should understand all possible stereoisomer that the present invention includes every kind of described compound.

Except above-claimed cpd and their pharmaceutically useful salt, when needed, the invention still further relates to and for example have, by suppressing C-proteinase activity and suppress, adjust and/or regulating the solvation of the generation of collagen protein and/or the compound of ripe ability and solvation form (hydrated form) not.

Above-claimed cpd can be prepared by being suitable for any known method of the compound that preparation is chemically relevant.Suitable method is illustrated by following representational embodiment.Required starting material can obtain by vitochemical standard method.

Single compound can be used known technology to measure as the related activity and the effect that affect the medicine of C-proteinase activity.Compound preferably carries out a series of screening and regulates, adjusts and/or the generation of inhibition collagen protein and ripe ability to measure compound.These screenings comprise biochemical test, cell culture test and animal model.5.2. indication

Can use compound of the present invention and inappropriate or irregular generation and/or the ripe relevant disease of compositions treatment with collagen protein, it comprises arthritis disease, fibrotic conditions and other connective tissue disease.

These diseases or symptom comprise pathology fibre modification or cicatrix, comprise endocardial sclerosis, idiopathic interstitial fibre modification, interstitial pnemnofibrosis, epimysium fibre modification, Symmer ' s fibre modification, the all fibre modifications of maincenter, hepatitis, dermatofibroma, biliary cirrhosis, alcoholic cirrhosis, acute pnemnofibrosis, idiopathic pulmonary fibrosis, acute respiratory distress syndrome, kidney fibre modification/glomerulonephritis, kidney fibre modification/diabetic nephropathy, scleroderma/systemic scleroderma, scleroderma/locality scleroderma, keloid, cicatrix is loose, serious joint accretion/arthritis, myelofibrosis, corneal scarring, vesical fibrosis, duchenne muscular dystrophy, cardiac fibers degeneration, myofibrosis/retina is separated, esophagostenosis, payr's disease.Other fibrotic conditions can cause or be brought out by operation, comprises that cicatrix correction/plastic operation, glaucoma, cataract fibre modification, corneal scarring, joint accretion, graft versus host disease, tendon operation, nerve are carried secretly, dupuytren contracture, OB/GYN adhesion/fibre modification, pelvis adhesion, epidural fibre modification, restenosis.Other fibrotic conditions can cause by chemotherapy, and it comprises such as pnemnofibrosis etc.5.3. pharmaceutical preparation and route of administration

The compound of identifying can compound itself or with the pharmaceutical compositions of itself and suitable carrier or mixed with excipients with the dosage for the treatment of or improving various diseases to patient's administration.Treatment effective dose refers to the amount of the compound that is enough to improve symptom.Can " Remington ' s Pharmaceutical Sciences " Mack PublishingCo. for the preparation of the application's compound and the technology of administration, Easton, PA, finds in latest edition.5.3.1. route of administration

Suitable route of administration can for example comprise per os, rectum, through film or through enteral administration; Parenteral, comprises intramuscular, subcutaneous, intramedullary injection, and in sheath, directly in ventricle, in intravenous, perineum, intranasal or intraocular injection.

In addition, people can be partly rather than are given capapie drug compound, for example, compound is injected directly in intraarticular or fibre modification tissue, conventionally to store or extended release preparation form.For the cicatrix of avoiding conventionally staying after operation for glaucoma, compound is administration partly, for example, as eye drop.

In addition, people can organize target administration to for example arthritis or fibre modification by target drug delivery system, for example, to have applied the liposome form of specific antibodies.Liposome will arrive target, and optionally be absorbed by tissue lesion.5.3.2. compositions/preparation

Pharmaceutical composition of the present invention can known method preparation itself, for example by conventional mixing, dissolving, pelletize, dragee processed, grinding, emulsifying, Rubber Capsule, carry secretly or freeze drying process.

Therefore, can conventional method for pharmaceutical composition of the present invention, use one or more pharmaceutically useful carrier preparations, excipient and adjuvant that described carrier contains the preparation that contributes to reactive compound to be machined for treatment, suitable preparation depends on selected route of administration.

While being used for injecting, medicine of the present invention can be mixed with aqueous solution, preferably in the compatible buffer of physiology, for example, Hank ' s solution, Ringer ' s solution or normal saline buffer solution.For when film administration, in preparation, use the penetrant that is suitable for having barrier to be infiltrated, this penetrant is normally known in the prior art.

For oral administration, compound can be by mixing reactive compound easily preparation with pharmaceutically useful carrier well known in the prior art.This carrier can be mixed with compound of the present invention tablet, pill, sugar-coat agent, capsule, liquid, gel, syrup, slurry agent, suspending agent etc., oral for treated patient.For oral pharmaceutical preparation, can be obtained by solid excipient, optionally grind the mixture obtaining, and adding as required suitable adjuvant post-treatment granulate mixture to obtain tablet or bran clothing ball core.Suitable excipient is filler especially, for example saccharide, for example lactose, sucrose, mannitol or sorbitol; Cellulose preparation, for example corn starch, wheaten starch, rice starch, potato starch, gelatin, Tragacanth, methylcellulose, hydroxypropyl emthylcellulose, sodium carboxymethyl cellulose and/or polyvinyl pyrrolidone (PVP).For example, for example, if need, can add disintegrating agent, crospolyvinylpyrrolidone, Colla Corii Asini or alginic acid or its salt, sodium alginate.

Dragee core is with suitable coating.For this reason, can use concentrated sugar juice, it optionally contains arabic gum, Talcum, polyvinylpyrrolidone, carbopol glue, Polyethylene Glycol and/or titanium dioxide, solution and suitable organic solvent or solvent mixture spray paint.Dyestuff or pigment can add in tablet or dragee coating, to identify or to distinguish the various combination of active compound doses.

Can be used for the capsule that oral pharmaceutical preparation comprises the propelling charging of being prepared by gelatin, and by gelatin and plasticizer, the soft seal capsule that for example prepared by glycerol or sorbitol.Advance the capsule of charging can contain and filler, for example lactose, binding agent, for example starch and/or lubricant, for example Talcum or magnesium stearate and, optionally, the active component that stabilizing agent mixes.In soft capsule, reactive compound solubilized or be suspended in suitable liquid, for example fatty oil, liquid paraffin or liquid macrogol.In addition, can add stabilizing agent.For oral all preparations, it should be the dosage form that is suitable for administration like this.

For buccal administration, compositions can adopt tablet or the lozenge form with conventional method preparation.

For passing through inhalation, the compound that the present invention uses is easily from pressurized package or aerosol apparatus, use suitable propellant, for example dichlorodifluoromethane, Arcton 11, dichlorotetra-fluoroethane, carbon dioxide or other suitable gas provide with aerosol spray form.In pressurized aerosol situation, can provide the valve of metered amounts to determine dosage device by installation.For the capsule of for example gelatin of inhaler or insufflator or cartridge case, can be mixed with and contain compound and suitable powder substrate, for example mixture of powders of lactose or starch.

