CN101434631B - Estra nitrate ester medicament for inhibiting angiogenesis - Google Patents

Abstract

The invention discloses a compound 1 or salts or ester thereof with a general formula as follows: T-(B-O-NO2)t1, wherein, t1 refers to 1 or 2; T-H refers to a steroidal compound; T refers to steroidal residue obtained from the T-H compound, H of which is removed; oxygen atoms on T are connected with H to form hydroxyl groups, namely, T is connected with H in the form of O-H.

Description

A kind of estra nitrate ester medicine of angiogenesis inhibiting

Technical field:

The invention belongs to field of medicaments, particularly formula (1) compound and compound method thereof.The present invention also relates to purposes and the medicinal prepns of this compound on medicine, particularly be directed against angiogenesis disease such as tumour, the newborn treatment of diseases of ocular angiogenesis.

Background technology:

Angiogenesis comprises two notions: brephic blood vessel take place (vasculogenesls, VG) with postnatal vasculogenesis (anglogenesls, AG).VG refers to not to be had under the situation of vascular system, by endothelial progenitor cell (endothellalprogenltor, cells, EPCs) or angioplast (angloblasts) be divided into endotheliocyte, and form vasoganglion.AG refers at the adult blood vessel, breaks through the migration of tube wall matrix by the mature endothelial cell that has existed, and propagation and reconstruct make the blood vessel branch continue to prolong with the germination mode.Angiogenesis among the present invention be meant postnatal vasculogenesis (anglogenesls, AG).Angiogenesis (anglogenesls) process that (like growth, wound healing) institute must have during still normal physiological changes, scientists finds that also the development of numerous diseases such as it and tumour, senile macular degeneration SMD, malignant hematologic disease has confidential relation in recent years.Suppress pathologic angiogenesis, can treat or slow down numerous diseases such as tumour, senile macular degeneration SMD, malignant hematologic disease.

The method of clinical treatment tumour is to adopt diverse ways to different tumours mostly at present, and the overwhelming majority is to treat to tumour cell.And tumor growth is the process of a complicacy, and it receives influence of various factors, comprising the foundation of tumor vessel net.Many researchs have proved that tumor growth must rely on vasculogenesis, through suppressing some link or the whole process of vasculogenesis, and then the control growth of tumor, to oncotherapy with prevent that the tumour metastasis is significant.Depend on the proposition of vascularization notion the beginning of the seventies along with tumor growth; The also corresponding notion that has proposed anti-angiogenic formation treatment; I.e. generation and/or the expansion of new vessel net and/or generation or the foundation that the destruction new vessel stops little solid tumor through stoping new vessel is to stop tumor growth, development and transfer.The more heparin of external report adds the hydrogenation KE. and be acknowledged as and can suppress vasculogenesis effectively.Experiment confirm, its two combined utilization can suppress the generation of blood vessel on the chick chorioallantoic membrane, and well can make tumor regression shift with stoping, and also can suppress the rabbit corneal angiogenesis that tumour causes.

And VEGF (VEGF) is one type of multifunctional growth factor, has the effect that promotes that endothelial cell proliferation, induction of vascular form, and it is generally acknowledged that suppressing VEGF (VEGF) can treat or slow down pathological.

Diseases such as VEGF (VEGF) and new vessel diseases associated such as tumour, eye neovascular disease, malignant hematologic disease, bronchial asthma have substantial connection, through suppressing VEGF (VEGF), can treat or slow down these diseases; Lot of documents all has the report of this respect like " constitutional features and the biological function of summary VEGF " (Chinese clinic study magazine; 2007 13 3 phases of volume, 388), " VEGF and acceptor thereof the progress in gynaecopathia " (2006 28 12 phases of volume of Hebei medicine, 1192-1194), " VEGF and tumour concern progress " (Guangxi Medical University's journal; 2006 23 2 phases of volume; 333-335), the present Research of antineoplaston " VEGF relevant " (Jilin medical science, 2006 27 5 phases of volume, 454-457), " progress of target VEGF treatment malignant tumour " (modern tumour medical science; 2006 14 3 phases of volume; 370-372), " VEGF and PEDF are to the common regulating effect of eyeground new vessel " (foreign medical science: clinical biochemistry and ecsomatics fascicle, 2005 26 11 phases of volume, 819-821), " eyeground new vessel control progress " (ophthalmology new development; 2000 20 6 phases of volume; 449-451), " VEGF and acceptor thereof and intraocular neovascularization property disease " (ophthalmology research, 2003 21 1 phases of volume, 103-106), " relation of VEGF and malignant hematologic disease " (foreign medical science: physiological and pathological science and clinical fascicle; 2004 24 2 phases of volume; 183-185), the research of patients serum VEGF level " alba loose " (the difficult disease magazine, 2007 61 phases of volume, 10-11), " VEGF and bronchial asthma " (practical medical journal; 2007 the 23rd the 3rd phases of volume, 433).

The known formation (vasculogenesis or new vessel form) that has multiple medicine can suppress neovascularity.For example; At people such as Crum " one type of new steroid suppresses vasculogenesis in the presence of heparin or heparin fragment " (A New Class of steroldslnhlblts Anglogenesls ln the Presence Of Heparln or a Heparln Fragment; Sclence; Vol.230:1375-1378, on December 20th, 1985) in the literary composition, the steroid that can in the presence of heparin or specific heparin fragment, suppress vasculogenesis is disclosed.The author is called said steroid " blood vessel suppresses (anglostatlc) " steroid.This type of comes to light and has dihydro and tetrahydro metabolites that KE and deoxidation KE are arranged included in the inhibiting steroid of blood vessel.In the follow-up study of test about the hypothesis of said mechanism, prove: heparin/angiostatic steroid compsn causes the membrane holder dissolving, on said support, is connected with the anchorage dependence endothelium, thereby causes kapillary atrophy (lnvolutlon); Wherein said mechanism is that steroid suppresses the mechanism of vasculogenesis through it; " the possible mechanism that suppresses vasculogenesis through angiostatic steroid: kapillary basilar membrane dissolved is induced " (A Posslble Mechanlsm forlnhlbltlon of Anglogenesls by Anglostatlc Sterolds:lnductlon of Caplllary BasementMembrane Dlssolutlon referring to people such as lngber; Endocrlnology Vol.119:1768-1775,1986).

U.S. Pat 4 people such as Arlstoff; One group of tetrahydro steroids that can be used for suppressing vasculogenesis is disclosed in 975,537. this patent discloses said compound and can be used for treating head trauma, spinal trauma, septic shock or traumatic shock, apoplexy and hemorrhagic shock.In addition, this patent has also been discussed these compounds and has been implanted and treated the effect in cancer, sacroiliitis and the arteriosclerosis the embryo.At USP us4771, disclose in people's such as Arlstoff the patent disclosed some steroid and heparin or heparin fragment in 042 and made up the vasculogenesis that suppresses warm-blooded animal.People such as L1; " angiostatic steroid that strengthens effectiveness through sulfated cyclodextrin suppresses cornea rebirth blood vessel formation " (Anglostatlc SteroldsPotentlated by Sulphated Cyclodextrln lnhlblt Corneal Neovascularlzatlon; Lnvestlgatlye ophthalmology and Vlsual Sclence; Vol 32 (11): 2898-2905, in October, 1991).The independent use of steroid can make new vessel formation how much alleviate, and new vessel forms but only independent use can not effectively be disappeared.

Urocortisone is as angiostatic steroid; " angiostatic steroid " (Anglostatlc sterolds people such as Folkman; Ann.Surg, Vol.206 (3), 1987) open in the literary composition; Wherein the document proposes angiostatic steroid and possibly can be used for treating by new vessel and form the disease of being controlled unusually, comprises diabetic retinopathy, neovascular glaucoma and retrolental fibroplasia.

Having introduced anecortave acetate among the CN03818826 is that a kind of exploitation is used to suppress the vasoinhibitor that the eyes neovascularity generates.Preparation and method that this invention relates to and is used to prevent AMD dependency visual loss, keeps AMD patient's eyesight and suppresses AMD dependency infringement development.Said preparation and method relate to the sclera side dressing with the anecortave acetate of 3-30mg or its corresponding alcohol so that trans-scleral drug release to be provided.

Summary of the invention:

A kind of compound 1 of following general formula or its ester or salt:

T-(B-O-NO2) _T1, t1 is 1 or 2, and T-H is a steroidal compounds, and T removes the steroidal residue of H for compound T-H, and T is to link to each other with H with the Sauerstoffatom on the T with H, the form that forms hydroxyl and be O-H links to each other.

B links to each other with the ester bond form with t1 hydroxyl on the TH; Be t1 on the TH substituent-O among OH respectively with t1 B on-the C=O-formation-O-CO-that links to each other; T1 is 2 o'clock; Two last hydroxyls of TH link to each other with two-B-O-NO2 respectively, and the B in this two-B-O-NO2 group can be identical or different in the scope of its definition.

Can there be substituting group in H or the H2 in the CH2 base in the CH base shown in the replacement general formula of TH, the substituting group that also can exist, and the substituting group of each position can be:

At the 3rd: can be OR3, R3 be H or ten carbon with interior hydrocarbon polymer or contain 1-2 heteroatomic ten carbon with interior hydrocarbon polymer, heteroatoms is N, O, one or both among the S;

At 9,11: can be two keys

At 11: can be hydroxyl

At 17: can be ketone group, OH or O-CO-R, R be ten carbon with interior hydrocarbon polymer or contains 1-2 heteroatomic ten carbon with interior hydrocarbon polymer that heteroatoms is N, O, one or both among the S

R1 can or have the straight or branched alkyl of 1 to 4 carbon atom for hydrogen

X can be O, a kind of among the S or do not exist

R2 can not exist or for H, ten carbon is with interior hydrocarbon polymer or contain 1-2 heteroatomic ten carbon with interior hydrocarbon polymer, heteroatoms is N, O, one or both among the S

B is-CO (O) _a(CR4R5) _{B '}D (CR4R5) _{B "}-

A is 0 or 1, b ', b " can be identical or different, and be 0 to 6 integer, R4; R5 is identical or different, and be selected from H, the hydrocarbon polymer of 1 to 6 carbon, D can not exist also can be for containing the hydrocarbon polymer of 0 to two heteroatomic 1 to 8 carbon; heteroatoms is N, O, one or both among the S

Arbitrary R4 substituting group described in the B can be identical or different in the scope of its definition.

Arbitrary R5 substituting group described in the B can be identical or different in the scope of its definition.

Preferably: R1 is a methyl.

Preferably: 17 of TH is OH

Preferably: X can be O

Preferably: 9,11 of TH is two keys.

Preferably: t1 is 1 o'clock, and last 17 of TH are hydroxyl, links to each other with-B-O-NO2, and B links to each other with the ester bond form with hydroxyl on the T, promptly on the TH-O among the OH respectively with B on-the C=O-formation-OCO-that links to each other,

T1 is 1 o'clock a compound 1

Preferably: t1 is 2 o'clock; Last 17 of TH are hydroxyl; Link to each other with one-B-O-NO2, last another hydroxyl of T with another-B-O-NO2 links to each other, B respectively with T on hydroxyl link to each other with the ester bond form; Be on the TH-O among the OH respectively with B on-the C=O-formation-OCO-that links to each other, the B in this two-B-O-NO2 group can be identical or different in the scope of its definition.

Preferably: R2 is that ten carbon are with interior hydrocarbon polymer

Preferably: R2 is a methyl

Preferably: R3 is H

Preferably: t1 is 2 o'clock; Last 17 of TH are hydroxyl, link to each other with one-B-O-NO2, and last 3 of TH is a hydroxyl; With another-B-O-NO2 links to each other; B respectively with TH on hydroxyl link to each other with the ester bond form, promptly on the TH-O among the OH respectively with B on-the C=O-formation-OCO-that links to each other, the B in this two-B-O-NO2 group can be identical or different in the scope of its definition.

Preferably: X is O, and R2 is a methyl, and R3 is H, and last 17 of TH are hydroxyl.

Preferably: X is O, and R2 is a methyl, and R3 is H, and last 17 of TH are hydroxyl, and 9,11 of TH is two keys.

Preferably: B is-CO (O) _a(CR4R5) _{B '}D (CR4R5) _{B "}-

A is 0 o'clock, R4, and R5 is identical or different, and is selected from H, and the straight or branched hydro carbons of 1 to 6 carbon, D can not exist also can be for containing the hydrocarbon of 0 to 2 heteroatomic 1 to 8 carbon.

Preferably: B is-CO (O) _a(CR4R5) _{B '}D (CR4R5) _{B "}-

A is 1 o'clock, R4, and R5 is identical or different, and is selected from H, and the hydrocarbon polymer of 1 to 6 carbon, D can not exist also can be for containing five yuan or six-ring hydrocarbon polymer of 0 to 2 heteroatomic 1 to 8 carbon.

Preferably: B is-CO (O) _a(CR4R5) _{B '}D (CR4R5) _{B "}-

A is 0 or 1 o'clock, b ', " b is 0, D is five yuan or the hexa-atomic cyclic hydrocarbon that contains 0 to two heteroatomic 1 to 8 carbon, heteroatoms is on ring.

Preferably: B is-CO (O) _a(CR4R5) _{B '}D (CR4R5) _{B "}-

A is 0 o'clock, and D can not exist also can be for containing five yuan or hexa-atomic cyclic hydrocarbon of 1 to two heteroatomic 1 to 8 carbon, and heteroatoms is on ring.

Formula 1 compound is preferably:

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-nitroxide) propionic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-methyl-4-nitroxide)-2-ester thiohenic acid

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-3-(2-nitroxide) acetic ester

3-methoxyl group female steroid-1,3,5 (10)-triolefins-17 β-(4-nitroxymethyl) benzoic ether

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester-17-(3-nitroxide) propionic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-oxyethyl group-17 β hydroxyl-17-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-oxyethyl group-17-ketone-3-(2-nitroxide) acetic ester

Acceptable salt or solvate are as the application in the medicine of treatment mammalian diseases on formula 1 compound or its physiology, and Mammals is preferably human.

For suppressing the medicine of mammal VEGF, Mammals is preferably human as the medicine of treatment mammalian diseases for acceptable salt or solvate on formula 1 compound or its physiology.