Compound can be mixed with by injection for parenteral, for example, by single fast injection or continuous infusion.For the preparation of injecting, can exist by unit dosage form, for example, at the ampulla that contains the antiseptic adding or multi-dose container.Compositions can adopt suspension, solution or the emulsion form in oil or aqueous carrier, and can contain formulation aid, for example suspending agent, stabilizing agent and/or dispersant.

The aqueous solution that comprises the reactive compound of water-soluble form for the pharmaceutical preparation of parenteral.In addition, the suspension of reactive compound can be prepared into suitable oily injectable suspensions.Suitable lipophilic solvent or excipient comprise fatty oil, for example Oleum sesami, or Acrawax, for example ethyl oleate or glyceride, or liposome.Moisture injectable suspensions can contain the material that increases suspension liquid viscosity, for example sodium carboxymethyl cellulose, sorbitol or glucose.The material that suspension also optionally contains suitable stabilizing agent or increases compound dissolution is to prepare highly concentrated solution.

In addition, active component can be powder type, for before use with suitable excipient, for example aseptic apirogen water is mixed.

Compound also can be mixed with rectal compositions, for example, contain conventional suppository bases, for example the suppository of cocoa butter or other glyceride or enema,retention.

Except above-mentioned preparation, compound is also mixed with storage preparation.This long effect preparaton can be for example, by implantation (subcutaneous or intramuscular) or by intramuscular injection administration.Therefore, for example compound can be used suitable polymerization or hydrophobic substance (for example, as the emulsion in available oil) or ion exchange resin, or as slightly soluble derivant, for example, as slightly soluble salt, prepares.

Pharmaceutically useful carrier for hydrophobic compound of the present invention is cosolvent system, and it contains benzyl alcohol, non-polar surfactant, the miscible organic polymer of water and water.Cosolvent system can be VPD cosolvent system.VPD is the solution of the dehydrated alcohol of 3%w/v benzyl alcohol, 8%w/v non-polar surfactant Tween 80 and 65%w/v Liquid Macrogol and surplus.VPD cosolvent system (VPD:5W) is by forming with the VPD of 5% D/W dilution in 1: 1.Be total to the abundant solubilizing hydrophobic compound of solution system, and this produces low toxicity when whole body administration.The ratio of cosolvent system naturally can consider not destroy its dissolubility and toxic characteristic changes.In addition, cosolvent component can change, and for example, other hypotoxicity non-polar surfactant can be used for replacing Tween 80; The fixed portion size of Polyethylene Glycol can change; Other biocompatible polymer can replace Polyethylene Glycol, for example polyvinylpyrrolidone; Can replace glucose with other sugar or polysaccharide glycosides.

In addition, can adopt other drug delivery system that is applicable to dewatering medicament compound.Liposome and emulsion are to be applicable to the administration excipient of dewatering medicament or the known embodiment of carrier.Some organic solvent, for example dimethyl sulfoxide also can be used, although cost is larger toxicity conventionally.In addition, compound can be used slow-releasing system administration, for example, contain the semi-permeable matrix of the solid hydrophobic polymer of medicine.Having confirmed various slow-release materials, is well known by persons skilled in the art, and according to its chemical property, slow releasing capsule discharges compound sustainable several weeks to over 100 days.Chemical property and biological stability according to treatment reagent, can adopt other strategy that is applicable to protein stabilization.

Pharmaceutical composition also can contain suitable solid or gel phase carrier or excipient.The example of this carrier or excipient comprises, be not restricted to, calcium carbonate, calcium phosphate, various sugar, cellulose derivative, gelatin and polymer, for example Polyethylene Glycol.

Many C-protease inhibiting compounds of the present invention can be used as the salt forming with pharmaceutically useful counter ion counterionsl gegenions and provide.This pharmaceutically useful base addition salts is the salt that has retained physiologic effect and the character of free acid, they by with inorganic or organic base, for example sodium hydroxide, magnesium hydroxide, ammonia, trialkylamine, dialkylamine, monoalkylamine, binary amino acid, sodium acetate, Potassium Benzoate, triethanol ammonium etc. reaction obtain.5.3.3. effective dose

Be applicable to pharmaceutical composition of the present invention and comprise compositions, its active component that contains effective dose is to obtain required object.More particularly, effective dose refer to prevent the existing Symptomatic development of treated patient or improvement effective dose.Those skilled in the art can determine effective dose well, especially after with reference to detailed description of the present invention.

For any compound using in the method for the invention, originally treatment effective dose can be determined by cell culture test.For example, this dosage can determine to obtain circulation composition scope in animal model, and it is included in the IC measuring in cell culture ₅₀(that is, test compound obtains the maximum concentration suppressing of half of C-proteinase activity).This data can be used for determining more accurately the using dosage in human body.

Treatment effective dose refers to the amount of the compound that causes the improvement of patient's symptom or existence prolongation.The toxicity of this compound and treatment effect can be definite by the standard drug experiment in cell culture or laboratory animal, for example, and for measuring LD ₅₀(overall 50% lethal dosage) and ED ₅₀(effectively treat overall 50% dosage).Dosage rate between toxicity and therapeutic effect is treatment coefficient, its available LD ₅₀with ED ₅₀ratio represent.The data that obtained by cell culture test and zooscopy can be used for determining the dosage range of human body.The dosage of this compound is preferably placed within the scope of circulation composition, and described concentration comprises having less or do not have a virose ED ₅₀.Dosage can change according to adopted dosage form and route of administration used.Definite preparation, route of administration and dosage can consider that patient's situation selects by clinician.Referring to, for example, Fingl etc., 1975, " pharmacological basis for the treatment of ", chapter 1, page 1.

Dosage and interval can regulate respectively to provide plasma content or the minimum effective drug concentration (MEC) that is enough to keep C-protease inhibition.For every kind of compound MEC, will change, but can be determined by vitro data; For example use test as herein described to determine that C-protease 50-90% suppresses required concentration.Obtain the required concentration of MEC by the feature and the route of administration that depend on separately.Yet HPLC test or biologic test can be used for determining plasma concentration.

Spacing of doses also can be used MEC value to determine.Compound should carry out administration by certain dosage regimen, and this scheme is for 10-90%, preferred 30-90%, and most preferably the time of 50-90% keeps plasma concentration higher than MEC.

The in the situation that of topical or selection absorption, effective local concentration of medicine may be irrelevant with plasma concentration.