Acceptable salt or solvate are the medicine of angiogenesis inhibitor as the medicine of treatment mammalian diseases on formula 1 compound or its physiology, and Mammals is preferably human.

Above-mentioned each compound is in the application of preparation treatment tumour class disease medicament.

Above-mentioned each compound in the medicine of preparation treatment ophthalmology eye neovascular disease application.

Above-mentioned each the application of compound in the medicine of preparation treatment malignant hematologic disease.

Above-mentioned each the application of compound in the medicine of preparation treatment bronchial asthma.

Above-mentioned each the application of compound in the medicine of the loose disease of preparation treatment alba.

A kind of medicinal compsns, said composition comprise acceptable salt or solvate on formula 1 compound or its physiology of above-mentioned each definition.On formula 1 compound of above-mentioned each definition or its physiology on acceptable salt or solvate and one or more physiology the acceptable drug auxiliary material mix.Above-mentioned medicinal compsns can be mixed with liquid preparation, sterilization preparation and sterile preparation, solid preparation, semi-solid preparation, aerosol.

Above-mentioned medicinal compsns, said composition also comprise the medicine of another kind of treatment tumour.

Above-mentioned compsn also comprises the medicine of another kind of treatment eye neovascular disease.

Above-mentioned compsn also comprises the medicine of another kind of treatment malignant hematologic disease disease.

Above-mentioned compsn also comprises the medicine of another kind of treatment bronchial asthma.

A kind of ophthalmic preparation, said preparation contain acceptable salt or solvate on formula 1 compound or its physiology of above-mentioned each definition, and as the pharmaceutical excipient of carrier.

A kind of oral prepns, said preparation contain acceptable salt or solvate on formula 1 compound or its physiology of above-mentioned each definition, and as the pharmaceutical excipient of carrier.

Medicinal compsns, said composition also comprise the medicine of another kind of treatment tumour.

A kind of above-mentioned formula 1 compound method for preparing, it comprises:

(1) TH is formed TBY with the acid anhydrides YBOBY of YBOH, carboxyl acyl chloride YBCl or carboxylic acylbromide YBBr reaction in the presence of alkaline organic solvent; Y is bromine or chlorine atom, then

(2) the product TBY that obtains in the above-mentioned reaction is reacted in organic solvent through the inorganic salt with nitric acid obtain T-B-O-NO2.

A kind ofly prepare the formula 1 compound method that above-mentioned B links to each other with 3 hydroxyls of TH, it comprises:

(1) TH is formed TBY with the acid anhydrides YBOBY of YBOH, carboxyl acyl chloride YBCl or carboxylic acylbromide YBBr reaction in the presence of alkaline organic solvent; Catalyzer can exist or not exist, and Y is bromine or chlorine atom, then

(2) the product TBY that obtains in the above-mentioned reaction is reacted in organic solvent through the inorganic salt with nitric acid obtain T-B-O-NO2.When being hydroxyl in 17 bit substituents of TH, can be earlier carry out the esterification protection to corresponding hydroxyl, form 3 again and slough the ester group protection after replacing nitric ethers, produce compound 1.

A kind ofly prepare the formula 1 compound method that above-mentioned B links to each other with 17 the hydroxyl of TH, it comprises:

(1) TH is formed TBY with the acid anhydrides YBOBY of YBOH, carboxyl acyl chloride YBCl or carboxylic acylbromide YBBr reaction in the presence of alkaline organic solvent;

Y is bromine or chlorine atom, then

(2) the product TBY that obtains in the above-mentioned reaction is reacted in organic solvent through the inorganic salt with nitric acid obtain T-B-O-NO2.

Above-mentioned preparation method is characterized in that the alkaline organic solvent in the reaction of (1) step is trialkylamine or pyridine etc., and catalyzer is DMAP, and the inorganic salt of the nitric acid in the reaction of (2) step are Silver Nitrate, and organic solvent is an acetonitrile.

Above-mentioned t1 is 2 formula 1 compound, and identical or different B links to each other with 3 hydroxyl with 17 of TH respectively, and it comprises:

(1) TH is formed TB ' Y ' with the acid anhydrides Y ' B ' OB ' Y ' of YBOH, carboxyl acyl chloride Y ' B ' Cl or carboxylic acylbromide Y ' B ' Br reaction in the presence of alkaline organic solvent; Catalyzer can exist or not exist, and Y ' is bromine or chlorine atom, and wherein B ' links to each other with hydroxyl on 17 bit substituents, then

(2) with above-claimed cpd TBY in the presence of alkaline organic solvent and Y " B " the acid anhydrides Y of OH " B " OB " Y ", carboxyl acyl chloride Y " B " Cl or carboxylic acylbromide Y " B " the Br reaction forms Y " B " TB ' Y '; Catalyzer can exist or not exist, Y " be bromine or chlorine atom, wherein B " link to each other with hydroxyl on 3 bit substituents, then

(3) with the product Y ' B ' TB that obtains in the above-mentioned reaction " Y " react in organic solvent through inorganic salt with nitric acid obtain O2N-O-B ' TB "-O-NO2, B ", the same B of the range of definition of B '.

B in wherein above-mentioned (1) to (3) steps ", B ' can be identical or different.

Above-mentioned preparation method is characterized in that (1), the alkaline organic solvent in the reaction of (2) step is trialkylamine or pyridine etc., and catalyzer is DMAP, and the inorganic salt of the nitric acid in the reaction of (3) step are Silver Nitrate, and organic solvent is an acetonitrile.

Neovascularization disease relates generally to tumour, eye new vessel, malignant hematologic disease, bronchial asthma, the loose disease of alba.

Tumour mainly is various noumenal tumours such as tumor stomach, lung tumors, liver's tumour, various body of gland tumour, nose tumour, ocular tumor, pharyngeal tumour, laryngeal neoplasm etc.; Malignant hematologic disease is meant the malignant tumour of blood system, mainly comprises white blood disease, lymphoma, multiple myeloma and malignant histocytosis etc.

This compound also can generate in the disease that causes at the eye new vessel and use:

The eye new vessel generates the disease cause and relates to cornea, iris, choroid and retina etc., and pathological changes such as it causes oozes out, hemorrhage and hyperplasia are the major reasons that causes visual disorder to the infringement of eye structure and function.All there are pathologic processes such as inflammation, ischemic, anoxic in most of ophthalmic; Therefore, illness in eye and eye new vessel are formed with confidential relation.Wherein the eye new vessel generates the disease cause and comprises:

Cornea rebirth blood vessel property illness in eye: in the eye disease relevant with cornea rebirth blood vessel; Modal is cornea rebirth blood vessel property disease due to the wearing of contact lens; Other cause that the eye disease of cornea rebirth blood vessel has the cornea wound; Alkali and other chemical substances burn, operation on cornea, various infection comprise infectation of bacteria, choamydiae infection, virus infection (herpes simplex virus and varicella zoster virus etc.), protozoan infection (leishmania) etc.

Iris neovascular illness in eye: common have neovascular glaucoma, and retinal detachment, wound, diabetic retinopathy, tumour (retinoblastoma), central vein of retina embolism etc. are its common inducements.

Retinal neovascularization property illness in eye: mellitus, tumour, retinal detachment, central retinal vein occlusion, periphlebitis of retina, systemic lupus erythematosus, Eales are sick, Coat is sick, Takavas disease etc. all can cause.

CNV property illness in eye: senile macular degeneration SMD, high myopia, the exudative retinochoroiditis of centrality, wound, tumour, ocular histoplasmosis's syndrome, serpiginous choroidopathy etc. all can cause this kind illness in eye.

Those skilled in the art can think, related " treatment " of this paper can extend to the prevention of disease and to confirm that disease is the treatment of purpose.

The dosage of formula (1) compound in the mammiferous disease of treatment is counted 0.001mg-5mg/kg/ days, preferred 0.005mg-2mg/kg/ days with TH's.Formula (1) compound suppresses the mammal VEGF in treatment drug dose is counted 0.001mg-5mg/kg/ days, preferred 0.005mg-2mg/kg/ days with TH.Formula (1) compound is counted 0.001mg-5mg/kg/ days, preferred 0.005mg-2mg/kg/ days at the dosage of treatment mammal neovascularization disease medicine with TH.During as treatment people's disease, people's body weight is generally calculated according to 50Kg, so the dosage of human described in the present invention can calculate according to people's ABW, also can calculate according to above-mentioned dosage X50.

The compounds of this invention can be mixed with any preparation of being convenient to medication, so the present invention also comprises The compounds of this invention in its scope, can with acceptable diluent on one or more physiology or carrier blended pharmaceutical composition.

The compounds of this invention can be mixed with the preparation of any people's of being convenient to medication, and its absorption of active ingredient is for counting 0.001mg-5mg/kg/ days, preferred 0.005mg-2mg/kg/ days with TH in the said preparation treatment disease.During as treatment people's disease, people's body weight is generally calculated according to 50Kg, so the consumption of people described in the present invention can calculate according to people's ABW, also can calculate according to above-mentioned consumption X50.

A kind of medicinal compsns, said composition comprise acceptable salt or solvate on formula 1 compound or its physiology of each definition.Above-mentioned medicinal compsns comprises on formula 1 compound or its physiology that the acceptable drug auxiliary material mixes on acceptable salt or solvate and one or more physiology.Above-mentioned medicinal compsns can be mixed with liquid preparation, sterilization preparation and sterile preparation, solid preparation, semi-solid preparation, aerosol; Above-mentioned preparation type can be according to pharmaceutics (the 5th edition; People's Health Publisher, Cui Fude chief editor) related definition in is understood.

Acceptable salt or solvate can be used as the medicine that treatment eye new vessel generates the disease that causes on formula 1 compound or its physiology.

A kind of injection formulations, said preparation contain acceptable salt or solvate on formula 1 compound or its physiology of each definition among the claim 1-2, and as the pharmaceutical excipient of carrier.

Non-active ingredient in the injection formulations contains water for injection or oil for injection.

A kind of oral prepns, said preparation contain acceptable salt or solvate on formula 1 compound or its physiology, and as the pharmaceutical excipient of carrier.This oral prepns can be according to the method preparation of the related preparations in the pharmaceutics (the 5th edition, People's Health Publisher, Cui Fude chief editor).

The present invention further provides the method for the such pharmaceutical composition of preparation, and this method comprises mixes various components.

The compounds of this invention can be pacified the preparation that ordinary method (for example) is mixed with oral, oral cavity, hypogloeeis, non-enteron aisle, injection, heeling-in, local application or rectal administration, the preparation of especially oral or injection.Wherein oral prepns comprises and is not limited only to the liquid of taking orally such as oral solid such as tablet, capsule, powder, granule and oral liquid; Injection comprises and is not limited only to the solid that injectable liquid such as injection, suspensoid, infusion solution or powder injection etc. can be used for injecting.

Non-active ingredient in the oral prepns contains one or more in starch, lactose, the water.

Compound of the present invention and pharmaceutical prepn can be selected from following other treatment agent with one or more and unite use or comprise one or more such therapeutical agents: the medicine, the treatment eye new vessel that suppress tumour generate the medicine of the disease that causes, the medicine of treatment asthma.

The medicine that suppresses tumour includes, without being limited to act on the medicine (comprising alkylating agent, anthracene nucleus class and platinum compound) of DNA chemical structure; Influence nucleic acid synthetic medicine (mainly being metabolic antagonist); Acting on dna profiling influences DNA and transcribes or suppress DNA dependenc RNA polysaccharase and suppress RNA synthetic medicine; Influence the medicine (like percephalotaxine, Japanese yew class, vincaleucoblastine and Podophyllum emodi var chinense bases etc.) of protein synthesis; Paclitaxel analog compound; The medicine of other types (like hormone, Asparaginase, R8923 etc.).

The preferred cis-platinum of medicine that suppresses tumour.

The medicine that treatment eye new vessel generates the disease that causes includes, without being limited to Endostatin (Endostatln), Angiostatin, anecortave etc.

The medicine of treatment asthma comprises and is not limited only to glucocorticosteroid, leukotriene inhibitors, broxaterol, xanthine drug, anticholinergic drug, anti-allergy agent.

The salt of formula (1) compound is meant that the OH on the TH is attached thereto the pharmaceutical salts that connects formation, and pharmaceutical salts preferably phosphoric acid, succsinic acid, toxilic acid, vitriolic salt more preferably refer to phosphoric acid, succsinic acid, toxilic acid, vitriolic sodium, potassium, magnesium, calcium salt.

The method for making of compound TH can be referring to one Chinese patent application 200710058413.5, the method in " a kind of female steroid medicine of treating tumour ".

DMAP is the 4-Dimethylamino pyridine.

Embodiment:

Column chromatography method among the present invention:

The minimum 70cm of the length of chromatography column, inner filling 254-silica gel, and will need isolating organism to be dissolved in minimum chloroform entirely: methyl alcohol=in 1: 1; This solution absorption is placed on the top of silica gel in the chromatography column with minimum 254-silica gel; Use the moving phase wash-out, chromatography column connects the solution that obtains through column chromatography with several 10ml test tubes down, and the control flow velocity is 10ml/3min; The solution of each test tube is analyzed with HPLC; The test tube solution that RT is identical merges, and the compound of getting principal point carries out recrystallization, obtains corresponding product.

Confirm the method for principal point: need analyze with HPLC by isolating organism, confirm as principal point except that the maximum point of the peak area of raw material point, its RT is the RT of principal point.

HPLC can be according to T-(COCH3) _T1The correlation method of (mode that T links to each other with COCH3 is identical with the mode that T links to each other with B) is measured, and also can measure through following condition, and the method minimum with principal point content is as the criterion:

Equipment: HP1084B liquid chromatograph, HP79850 BLC terminal and UV detector

Column material: Hypersll C18,5um, 125 * 4.6mm

Detect wavelength: 242nm

Moving phase: methyl alcohol: water=5.8: 4.2

Column temperature: 45 ℃

Flow velocity: about 1.2ml/ branch

Embodiment 1 3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β hydroxyl-17-(3-nitroxide) propionic ester

1.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-chlorine) propionic ester

Method 1:

The triethylamine of 40ml and the 3-chlorpromazine chloride of 0.015mol are stirred and be cooled to-20 degree in reacting also, slowly add 0.01mol3-hydroxy-estra-1,3; 5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy also keeps temperature-20 to-15 degree, and this reaction is thermopositive reaction; So adding speed and temperature all need careful control, and above-mentioned reacting liquid temperature is remained on below-10 ℃, finish the back and in 1 hour, slowly rise to the 0-5 degree; With this temperature insulation after 10 hours, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature, slowly adds hydrochloric acid and regulates PH for neutral; This liquid uses methylene dichloride 90ml to extract three times (30ml * 3), and water 90ml three times (30ml * 3) washs dichloromethane layer again, and the pH value is transferred to neutrality; Dichloromethane layer concentrates, and pours methyl alcohol and carries out recrystallization, obtains title compound bullion 0.0053mol.