Certainly, the amount of administration composition will depend on the body weight of treated receptor, receptor, the seriousness of illness, administering mode and clinician's judgement.5.3.4. packing

If needed, compositions can exist in packing or dispensing device, and it can contain one or more unit dose that contain active component.Packing can for example comprise metal or plastic foil, for example vesicatory packing.Packing or a minute device for dispensing can attach the device for administration.Contain the compositions that is formulated in the compound of the present invention in pharmaceutically useful carrier and also can in suitable container, prepare, place, stick the labelling of symptom shown in treatment.Suitable symptom shown on labelling can comprise treatment of arthritis or any other fibrotic conditions.6. embodiment

Compound of the present invention can be synthetic according to known method.Following expression for the synthesis of the method for optimizing with test compound of the present invention.6.1. embodiment 1: compou nd synthesis 6.1.1. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide synthetic

For the synthesis of N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide (5) (and for name object; concrete in this manual compound can be described as " C# "; wherein " # " is Arabic numerals); also referred to as FG047 (embodiment of inhibitor A), method for optimizing as follows:

2-[(4-chlorobenzyl) amino] acetic acid ethyl ester (3) synthetic.At glycine ethyl ester hydrochloride (1) (1.00g, 7.16mmol) and 4-chlorobenzaldehyde (2) (1.00g, 7.16mmol), in the cooling solution of methanol (10ml), add anhydrous zinc chloride (75mg, 0.55mmol) and NaBCNH ₃(0.45g, 7.16mmol).At room temperature stir after 18 hours, by 1N aqueous hydrochloric acid solution (20ml) reaction mixture, then stir 30 minutes.With rotary evaporator enriched mixture, to remove most of methanol solvates, then use ether (20ml) to extract.Water layer uses carefully 45% (w/w) KOH aqueous solution to alkalize to pH10 in ice bath, by ethyl acetate (2 * 50ml), extracts.The ethyl acetate organic layer salt water washing merging, by dried over sodium sulfate, and concentrated.Crude product silica gel chromatography (4/1, hexane/ethyl acetate) purification obtains 2-[(4-chlorobenzyl) amino] acetic acid ethyl ester (3) grease.

2-[[N-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] acetic acid ethyl ester (4) synthetic.By triethylamine (294mg, 2.90mmol) drop to 2-[(4-chlorobenzyl) amino] acetic acid ethyl ester (3) (600mg, 2.64mmol) and the solution of p-methoxybenzene sulfonic acid chloride (545mg, 2.64mmol) in anhydrous methylene chloride (7ml) in.Mixture at room temperature stirs 15 hours, by 1N hydrochloric acid (20ml) solution-treated.It is biphase that separation obtains, water layer dichloromethane extraction.The organic layer salt water washing merging, by dried over mgso concentrated.Silica gel chromatography for crude product (3/1-2/1 hexane/ethyl acetate) purification obtains 2-[[N-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] acetic acid ethyl ester (4) grease.

N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide (5) synthetic.Under boiling point, prepare respectively hydroxy amine hydrochloric acid salt (171mg, 2.46mmol) solution in methanol (1.3ml) and KOH (207mg, 3.69mmol) solution in methanol (1.3ml), is cooled to 40 ℃, then the latter solution is added to the former.In ice bath, reaction mixture, after 30 minutes, filters out solid potassium chloride.In filtrate, add ethyl ester (4) (487mg, 1.23mmol).At room temperature stir after 6 hours, 1N hydrochloric acid (20ml) solution-treated for reactant mixture, uses dichloromethane extraction.Organic layer salt water washing, by dried over mgso, the concentrated crude product that obtains.Residue is developed in ether, collects white solid and obtains hydroxamic acid (hydroxamate), be i.e. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide (5), also referred to as FG-047.

mp：124-125℃；MS(ES)(M+H) ⁺：385；1H?NMR(360MHz，

DMSO-d6)δ10.47(s，1H，OH)，8.81(s，1H，NH)7.81-7.08(m，8H，

Ph), 4.34 (s, 2H, CH ₂cO), 3.85 (s, 3H, OMe), 3.63 (s, 2H, CH ₂ph) .6.1.2. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-luorobenzyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-luorobenzyl)] amino] acetamide (6); also referred to as the method for optimizing of FG053 (embodiment of inhibitor A) substantially with described synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] method of acetamide is identical, uses starting material 4-fluorobenzaldehyde to replace 4-chlorobenzaldehyde.Referring to, 6.1.1. part:

mS (ES) (M-1) ^-: 367; ¹hNMR (360MHz, DMSO-d6) δ 10.45 (s, 1H, OH), 8.83 (s, 1H, NH), 7.81-7.08 (m, 8H, Ph), 4.33 (s, 2H, CH ₂cO), 3.85 (s, 3H, OMe), 3.62 (s, 2H, CH ₂ph) method for optimizing of the synthetic hydroxamic acid (11) (embodiment of inhibitor B) of .6.1.3. hydroxamic acid is as follows:

with bromoacetic acid tertiary butyl ester alkylation malonate (7), obtain four esters (8).Through saponification and decarboxylic reaction, the acid obtaining (9) obtains (10) with Glu (OBn) NHMe through the coupling of peptide coupling reaction.Hydrolysis, acid anhydride subsequently form and NH ₂after OH addition, (10) can be converted into required hydroxamic acid (11).6.1.4. N-carboxymethyl dipeptides is synthetic

For the synthesis of N-carboxymethyl dipeptides (16), as follows also referred to as the method for optimizing of FG057 (embodiment of inhibitor D):

The construction unit easily being obtained by literature method (12) and (13), obtain dipeptides (14) by classical DCC/HOBt method coupling with 78% yield.(14) be used in the TFA deprotection in dichloromethane, form N-carboxymethyl dipeptides (15) subsequently under NMM exists with the alkylation of bromoacetic acid benzyl ester, it is converted into free acid (16) by catalytic hydrogenation.

Boc-Glu (OBn) OH and Boc-Asp (OBn) OH are used method well known in the prior art synthetic.

Boc-Glu (OBn) NHMe's (13) is synthetic: triethylamine (2.0g, 20mmol) is added dropwise in the solution of Boc-Glu (OBn) OH (6.7g, 20mmol) in anhydrous THF (100ml).Solution is cooled to-78 ℃ under argon atmospher, then drips ethyl chloroformate (2.2g, 20mmol).Make reactant mixture in 2 hours, be warmed to-30 ℃, add 40% aqueous solution of methylamine (22mmol), make reactant mixture be warmed to room temperature.Reaction is stirred after 1 hour again, add ether (50ml) and water (70ml).Isolate organic layer, with 1M sodium bicarbonate, 10% citric acid and saturated nacl aqueous solution washing, use dried over mgso.Vacuum evaporating solvent obtains white solid (5.0g, 72% yield).

¹H-NMR(200MHz，CDCl ₃)：δ1.38(s，9H，CH ₃)；1.85-2.59(m，

4H，CH ₂)；2.75(d，3H，NCH ₃)；4.17(m，1H，CH)；5.08(δ，2H，OCH ₂)；5.33

(bδ，1H，NH)；6.30(bs，1H，NH)；7.31(s，5H，Ph-H).