Method 2:

With 0.01mol3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy add in the 60ml pyridine;-20 ℃ of addings stir, and slowly add 3-chlorpromazine chloride 0.0015mol at-20 to-15 ℃, and this reaction is thermopositive reaction; So adding speed and temperature all need careful control, and above-mentioned reacting liquid temperature is remained on below-10 ℃, add the back and under-5 degree, be incubated 10 hours; Then reaction solution is cooled to 0-5 ℃, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature, and slowly adds HCl in cooling with under stirring; Above-mentioned reaction is thermopositive reaction, but temperature can not surpass 10 ℃, and certain amount of H Cl is used for the pH value is transferred to neutrality.This liquid uses methylene dichloride 90ml to extract three times (30ml * 3), and water 90ml three times (30ml * 3) washs dichloromethane layer again, and the pH value is transferred to neutrality, and dichloromethane layer concentrates, and pours methyl alcohol and carries out recrystallization, obtains title compound bullion 0.0051mol.

Also can use DMAP in the reaction method 1 and 2, corresponding content can reduce by 40%, and the product that reaction produces can obviously increase 3-hydroxyl-(3-chlorine) propionic ester and 3, the impurity of the two hydroxyls of 17--two (3-chlorine) propionic ester.

2.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-nitroxide) propionic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-chlorine) propionic ester 0.005mol is in the solution of acetonitrile 40ml, with AgNO ₃0.14 gram adds and refluxed 10 hours; After solution down removed to desolvate in decompression carry out column chromatography, be the moving phase wash-out with methyl alcohol and chloroform (1: 1), getting wherein, the principal point compound carries out concentrating under reduced pressure; Pour methyl alcohol and carry out recrystallization, get title compound 0.0031mol.

Ultimate analysis calculated value (%): C, 63.30; H, 6.52; N, 3.36; 0,26.83

Determination of elemental analysis value (%): C22H27NO7 C, 63.35; H, 6.57; N, 3.33; O, 26.75

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

112.1

?147.8

?150.9

115.4

?130.3

?30.6

?21.8

?39.6

The position of C

9

?10

?11

12

?13

?14

?15

?16

? 13C-NMR

128.3

?132.7

?118.0

29.7

?40.5

?47.6

?25.9

?28.4

The position of C

17

?18

?19

20

?21

?22

?23

?24

? 13C-NMR

84.5

?11.7

?175.0

29.8

?68.7

?56.9

Embodiment 2 3-hydroxy-estras-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-methyl-4-nitroxide)-2-ester thiohenic acid

With 3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy is a raw material, according to the method for embodiment 1 and 4-bromo-3-methyl-2-thiophene chloride, AgNO ₃Reaction obtains title compound 0.0030mol.

1.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-methyl-4-bromine)-2-ester thiohenic acid

Method 1:

The triethylamine of 40ml, 4-bromo-3-methyl-2-thiophene chloride of 0.015mol are stirred and be cooled to 0 degree in reacting also, slowly add 0.01mol3-hydroxy-estra-1,3; 5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy also keeps temperature 0-5 degree, and this reaction is thermopositive reaction; So adding speed and temperature all need careful control, and above-mentioned reacting liquid temperature is remained on below-10 ℃, add the back and under-5 degree, be incubated 10 hours; Then reaction solution is cooled to 0-5 ℃, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature, slowly adds hydrochloric acid and regulates PH for neutral; This liquid uses methylene dichloride 90ml to extract three times (30ml * 3), and water 90ml three times (30ml * 3) washs dichloromethane layer again, and the pH value is transferred to neutrality; After dichloromethane layer down removed to desolvate in decompression carry out column chromatography, be the moving phase wash-out with methyl alcohol and ETHYLE ACETATE (1: 1), getting wherein, the principal point compound carries out concentrating under reduced pressure; Pour methyl alcohol and carry out recrystallization, get title compound 0.0057mol.

Method 2:

With 0.01mol3-hydroxy-estra-1,3,5 (10); 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy add in the 60ml pyridine, and 0-10 ℃ of adding stirs, and slowly adds 4-bromo-3-methyl-2-thiophene chloride 0.0015mol at-15 ℃; This reaction is thermopositive reaction, so adding speed and temperature all need careful control, above-mentioned reacting liquid temperature is remained on below-10 ℃; Add the back and under-5 degree, be incubated 10 hours, then reaction solution is cooled to 0-5 ℃, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature; And under cooling and stirring, slowly add HCl; Above-mentioned reaction is thermopositive reaction, but temperature can not surpass 20 ℃, and certain amount of H Cl is used for the pH value is transferred to neutrality.This liquid uses methylene dichloride 90ml to extract three times (30ml * 3); Water 90ml three times (30ml * 3) washing dichloromethane layer again, pH value is transferred to neutrality, after dichloromethane layer is carried out column chromatography except that desolvating down in decompression; With methyl alcohol and ETHYLE ACETATE (1: 1) is the moving phase wash-out; Get wherein that the principal point compound carries out concentrating under reduced pressure, pour methyl alcohol and carry out recrystallization, title compound 0.0054mol.

Also can use DMAP in the reaction method 1 and 2, corresponding content can reduce by 30%, and the product that reaction produces can obviously increase 3-ester and 3, the impurity of 17-dibasic acid esters.

2.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-methyl-4-nitroxide)-2-ester thiohenic acid

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-methyl-4-bromine)-2-ester thiohenic acid 0.005mol is in the solution of acetonitrile 40ml, with AgNO ₃0.12 gram adds and refluxed 20 hours; After solution down removed to desolvate in decompression carry out column chromatography, be the moving phase wash-out with methyl alcohol and ETHYLE ACETATE (1: 1), getting wherein, the principal point compound carries out concentrating under reduced pressure; Pour methyl alcohol and carry out recrystallization, get title compound 0.0029mol.

Ultimate analysis calculated value (%): C, 61.84; H, 5.60; N, 2.88; 0,23.07; S, 6.60

Determination of elemental analysis value (%): C25H27NO7S C, 61.80; H, 5.59; N, 2.90; S, 6.62, O, 23.09

¹³C-NMR (CDCl ₃): 1 numerical value to 25 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 25 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?112.1

?147.8

?150.6

?115.3

?130.3

?30.6

?21.8

?39.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?128.5

?132.7

?118.0

?29.7

?40.5

?47.6

?25.9

?28.4

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?83.6

?11.7

?162.0

?149.1

?137.2

?161.1

?103.8

?6.3

The position of C

?25

?26

?27

?28

?29

?30

?31

?32

? 13C-NMR

?56.9

Embodiment 3 3-hydroxy-estras-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-3-(2-nitroxide) acetic ester

1.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-3-(2-chlorine) acetic ester

Method 1:

The triethylamine of 40ml, the DMAP of 0.05g and the 3-chlorpromazine chloride of 0.015mol are stirred and be cooled to 0 degree in reacting also, slowly add 0.01mol3-hydroxy-estra-1,3; 5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone also keeps temperature 0-5 degree, and this reaction is thermopositive reaction; So adding speed and temperature all need careful control, and above-mentioned reacting liquid temperature is remained on below 10 ℃, finish the back and in 1 hour, slowly rise to the 10-15 degree; With this temperature insulation after 15 hours, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature, slowly adds hydrochloric acid and regulates neutral; This liquid uses methylene dichloride 90ml to extract three times (30ml * 3), and water 90ml three times (30ml * 3) washs dichloromethane layer again, and the pH value is transferred to neutrality; Dichloromethane layer concentrates, and pours methyl alcohol and carries out recrystallization, obtains above-claimed cpd bullion 0.0060mol.

Method 2:

With 0.01mol3-hydroxy-estra-1,3,5 (10); The DMAP of 9 (11)-tetraenes-2-methoxyl group-17-ketone, 0.05g adds in the 60ml pyridine, and 0-10 ℃ of adding stirs, and slowly adds 3-chlorpromazine chloride 0.0015mol at 0-10 ℃; This reaction is thermopositive reaction, so adding speed and temperature all need careful control, above-mentioned reacting liquid temperature is remained on below 10 ℃; Add the back and under this temperature, be incubated 15 hours, then reaction solution is cooled to 0-5 ℃, being diluted to 50mlPH is in the water of 0 degree for the 1-2 temperature; And under cooling and stirring, slowly add HCl; Above-mentioned reaction is thermopositive reaction, but temperature can not surpass 20 ℃, and-quantitative HCl is used for the pH value is transferred to neutrality.This liquid uses methylene dichloride 90ml to extract three times (30ml * 3), and water 90ml three times (30ml * 3) washs dichloromethane layer again, and the pH value is transferred to neutrality, and dichloromethane layer concentrates, and pours methyl alcohol and carries out recrystallization, obtains above-claimed cpd bullion 0.0054mol.

Also can not use DMAP in the reaction method 1 and 2, the content of above-claimed cpd can reduce by 35%, and unreacted raw material can increase.

2.3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-3-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-(2-chlorine) acetic ester 0.005mol is in the solution of acetonitrile 40ml, with AgNO ₃0.12 gram adds and refluxed 20 hours; After solution down removed to desolvate in decompression carry out column chromatography, be the moving phase wash-out with methyl alcohol and ETHYLE ACETATE (1: 1), getting wherein, the principal point compound carries out concentrating under reduced pressure; Pour methyl alcohol and carry out recrystallization, get title compound 0.003mol.

Ultimate analysis calculated value (%): C, 62.83; H, 5.78; N, 3.49; O, 27.90

Determination of elemental analysis value (%): C21H23NO7 C, 62.91; H, 5.73; N, 3.44; O, 27.92

¹³C-NMR (CDCl ₃): 1 numerical value to 21 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

13C-NMR

?110.9

?149.2

?140.1

?120.9

?130.3

?30.2

?25.9

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?128.0

?136.7

?117.2

?34.8

?48.5

?47.9

?22.8

?35.9

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?221.4

?14.9

?180.4

?76.7

?55.9

Embodiment 4 3-methoxyl group female steroids-1,3,5 (10)-triolefins-17 beta-hydroxyl-17-(4-nitroxymethyl) benzoic ether

Think 3-methoxyl group female steroid-1,3,5 (10)-triolefins-17 beta-hydroxy is a raw material, according to method and (4-chloromethyl) Benzoyl chloride 99min., the AgNO of embodiment 1 ₃Reaction obtains title compound.

Ultimate analysis calculated value (%): C, 69.66; H, 6.71; N, 3.01; O, 20.62

Determination of elemental analysis value (%): C27H31NO6 C, 69.67; H, 6.73; N, 3.02; O, 20.58

¹³C-NMR (CDCl ₃): 1 numerical value to 27 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 27 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?126.5

?111.7

?158.3

?114.2

?138.1

?30.0

?28.2

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?44.7

?133.2

?27.1

?36.7

?42.9

?51.7

?23.7

?27.9

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?83.0

?12.4

?166.8

?130.4

?131.8

?130.5

?142.6

?130.5

The position of C

?25

?26

?27

?28

?29

?30

?31

?32

? 13C-NMR

?131.8

?79.3

?56.1

Embodiment 5 3-hydroxy-estras-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester-17-(3-nitroxide) propionic ester

With 3-hydroxy-estra-1,3,5 (10); 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxy is a raw material, and method 1 and the reaction of (3-chlorine) propionyl chloride according to embodiment 1 obtain 3-hydroxy-estra-1; 3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxyl-17-(3-chlorine) propionic ester; Method 1 according to embodiment 3 obtains 3-hydroxy-estra-1,3,5 (10) with (2-chlorine) Acetyl Chloride 98Min. again; 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxies-3-(2-chlorine) acetic ester-17-(3-chlorine) propionic ester, last according to method among the embodiment 1 and AgNO ₃(consumption is the twice of former method) reaction obtains title compound 3-hydroxy-estra-1,3, and 5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester-17-(3-nitroxide) propionic ester.

Ultimate analysis calculated value (%): C, 55.38; H, 5.42; N, 5.38; O, 33.81

Determination of elemental analysis value (%): C24H28N2O11 C, 55.44; H, 5.44; N, 5.35; O, 33.77

¹³C-NMR (CDCl ₃): 1 numerical value to 24 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 24 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?110.4

?149.0

?139.9

?120.7

?129.8

?30.0

?21.9

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?128.4

?135.1

?118.2

?28.4

?40.9

?47.1

?25.7

?27.8

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?83.3

?11.5

?173.6

?30.0

?69.2

?180.2

?76.5

?56.4

Embodiment 6 3-hydroxy-estras-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester

With 3-hydroxy-estra-1; 3; 5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester is a raw material, obtains 3-hydroxy-estra-1,3 according to the method for embodiment 3 and (2-chlorine) excess acetyl chloride; 5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-chlorine) acetic ester is again according to method among the embodiment 3 and AgNO ₃Reaction obtains title compound 3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester.

Ultimate analysis calculated value (%): C, 61.73; H, 6.53; N, 3.13; O, 28.60

Determination of elemental analysis value (%): C23H29NO8 C, 61.70; H, 6.55; N, 3.11; O, 28.64

¹³C-NMR (CDCl ₃): 1 numerical value to 23 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 23 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?109.4

?149.0

?137.3

?122.7

?129.8

?30.0

?27.9

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?44.7

?139.2

?26.9

?36.7

?42.9

?51.9

?23.7

?31.8

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?83.2

?12.2

?179.8

?76.9

?56.1

?171.5

?21.9

Embodiment 7 3-hydroxy-estras-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester

With 0.002mol3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester is dissolved among methyl alcohol and chloroform (1: the 1) 10ml; Drip the 0 degree aqueous sodium carbonate of saturated 0.0025mol down in 0 degree under the nitrogen protection, stirs after 5 hours, it is concentrated to pressurize after using hydrochloric acid conditioned reaction system PH as neutrality; Remove chloroform; This solution dilution in the 40ml frozen water, is filtered the dry title compound bullion that gets.Bullion is carried out column chromatography, is the moving phase wash-out with methyl alcohol and ETHYLE ACETATE (3: 2), gets wherein that the principal point compound carries out concentrating under reduced pressure, pour methyl alcohol and carry out recrystallization, title compound 0.0011mol.