Boc-Asp (OBn)-Glu (OBn) NHMe's (14) is synthetic: at t-butoxycarbonyl amino ester (13) (700mg, 2mmol) in the solution in 10ml dichloromethane, add 1.5mlTFA, reactant mixture at room temperature stirs 1 hour under argon atmospher.The acid that vacuum evaporation is excessive, residue is processed for several times with ether, and concentrating under reduced pressure, obtains colorless oil, and it is without further purification.Be added in tfa salt, Boc-Asp (OBn) OH (466ml, 2mmol), HOBt (170mg, 2mmol) and NMM (202mg, 2mmol) in dichloromethane (10ml), reactant stirs and spends the night under room temperature and argon atmospher.Remove by filter 1,3-Dicyclohexylurea, concentrating under reduced pressure filtrate.Ethyl acetate for residue (30ml) dilution, filters again, with 1M sodium bicarbonate, 10% citric acid and saturated nacl aqueous solution washing.Organic layer dried over mgso, vacuum concentration obtains white solid, yield 78% (867mg).

¹H-NMR(200MHz，CDCl ₃)：δ1.41(s，9H，CH ₃)；1.85-3.05(m，

9H，3CH ₂，NCH ₃)；4.40(m，2H，CH)；5.01(m，4H，OCH ₂)；5.53(d，1H，

NH)；6.48(bs，1H，NH)；7.15-7.38(m，10H，Ph-H).

Synthesizing of N-carboxymethyl dipeptides (15): to Boc-Asp (OBn)-Glu (OBn) NHMe (14) (555mg, 1mmol) in the solution of 10ml dichloromethane, add 1mlTHF, reactant mixture at room temperature stirs 1 hour in argon atmospher.The acid that vacuum evaporation is excessive, residue is processed several times with ether, and concentrating under reduced pressure, obtains colorless oil, is dissolved in anhydrous THF (20ml).In this solution, add NMM (101mg, 1mmol) and bromoacetic acid benzyl ester (230mg, 1mmol), reactant mixture at room temperature stirs and spends the night in argon atmospher.Concentrating under reduced pressure reactant mixture, ethyl acetate for residue (20ml) dilution, with 1M sodium bicarbonate, 10% citric acid and saturated nacl aqueous solution washing.Organic layer dried over mgso, then concentrates and obtains colorless oil, and it is obtained to (15) (410mg) white solid, yield 88% through fast silica gel chromatogram method (ethyl acetate/methanol 10: 1) purification.

¹H-NMR(200MHz，CDCl ₃)：δ1.83-2.95(m，9H，3CH ₂，NCH ₃)；3.30-3.60(m，4H，CH?&?N?CH ₂)；4.39(m，2H，CH)；5.05(m，6H，OCH ₂)；6.64(bs，1H，NH)；7.29-(m，15H，Ph-H)8.02(bs，-1H，NH)； ¹³C-NMR(60MHz，CDCl ₃)；δ26.17，27.24，30.59，36.57(3CH ₂，NCH ₃)；49.59(NCH ₂)；52.54，59.07(2CH)；66.46，66.71，66.86(3OCH.)；128.17，128.26，128.34，128.55，128.80，135.27，135.44，135.79(Ph-C)；171.25，171.34，172.01，172.61，173.05(5C＝O).

N-carboxymethyl-Asp-Glu-NMe's (16) is synthetic: the N-carboxymethyl dipeptides (15) (68mg, 0.113mg) to benzyl protection adds Pd/C powder (50mg) by criticizing in the solution of methanol (5ml).Mixture under hydrogen atmosphere (airbag pressure) stirring at room 20 hours.By Celite pad filtering catalyst, use washed with methanol.Concentrated filtrate, thick solid by ethyl acetate/methanol recrystallization, obtains product N-carboxymethyl-Asp-Glu-NMe (16) (23mg, 0.065mmol) white solid, yield 58% at-20 ℃.

Mp:147-149 ℃; NMR (360MHz, DMSO-d6) δ 8.19 (d, J=8.5Hz1H), 7.72 (m, 1H), 4.20 (m, 1H), 3.45-3.20 (m, 4H), 2.63-2.44 (m, 5H), 2.20 (m, 2H), 1.93 (m, 1H), synthesizing of 1.72 (m, 1H) .6.1.5. sulfhydryl compound

Synthetic sulfhydryl compound (22), as follows also referred to as the method for optimizing of FG-O74 (embodiment of inhibitor C):

With bromoacetic acid tertiary butyl ester alkylation di(2-ethylhexyl)phosphate ethyl ester (17) under sodium hydride exists, the yield with 90% obtains phosphonate ester (18).Use potassium carbonate as alkali, (18) are carried out Horner-Emmons with formaldehyde and are reacted formation beta-unsaturated esters (19).(19) with LiOH, carry out saponification, then carry out thiacetic Michael addition and obtain acid (20).(20) adopt the coupling of DCC/HOBt method with Glu (OBn) NHMe, by flash chromatography, obtain the dipeptides (21) of non-enantiomer mixture.(21) the TFA deprotection being used in dichloromethane obtains single acid (22).

Synthesizing of 1-ethyl-2-diethyl phosphonyl succinic acid 4-tertiary butyl ester (18): to sodium hydride (0.48g; 20mmol) in the suspension in anhydrous THF (80ml), drip phosphoryl acetic acid triethyl group ester (4.48g; 20mmol) the solution in THF (20ml), is at room temperature added in the bromoacetic acid tertiary butyl ester in THF (20ml) after 3 hours.The suspension obtaining is at room temperature stirred 3 hours, add water (50ml), reactant mixture uses 1M hcl acidifying to pH3.After adding ether (70ml), isolate organic layer, with saturated nacl aqueous solution washing, by dried over mgso, vacuum evaporating solvent obtains colorless oil, yield 90%.

Synthesizing of 1-ethyl-2-methylene-succinic acid 4-tertiary butyl ester (19): by compound (18) (5.4g, 16mmol), potassium carbonate (6.9g, 50mmol) and the mixture of 30% aqueous solution of formaldehyde (3.2g, 100mmol) reflux 3 hours.After cooling, mixture hexane extraction, organic layer water and salt water washing, use dried over mgso.After filtration and vacuum evaporation, obtain oily residue, yield 80% (2.7g).

¹H-NMR(200MHz，CDCl ₃)：δ1.29(t，j＝7Hz，3H，CH ₂)；1.43(s，9H，CH ₃)；3.24(s，2H，CH ₂；4.20(q，J＝Hz，2H，OCH ₂)；5.63，6.28(s，2H，＝CH ₂).