Ultimate analysis calculated value (%): C, 62.21; H, 6.71; N, 3.45; O, 27.62

Determination of elemental analysis value (%): C21H27NO7 C, 62.24; H, 6.67; N, 3.42; O, 27.67

¹³C-NMR (CDCl ₃): 1 numerical value to 21 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 21 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?109.4

?149.0

?137.3

?122.7

?129.8

?30.0

?27.9

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

13C-NMR

?44.7

?139.2

?26.9

?36.7

?42.9

?51.9

?23.7

?31.8

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

13C-NMR

?81.5

?12.2

?179.8

?76.9

?56.0

Embodiment 8 3-hydroxy-estras-1,3,5 (10), 9 (11)-tetraenes-2-oxyethyl group-17 β-(2-nitroxide) acetic ester

Think 3-hydroxy-estra-1,3,5 (10); 9 (11)-tetraenes-2-oxyethyl group-17 beta-hydroxy is a raw material; Method according to embodiment 1 obtains 3-hydroxy-estra-1,3,5 (10) with (2-chlorine) excess acetyl chloride; 9 (11)-tetraenes-2-oxyethyl group-17 β-(2-chlorine) acetic ester, the back is according to method and the AgNO of a kind of embodiment ₃Reaction obtains title compound 3-hydroxy-estra-1,3, and 5 (10), 9 (11)-tetraenes-2-oxyethyl group-17 β-(2-nitroxide) acetic ester.

Ultimate analysis calculated value (%): C, 63.30; H, 6.52; N, 3.36; O, 26.83

Determination of elemental analysis value (%): C22H27NO7 C, 63.27; H, 6.55; N, 3.38; O, 26.80

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

The position of C

1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?112.1

?145.8

?147.0

?115.4

?130.3

?30.6

?21.8

?39.6

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?128.3

?132.7

?118.0

?29.7

?40.5

?47.6

?25.9

?28.4

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?84.5

?11.7

?170.7

?76.9

?65.2

?14.9

Embodiment 9:3-hydroxy-estra-1,3,5 (10)-triolefins-2-oxyethyl group-17-ketone-3-(2-nitroxide) acetic ester

With 3-hydroxy-estra-1,3,5 (10)-triolefins-2-oxyethyl group-17-ketone is raw material, according to method and (2-chlorine) Acetyl Chloride 98Min., the AgNO of embodiment 3 ₃Reaction obtains 3-hydroxy-estra-1,3,5 (10)-triolefins-2-oxyethyl group-17-ketone-3-(2-nitroxide) acetic ester.

Ultimate analysis calculated value (%): C, 63.30; H, 6.52; N, 3.36; O, 26.83

Determination of elemental analysis value (%): C22H27NO7 C, 63.29; H, 6.53; N, 3.34; O, 26.85

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

¹³C-NMR (CDCl ₃): 1 numerical value to 22 carbon:

The position of C

?1

?2

?3

?4

?5

?6

?7

?8

? 13C-NMR

?109.4

?149.0

?137.3

?137.7

?123.1

?30.0

?27.9

?38.9

The position of C

?9

?10

?11

?12

?13

?14

?15

?16

? 13C-NMR

?44.7

?139.2

?26.9

?33.1

?48.9

?52.0

?21.7

?36.6

The position of C

?17

?18

?19

?20

?21

?22

?23

?24

? 13C-NMR

?220.5

?14.0

?179.8

?76.8

?65.2

?14.9

Embodiment 10: injection

Main ingredient:

3-methoxyl group female steroid-1,3,5 (10)-triolefins-17 beta-hydroxyl-17-(4-nitroxymethyl) benzoic ether 5g

Auxiliary material:

Vitamin PP 70g

Phenylcarbinol 7.5ml

Water for injection adds to 1000ml

[preparation] with 3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 β-alcohol-3, and 17-two (2-nitroxide) acetic ester is mixed well for use with a small amount of water for injection earlier; Vitamin PP is dissolved in an amount of water for injection again, adds gac 0.1g, 15mln is placed in the back that stirs; The coarse filtration decarburization adds the injection water to about 900ml, is heated to 80 ~ 90 ℃ in the water-bath; Slowly add pregnant steroid-4,9 (11)-diene-3 that has mixed up with water for injection, 20-diketone-17; Two hydroxyl-21-(2-nitroxide) acetic esters of 21-, insulation 20 ~ 30mln dissolves postcooling fully to room temperature.Add phenylcarbinol, regulate pH to 6.5 ~ 6.0, the adjustment volume is to 1000ml, then 10 ℃ with held 8h, be filtered to clear and bright, embedding, 100 ℃ of circulation vapor sterilization 15 min get final product.

Embodiment 11: oral capsule

With 20mg3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester diluted mixture becomes the pharmaceutical composition of 200mg with starch in small, broken bits, in No. 4 hard capsule of packing into.

Pharmacology embodiment 1: experiment in vitro

Adopt the influence of chick chorioallantoic membrane (CAM) blood vessel hyperplasia model observation type (1) compound of growth factor-induced to blood vessel hyperplasia, understanding formula (1) compound is to the effect of vasculogenesis.

Experiment material:

The instar chicken embryo on the 3rd of being fertilized

Glass fiber filter paper

Vascular endothelial growth factor (VEGF) Chemlcon Company products

The compound that embodiment 1-9 is made with the DMSO dissolving, is diluted to needed concentration (DMSO concentration＜0.1%) with PBS earlier again

Experimental technique

1.CAM the foundation of model:

75% ethanol cleans chicken ovigerm shell and in super clean bench, dries up, and behind the horizontal positioned 5min, carefully chorion is broken into pieces at 100mm petridish edge, and the ovum content places petridish (petridish is added with 10ml DMEM substratum in advance).This petridish is put into the big petridish of 150mm (big petridish adds little water), cover the ware lid, place 37 ℃, 5%CO ₂Cell culture incubator in cultivate.

2. to the influence of CAM blood vessel hyperplasia

With tapping and plugging machine glass fiber filter paper is processed the sequin of diameter 3mm, moist heat sterilization, each 10 μ l drips on glass fiber filter paper with reagent, processes the medicine film, dries up subsequent use.Behind the chicken embryo culture 3d; Be divided into 13 groups at random; Every group 8, the compound of embodiment 1-9 gained is made as 9 administration groups (1g/L), give growth factor (2 μ g/L) simultaneously; And the independent stimulating group (positive controls) of growth factor (VEGF) and PBS (phosphate buffered saline buffer, PH7.4) stimulating group (negative control group).The medicine film is affixed on CAM and yolk cyst membrane (yolk sacmembrane, YSM) the outer less position of 2/3 place's blood vessel.Behind dosing 48 h, microscopically is observed, large, medium and small blood vessel number in the 5mm around the counting medicine film.