2-acetyl group sulfenyl methyl-succinic acid 4-tertiary butyl ester (20): beta-unsaturated esters (19) (2.3g, 11mmol) is dissolved in THF (50ml), adds 0.2MLiOH (264mg, 11mmol) aqueous solution at 0 ℃.Make mixture be warmed to room temperature, then stir 3 hours.Evaporation organic solvent, ether for water layer (30ml) extracts.Water layer, extracts by ethyl acetate to pH3 with 1M hcl acidifying.Organic layer water and salt water washing, use dried over mgso.After filtration and evaporating solvent, obtain acid, white solid, 60% yield.

¹H-NMR(200MHz，CDCl ₃)；δ1.44(s，9H，CH ₃)，3.26(s，2H，CH ₂)；5.78，6.42(s，2H，＝CH ₂)：10.01(bs，1H，COOH).

Acid (1.0g, 5.4mmol) is dissolved in chloroform, adds thiacetic acid. (1.45g, 19mmol).Mixture stirs 49 hours at 60 ℃, and vacuum evaporating solvent, obtains compound (20) with quantitative yield, is colorless oil.

¹H-NMR(200MHz，CDCl ₃)；δ1.44(s，9H，CH ₃)；2.34(s，3H，CH ₃)；2.45-2.75(m，2H，CH ₂；2.96-3.33(m，3H，CH ₂，CH)；9.74(bs，1H，COOH).

3-acetyl group sulfidomethyl-4-oxo-5-azepine-6-(R)-methylamino formoxyl-8-benzyloxycarbonyl-sad 4-tertiary butyl ester (21): at tert-butoxycarbonyl amino ester (13) (700mg; 2mmol) in the solution in 10ml dichloromethane, add 1mlTFA, reactant mixture at room temperature stirs 1 hour under argon atmospher.The acid that vacuum evaporation is excessive, ether for residue (15ml) is processed for several times, and concentrating under reduced pressure, obtains colorless oil, and it is without further purification.Tfa salt, acid (20) (525mg, 2mmol), HOBt (170mg, 2mmol) and NMM (202mg, 2mmol) are dissolved in dichloromethane (40ml), add the CH of DCC (412mg, 2mmol) ₂cl ₂(10ml) solution, reactant stirs and spends the night under room temperature and argon atmospher.Remove by filter 1,3-Dicyclohexylurea precipitation, concentrating under reduced pressure filtrate.Ethyl acetate for residue (30ml) dilution, filters again, with 1M sodium bicarbonate, 10% citric acid and saturated nacl aqueous solution washing.Organic layer dried over mgso, vacuum concentration obtains (21) colorless oil, with silica gel chromatography, purifies, and uses ethyl acetate to make eluant, obtains 72% yield (710mg), is the mixture of diastereomer. ¹H-NMR(200MHz，CDCl ₃)；δ1.36，1.39(2s，9H，CH ₃)；1.87-3.2(m，14H，4CH ₃，CH ₃，NCH ₃)；4.40(m，2H，CH)；5.08，5.11(2s，2H，OCH ₂)；6.43，6.76，7.07(bs，-2H，NH)；6.76(2d，1H，NH)；7.31(m，15H，Ph-H).

Synthesizing of 3-thiopurine methyltransferase-4-oxo-5-azepine-6-(R)-methylamino formoxyl-8-carbonyl-sad (22) (FG 074): at tert-butoxycarbonyl amino ester 21 (495mg; 1mmol) in the solution in 15ml dichloromethane, add TFA (1.5ml), reactant mixture at room temperature stirs 1 hour under argon atmospher.Vacuum evaporating solvent and excessive acid, ether for residue (10ml) is processed for several times, concentrating under reduced pressure, obtain colorless oil, it is purified and obtains the white solid of intermediate, yield 84% (370mg) with silica gel chromatography (ethyl acetate/methanol 10: 1 contains 1% acetic acid).

At room temperature at intermediate (30mg, 0.07mmol), in the solution in 1ml (1.5/1) methanol/water, add LiOH.H ₂o (12mg, 0.27mmol).At room temperature stir after 3 hours, reactant mixture is processed with 0.5ml1N aqueous hydrochloric acid solution, by ethyl acetate (2 * 10ml), extracts.The organic layer salt water washing merging, by dried over mgso, concentrates and obtains sulphur compound 22, is colloidal solid (19mg, 0.06mmol) yield 86%.

MS (ES) (M+H) ⁺: 307

6.1.6. 2-[[N-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] acetic acid synthetic

Synthetic 2-[[N-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] acetic acid (23), also referred to as GF046, the method for optimizing of (embodiment of inhibitor A) is as follows: 2-[[N-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] acetic acid (23) synthetic: in the suspended mixture at ethyl ester (4) (300mg, 0.75mmol) in 1.5: 1 methanol/water (4ml), add Lithium hydrate/water.At room temperature stir after 5 hours, mixture is processed with 1N hydrochloric acid (20ml), with dichloromethane (2 * 20ml), extracts.The organic layer salt water washing merging, by dried over mgso, and concentrated.Thick solid recrystallization from hot ether obtains (23) white solid.

mp：139.5-140℃； ¹H?NMR(360MHz，DMSO-d6)δ7.79-7.08(m，4H，Ph)，4.37(s，2H，CH ₂)，3.85(s，3H，OCH ₃)，3.83(s，2H，CH ₂)

6.1.7. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(carboxymethyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(carboxymethyl)] amino] acetamide (30), also referred to as GF055, the preferred synthetic method of (embodiment of inhibitor A) is as follows:

2-[[N-(4-methoxybenzene sulfonyl)] amino] ethyl acetate (26) synthetic: to glycine ethyl ester hydrochloride (24) (3.0g; 21.5mmol) with 4-methoxybenzene sulfonyl chlorine (25) (4.4g; 21.3mmol) in the mixture in anhydrous methylene chloride (60ml), add triethylamine (4.79g, 47.3mmol).At room temperature stir after 15 hours, reactant mixture is processed with 1N hydrochloric acid (120ml), with dichloromethane (2 * 100ml), extracts.The organic layer salt water washing merging, by dried over mgso, and concentrated.Thick solid recrystallization from hot ethyl acetate/hexane obtains sulfonamide (26) white solid.

2-[[N-(4-methoxyphenyl sulfonyl)-N-tert-butoxycarbonyl methyl] amino] ethyl acetate (27) synthetic: in ice bath, to sodium hydride, (be dispersed in mineral oil; 60%) (162mg; 4.03mmol) in the slurry in anhydrous THF (10ml), add sulfonamide (26) (1.0g; 3.66mmol); then add bromoacetic acid tert-butyl group ethyl ester (785mg; 4.03mmol) mixture vigorous stirring 30 minutes at 0 ℃, then at room temperature stirs 15 hours.In ice bath, after reaction mixture, its water (25ml) is processed, with ether (2 * 25ml), extracted.The organic layer salt water washing merging, by dried over mgso concentrated.(2: 1 hexanes: ethyl acetate) purification, obtains (27) colourless slurry of silica gel chromatography for residue.

N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(tert-butoxycarbonyl methyl)] amino] acetamide (28) synthetic: under boiling point, prepare respectively hydroxy amine hydrochloric acid salt (377mg; 5.43mmol) solution in methanol (2.7ml) and KOH (456mg; 8.13mmol) the solution in methanol (2.7ml); be cooled to 40 ℃, then the latter solution added to the former.In ice bath, after cooling 30 minutes, filter out solid potassium chloride.In filtrate, add ethyl ester (27) (1.05g, 2.71mmol).At room temperature stir after 6 hours, reactant mixture is neutralized to pH4 with 1N hydrochloric acid solution, between dichloromethane (60ml) and water (20ml), distributes.It is biphase that separation obtains, and dichloromethane for water layer (60ml) extracts, the salt water washing of the organic layer of merging, and by dried over mgso, and concentrated.Silica gel chromatography for residue (11: 1 dichloromethane: methanol) purification obtains ester hydroxamic acid (28), also referred to as GF-058, white consubstantiality.

N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(carboxymethyl)] amino] acetamide (29) synthetic: by tertiary butyl ester hydroxamic acid (28) (520mg; mmol) solution in 35% trifluoroacetic acid/dichloromethane (9ml) stirs 10 minutes at 0 ℃, then at room temperature stirs 1.5 hours.Enriched mixture vacuum drying.Residue is developed in ethyl acetate, collects solid, and from ethyl acetate/methanol/hexane, recrystallization obtains sour hydroxamic acid (29) white solid.

Mp:160-161 ℃; MS (ES) (M+H) ⁺: 319; ¹h NMR (360MHz, DMSO-d6) δ 12.08 (brs, 1H, CO ₂h), 10.69 (s, 1H, OH), 8.96 (s, 1H, NH), 7.76 (d, J=8.7Hz, 2H, Ph), 7.09 (d, J=8.7Hz, 2H, Ph), 4.01 (s, 2H, CH ₂), 3.84 (s, 3H, OMe), 3.83 (s, 2H, CH ₂) .6.1.8. N-(4-methoxybenzene sulfonyl)-L-PROLINE hydroxamic acid synthetic

Synthetic N-(4-methoxybenzene sulfonyl)-L-PROLINE hydroxamic acid, also referred to as FG054, the method for optimizing of (embodiment of inhibitor A) is as follows:

Synthesizing of N-(4-methoxybenzene sulfonyl)-L-PROLINE methyl ester (32): to L-PROLINE methyl ester hydrochloride (30) (1.00g; 6.03mmol) with 4-methoxybenzene sulfonyl chlorine (31) (1.19g; 5.75mmol) in the solution in anhydrous methylene chloride (17ml), add triethylamine (1.22g, 12.06mmol).At room temperature stir after 15 hours, reactant mixture is processed with 1N hydrochloric acid (30ml) aqueous solution, by ethyl acetate (2 * 100ml), extracts.The organic layer salt water washing merging, by dried over mgso, concentrated obtain (32).This product is directly used in next reaction without being further purified.

Synthetic (33) of N-(4-methoxybenzene sulfonyl)-L-PROLINE hydroxamic acid: prepare respectively hydroxylamine hydrochloride (465mg under boiling point; 6.68mmol) solution in methanol (3.4ml) and KOH (561mg; 10.0mmol) the solution in methanol (3.4ml); be cooled to 40 ℃, then the latter solution added to the former.In ice bath, after cooling 30 minutes, filter out solid potassium chloride.In filtrate, add ethyl ester (32) (1.0g, 3.34mmol).At room temperature stir after 15 hours, 1N hydrochloric acid (40ml) solution-treated for reactant mixture, extracts by methylene chloride/methanol (10: 1).Organic layer salt water washing, by dried over mgso, and the concentrated crude product that obtains.Crude product recrystallization in hot methanol/ethyl acetate obtains hydroxamic acid (33) solid.

6.1.9. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-trifluoromethyl benzyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-trifluoromethyl benzyl)] amino] acetamide (34); also referred to as the method for optimizing of FG066 (embodiment of inhibitor A) substantially with described synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] method of acetamide is identical, but uses starting material 4-trifluoromethylated benzaldehyde to replace 4-chlorobenzaldehyde.Referring to, 6.1.1. part:

¹hNMR (360MHz, DMSO-d6) δ 10.50 (s, 1H, OH), 8.83 (s, 1H, NH), 7.81-7.08 (m, 8H, Ph), 4.45 (s, 2H, CH ₂cO), 3.86 (s, 3H, OMe), 3.67 (s, 2H, CH ₂ph). 6.1.10. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-methoxy-benzyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-methoxyphenyl sulfonyl)-N '-(4-methoxy-benzyl)] amino] acetamide (35); also referred to as the method for optimizing of FG067 (embodiment of inhibitor A) substantially with described synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N-(4-chlorobenzyl)] amino] method of acetamide is identical, but uses starting material 4-methoxybenzaldehyde to replace 4-chlorobenzaldehyde.Referring to, 6.1.1. part: ¹hNMR (360MHz, DMSO-d6) δ 10.42 (s, 1H, OH), 8.79 (s, 1H, NH) 7.81-6.86 (m, 8H, Ph), 4.29 (s, 2H, CH ₂cO), 3.85 (s, 3H, OMe), 3.73 (s, 3H, OMe), 3.28 (s, 2H, CH ₂ph). 6.1.11. N-hydroxyl-2-[[N '-(4-benzenesulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-benzenesulfonyl)-N '-(4-chlorobenzyl)] amino] acetamide (36); also referred to as the method for optimizing of FG-080 (embodiment of inhibitor A) substantially with described synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(4-chlorobenzyl)] amino] method of acetamide is identical, but uses starting material benzenesulfonyl chloro for 4-methoxybenzene sulfonyl chlorine.Referring to above-mentioned 6.1.1. part.

MS (ES) (M+H) ⁺: 355

36 (FG-080) 6.1.12. N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(benzyl)] amino] acetamide synthetic

Synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(benzyl)] amino] acetamide (37); also referred to as the method for optimizing of FG-061 (embodiment of inhibitor A) substantially with described synthetic N-hydroxyl-2-[[N '-(4-methoxybenzene sulfonyl)-N '-(carboxymethyl)] amino] method of acetamide is identical, but uses starting material benzyl bromide a-bromotoluene to replace bromoacetic acid tertiary butyl ester.Referring to above-mentioned 6.1.7. part.

MS (ES) (M-H) ^-: 349 synthetic N-(4-methoxybenzene sulfonyl)-β-benzyl-(the L)-aspartic acid (42) of 37 (FG-061) 6.1.13. N-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-aspartic acid; also referred to as FG084, the method for optimizing of (embodiment of inhibitor A) is as follows: FG084's (inhibitor A) is synthetic

N-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-aspartic acid (40).At room temperature; to β-benzyl-(L)-aspartic acid hydrochlorate (38) (2.00g; 8.96mmol) with to methoxybenzene sulfonyl chlorine (39) (1.76g; 8.53mmol) in the solution in anhydrous methylene chloride, add triethylamine (1.81g, 17.91mmol).Stir after 15 hours, reactant mixture is processed with 1N hydrochloric acid (60ml), by ethyl acetate (3 * 50ml), extracts.The organic layer salt water washing merging, by dried over mgso, concentrated N-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-aspartic acid (3.14g, 7.99mmol, 94% yield), the gluey product of obtaining.

N-benzyloxy-N '-(4-methoxyphenyl sulfonyl)-β-benzyl-(L)-agedoite (41).At room temperature; to N-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-aspartic acid (300mg; 0.76mmol) He in the mixture of O-benzyl hydroxylamine/hydrochloric acid in the anhydrous solution of (7/3) THF/DMF (10ml) add N-hydroxybenzotriazole (HOBT) (103mg; 0.76mmol), N-ethylmorpholine (204mg; 1.68mmol) with diisopropyl carbodiimides (106mg, 0.84mmol).After stirring a weekend (2.5), (1/1) hexane/ethyl acetate (40ml) dilution for reactant mixture, uses 1N hydrochloric acid (2 * 20ml), saturated sodium bicarbonate aqueous solution (2 * 20ml) and salt water washing successively.Dried over mgso concentrated for organic layer.Residue obtains N-benzyloxy-N '-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-agedoite (114mg through flash chromatography on silica gel method ((1/1) ethyl acetate/hexane) purification; 0.22mmol; 30% yield), white solid.mp：128-129℃；MS(ES)(M+S) ⁺：499

N-hydroxy-n '-(4-methoxybenzene sulfonyl)-(L)-agedoite (42).N-benzyloxy-N '-(4-methoxybenzene sulfonyl)-β-benzyl-(L)-agedoite (102mg, 0.20mmol) and the mixture of 10%Pd/C (43mg) in methanol (7ml) are descended to vigorous stirring 20 hours at hydrogen atmosphere (airbag pressure).With Celite pad, cross elimination catalyst, concentrated filtrate.Residue is lyophilizing in water, obtains N-hydroxy-n '-(4-methoxybenzene sulfonyl)-(L)-agedoite (50mg, 0.16mmol, 77% yield), flying upward property of moisture absorption powder.

1H NMR (360MHz, DMSO-d6) δ 10.60 (s, 1H), 8.78 (s, 1H), 7.87 (d, J=8.2Hz, 1H), 3.93 (m, 1H), 3.82 (s, 3H), 2.50 (m, 1H), 2.21 (dd, J=16.0,6.5Hz, 1H) .6.2. embodiment 2:C-proteinase activity test 6.2.1. is for measuring C-proteinase activity and the IC of inhibitor ₅₀in vitro tests

Following test can be used for measuring the different compounds of the present invention to the activity level of C-proteinase activity and effect.

By approximately 125 μ g radiolabeled ( ¹⁴c) to add cumulative volume be in the chicken C-protease in 0.1M Tris hydrochloric acid, 0.1M sodium chloride, 0.02%Brij-35 and 5mM calcium chloride solution of 10 μ l to tropocollagen.Reaction is carried out 15 minutes at 35 ℃, with the 3x termination/load buffer liquid (30mM EDTA, 30% glycerol, 6%SDS, 0.006% bromophenol blue) of half volume, stopped.Then sample is heated to 100 ℃ 4 minutes, use 6% polyacrylamide gel to differentiate by SDS-PAGE (Novex).By autoradiography, detect protein spectra.The disappearance of the amount of enzymatic activity based on corresponding to uncracked tropocollagen bands of a spectrum.

The IC of inhibitor ₅₀can be by drawing the curve of % activity to inhibitor concentration, and estimation produces the inhibitor concentration of 50% activity.

The IC of the inhibitor of test ₅₀value is shown in Table I.

Table I

The IC of the C-protease inhibitor that each is identified ₅₀

Inhibitor Generic organizes IC ₅₀

FG-047??????????????????????????A??????????50μM

FG-061??????????????????????????A?????????100μM

FG-053??????????????????????????A?????????100μM

FG-052 ¹????????????????????????A?????????125μM

FG-066??????????????????????????A?????????150μM

FG-067 A 150 μ MFG-086 (cbz-Pro-Leu-Gly-hydroxamic acid ²) A 200 μ M FG-087 (Ac-PYYG-hydroxamic acid) A 335 μ M

FG-088 (radiation amide element ³) A 350 μ M

FG-054??????????????????????????A?????????400μM

FG-057??????????????????????????A????????1500μM

FG-058 ⁴????????????????????????A????????2100μM

FG-055??????????????????????????A????????2600μM

FG-051 ⁵????????????????????????A??＞＞???100μM

FG-046 A > > 1000 μ M1 are commercial is obtained by Peptides International that (IHN-3850-PI) 2 is commercial is obtained by Sigma that (C-8537) 3 is commercial obtains (A-6671) 4 midbody compounds (28) by Sigma, referring to 6.1.7 part 5 is commercial, obtains (ISN-3835-PI) 6.2.2. be used for measuring inhibitor C-proteinase activity and IC by Peptides International ₅₀external ELISA test

The IC of inhibitor ₅₀value can also be by filtering ELISA test determination.In this test, the unlabelled human body tropocollagen of about 25ng I partly cultivates 1 hour as 6.2.1 with C-protease.By adding 40 μ l precipitation buffering liquid (0.5X reaction buffer, 0.1mg/ml chicken collagen protein H, 10 μ g/mlBAS, 7.5mM EDTA) cessation reaction.Add 25 μ l75% ethanol, hybrid reaction mixture, cultivates 1 hour with precipitinogen collagen protein on ice., use Milliporemultiscreen vacuum valve, by Millipore multiscreen-HV 0.45 μ m hydrophilic plate, filter the c-propetide of separate dissolved in the collagen protein precipitating.Remove the filtrate of 20 μ l, by using the amount of C-propetide of tropocollagen I type C-peptide (PIP) the EIA kit measurement cracking of Takara Biomedicals.

For suppressing hBMP-1, by about 20ng radiolabeled ( ¹²⁵i) human body tropocollagen I is added in the 5 times of concentrated restructuring hBMP-1 cell culture mediums of 1-2 μ l (Kessler etc., (1996) science 271:360) in the reaction solution of cumulative volume 10 μ l.Reaction is carried out 1 hour at 35 ℃, with half volume 3x termination/load buffer liquid, stopped subsequently, with SDS-PAGE, analyze as mentioned above.

The IC of inhibitor ₅₀can be by drawing the curve of % activity to inhibitor concentration, and estimation produces the inhibitor concentration of 50% activity.IC ₅₀value is shown in Table II.

Table II

The IC of the C-protease inhibitor that each that measured by ELISA identified ₅₀inhibitor Generic organizes IC ₅₀eLISAFG-061 A 12 μ MFG-047 A 13 μ MFG-053 A 22 μ MFG-067 A 37 μ MFG-066 A 48 μ M6.2.3. are used for measuring inhibitor C-proteinase activity and IC ₅₀tissue culture test

C-proteinase activity and IC in the body of inhibitor ₅₀in Zhi Ke tissue culture test, by being determined at the generation of tropocollagen and ripe collagen protein in conditioned medium of compound treatment front and back, measure.The ratio of collagen protein and tropocollagen is by directly relevant to the cell transformation of ripe collagen protein product with precursor, and it represents C-proteinase activity.

In addition, can measure the culture medium content of C-propetide/cell, and the cell of more untreated cell and inhibitor processing.6.3. for measuring the animal model of inhibitor C-proteinase activity and effect

Simulation is in the prior art known with some animal models that irregular or inappropriate collagen protein produces relevant clinical disease, can be used for measuring the interior effect of body of compound of the present invention.These material models comprise rat wound chamber (chamber) model (Schilling etc., 1959, surgery 46:702-210), the metreurysma model (Mandell etc. that male glycol stimulates, 1982, journal of biological chemistry 257:5268-5273) and induction angiogenesis model (Matrigel) (Passaniti etc., 1992, experimentation 67:519-528).Other animal model comprises clinical disease model, as hepatic fibrosis model (Tsukamoto etc., 1990, hepatopathy seminar 10:56-65; Kock-Weser, 1952, experimentation 1:324-331; Marrione, 1949, U.S. pathology magazine 25:273-285; Tams, 1957, U.S. pathology magazine 33:13-27; Wahl etc., 1986, The Journal of Experimental Medicine 163:884-902), pnemnofibrosis model (Kelly etc., 1980, experiment Clinical Medical Journals 96:954-964), arterial restenosis model (Jackson, 1994, Trends of Cardiovascular Medicine4:122-130; Clowes etc., 1983, experimentation 49:327-333), kidney fibre modification model (Yamamoto etc., 1987, Kidney International32:514-525), tendon recovers model (Fanklin etc., 1986, experiment and Clinical Medical Journals 108:103-108), tumor growth model (kiohs etc., 1985, JNCL75:353-359), trabeculectomy model (Lahery etc., 1989, pathology of eye research 5:155-179) and abdomen adhesion model (Williams etc., 1992, surgery research magazine 52:65-70).6.4. embodiment 4: cytotoxic assay

The cytotoxicity of research potential inhibitor is to measure them whether to cells survival or breed influential.These tests comprise use the Fast-propagation phase or resting stage cell.Inoculate datum object cell, and with the time increasing, be exposed to the concentration range of potential inhibitor.For example, by cell counting or staining (crystal violet), measure cell number.

Cytotoxicity is as the function evaluation of cells survival and cell proliferation.Cells survival comprises use cell resting stage, and measures by cell number (counting or staining).The reduction of cell number represents loss cell, therefore, influential to cells survival.Cell proliferation comprises uses Fast-propagation phase cell, also by cell number, is measured.Now, the reduction with respect to untreated cell number represents cell proliferation influential.

The present invention is not restricted to the scope of embodiment explanation, and embodiment is for the explanation of the single aspect of the present invention, and any compound that function is suitable and their using method are within the scope of the invention.In fact, except above-mentioned, by following description and accompanying drawing, various modifications of the present invention it will be apparent to those skilled in the art that.This modification also within the scope of the appended claims.

It classifies list of references herein in full as all lists of references of listing.

Claims

1. have the compound of C-protease inhibition and its pharmaceutically useful salt, described compound has and is selected from following structural formula: or

Wherein

R ₂be selected from H, low alkyl group;

X is selected from SO ₂, C=O;

Y is selected from OH, HOHN (azanol), H ₂n, alkyl amino;

Z is direct bond, methylene, oxygen, sulfur, amino;

N is 0 or 1.

2. have the compound of C-protease inhibition and its pharmaceutically useful salt, described compound has and is selected from following structural formula:

Wherein

R ₄be selected from H, low alkyl group; With

3. have the compound of C-protease inhibition and its pharmaceutically useful salt, described compound has and is selected from following structural formula:

Wherein

R ₄be selected from H, low alkyl group; With

4. have the compound of C-protease inhibition and its pharmaceutically useful salt, described compound has and is selected from following structural formula:

Wherein

R ₄select white H, low alkyl group;

5. have the compound of C-protease inhibition and its pharmaceutically useful salt, described compound has and is selected from following structural formula:

Wherein

R ₁be selected from OH, alkoxyl, low alkyl group, alkyl amino, peptide;

X is selected from N, C;

R ₄be selected from H, low alkyl group;

6. the compound of claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

7. the compound of claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

8. the compound of claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

9. the compound of claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

10. the compound of claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 11. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 12. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 13. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 14. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 15. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 16. claim 1 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 17. claim 3 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

The compound of 18. claim 3 and its pharmaceutically useful salt, wherein above-claimed cpd has following formula:

19. pharmaceutical compositions, it contains and is selected from following compound: or

Wherein

R ₂be selected from H, low alkyl group;

R ₄be selected from aryl, heteroaryl, alkyl, aralkyl, heteroarylalkyl, alkyl amino, aryl alkyl amino;

X is selected from SO ₂, C=O;

Y is selected from OH, HOHN (azanol), H ₂the group of N, alkyl amino;

Z is direct bond, methylene, oxygen, sulfur, amino;

N is 0 or 1;

With pharmaceutically useful carrier or excipient;

Or:

Wherein

R ₄be selected from H, low alkyl group; With

With pharmaceutically useful carrier or excipient; Or:

Wherein

R ₄be selected from H, low alkyl group; With

With pharmaceutically useful carrier or excipient;

Or:

Wherein

R ₄be selected from H, low alkyl group;

With pharmaceutically useful carrier or excipient; Or:

Wherein

R ₁be selected from OH, alkoxyl, low alkyl group, alkyl amino, peptide;

X is selected from N, C;

R ₄be selected from H, low alkyl group;

With pharmaceutically useful carrier or excipient.

20. pharmaceutical compositions, it contains and is selected from following compound: with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or: with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or: with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or:

with pharmaceutically useful carrier or excipient;

Or:

With pharmaceutically useful carrier or excipient.

The method of the disease that 21. treatments are relevant with the inappropriate or irregular generation of collagen protein, it comprises the compositions of the claim 20 of using effective dose.

The method of the disease that 22. treatments are relevant with the inappropriate or irregular generation of collagen protein, it comprises the compositions of the claim 21 of using effective dose.

23. the method for claim 22, wherein fibrotic conditions is selected from hepatitis interstitialis chronica and arthritis.

24. the method for claim 23, wherein fibrotic conditions is selected from hepatitis interstitialis chronica and arthritis.