Table 1 pair CAM blood vessel hyperplasia influence table (

n=8)

Group name	Aorta	Medium vessels	Little blood vessel	P (each group is to the VEGF group in the little blood vessel)
					PBS	?1.00±0.53	?2.50±0.67	?8.25±0.35	＜0.01
?VEGF	?1.50±0.85	?2.50±0.33	?17.50±0.81
					Embodiment 1	?1.00±0.53	?2.50±0.34	?6.75±0.93	＜0.01
Embodiment 2	?1.10±0.48	?2.50±1.05	?7.50±0.71	＜0.01
					Embodiment 3	?1.20±0.59	?2.50±0.78	?7.50±1.36	＜0.01
Embodiment 4	?1.0±0.62	?2.50±0.40	?6.75±1.41	＜0.01
					Embodiment 5	?1.00±0.53	?2.25±0.33	?6.00±0.68	＜0.01
Embodiment 6	?1.00±0.38	?2.25±0.47	?6.25±1.06	＜0.01
					Embodiment 7	?1.00±0.38	?2.25±0.40	?6.25±1.36	＜0.01
Embodiment 8	?1.2±0.69	?2.50±0.59	?7.00±1.14	＜0.01
					Embodiment 9	?1.2±0.43	?2.50±0.91	?7.50±1.23	＜0.01

Annotate: the statistical method data are represented with

. relatively use the t check between group

3. result

1 formula (1) compound is to VEGF inductive CAM

Blood vessel hyperplasia influence table 1 as a result, it is obvious that the good .VEGF of PBS group angiogenic growth organizes little blood vessel hyperplasia, the blood vessel hyperplasia of respectively organizing of formula (1) compound obviously is suppressed. little blood vessel significantly reduces, and gets back to normal level basically.

CAM is classical vasculogenesis evaluation model, have method easy, advantage such as be prone to observe, inexpensive. be at present the most frequently used in the phantom type.VEGF is important short angiogenesis factor, and the hyperplasia and the migration of ability stimulating endothelial cell promote vasculogenesis, and in the kinds of tumors tissue, find over-expresses.This research shows: formula (1) compound suppresses VEGF inductive CAM blood vessel hyperplasia, shows that formula (1) compound has the short angiogenic action that suppresses VEGF.

Formula (1) compound has restraining effect to vasculogenesis, has further confirmed to have the potentiality that suppress tumor-blood-vessel growth.

Pharmacology embodiment 2: experiment in the body

1. laboratory animal and knurl strain

Kunming mouse is selected in experiment for use, and male, body weight (20 ± 2) g, divides 16 groups, 10 every group by 160.

Murine sarcoma knurl strain S180

2. medicine

Cis-platinum (DDP) injection liquid: 20ml: 20mg/ props up.

3. method

3.1 the foundation of knurl mouse model: the strain of murine sarcoma S180 knurl, the abdominal cavity inoculation of going down to posterity.When treating the ascites well-grown, extract ascites out, cell counting, the adjustment cell concn is 2 * 107 cell/ml, at mouse oxter subcutaneous injection S180 sarcoma cell, every inoculation 0.25ml observed the local tumor growing state in the 11st day.

3.2 treatment and grouping: 160 mouse were divided into 16 groups, 10 every group in inoculation the same day at random.Pressing table gives and drug test grouping information slip

Group number	Activeconstituents	Administering mode	Dosage
				Control group	Saline water	Every day is once oral	0.4ml
The DDP group	Cis-platinum	The next day abdominal injection once	0.5mg/kg
				?1	Embodiment 1	Every day is once oral	1mg/kg
?2	Embodiment 2	Every day is once oral	1mg/kg
				?3	Embodiment 3	Every day is once oral	1mg/kg
?4	Embodiment 4	Every day is once oral	1mg/kg
				?5	Embodiment 5	Every day is once oral	1mg/kg
?6	Embodiment 6	Every day is once oral	1mg/kg
				?7	Embodiment 7	Every day is once oral	1mg/kg
?8	Embodiment 8	Every day is once oral	1mg/kg
				?9	Embodiment 9	Every day is once oral	1mg/kg
?10	Embodiment 5	Every day is once oral	0.01mg/kg
				?11	Embodiment 5	Every day is once oral	0.1mg/kg
?12	Embodiment 5	Every day is once oral	10mg/kg
				?13	Embodiment 5	Every day is once oral	20mg/kg
The DDP combined group	Embodiment 5 and DDP	Embodiment is once oral 5 every days, and abdominal injection once next day of DDP	Embodiment 5,1mg/kg DDP, 0.5mg/kg

Annotate: dosage is by activeconstituents in the embodiment test group.

According to document " cis-platinum is to the genotoxic research of mouse bone marrow cells " (" canceration. distortion. sudden change ", 1996 86 phases of volume, the dosage of 0.5mg/kg is adopted in the introduction in 362-365) to mouse.

Medication 10d, 60min behind the last filling stomach puts to death and respectively organizes mouse, strips the knurl piece, weighs, and the knurl piece is made the pathology tissue slice.By formula calculate tumour inhibiting rate: tumour inhibiting rate=(the average knurl of the average knurl weight-experimental group of control group is heavy)/average knurl of control group heavy * 100%.

3.3 capillary blood vessel dyeing: immunohistochemical methods SABC method dyeing blood vessel, instant capillary blood vessel staining kit (CD31) is Wuhan doctor's moral biotech firm product.After each is organized the knurl body and peels off, cut sample, fixing, embedding, section.Section after dyed, endotheliocyte is brown to be dyed, and blood vessel is tawny, is easy to identification.

The mensuration of MVD: MVD is according to people such as Weldner (Weldner N, Semple JP, Welch WR.Et al.Tumor anglogeneslsand Metastastasls correlatlon ln lnvaslve breast carclnonma; N Engl J Med, 1991,324; Method 1-8) and judgement criteria are carried out; Calculate the capillary vessel and the tiny blood vessels of tumour intrinsic color, promptly the most intensive capillary blood vessel mark zone is selected by the whole tumor tissue section of pan under low-power field; The all single clearly endotheliocyte of tawny mark or endotheliocyte strings of appearing have the aorta of thicker flesh wall and tube chamber area not to count greater than the blood vessel of 8 red blood cell diameters all as an isarithmic capillary blood vessel.Method of counting: under low power lens (10 * 10) visual field, sweep whole tissue slice earlier; Select the most intensive visual field, 3 capillary blood vessel mark zones in tumor-infiltrated district; Promptly so-called " new vessel hot zone " is then at the bottom of the identical back of the body, under the back of the body concrete conditions in the establishment of a specific crime, with high power lens (20 * 20) visual field (0.72mm ²) for all painted MVCs of standard counting, get the measured value of its MV as this sample.

3.4 VEGF immunohistochemical methods: VEGF immunologic combined detection reagent kit (instant), opticmicroscope is observed down: the VEGF positive staining the positive expression of brown yellow granule occurs with tumour cell and near vascular endothelial cell slurry or after birth thereof.Every stained is carried out absorbancy (area density) and intensity (gray scale) mensuration through MP1AS-1000 high-definition color pathological image analytical system then.

3.5 statistical method: utilization SPSS statistical software, adopt the q check.There is statistical significance P＜0.05 for difference.

4. result

Heavily change comparison 4.1S180 respectively organize mice-transplanted tumor

The variation that knurl is heavy: each treatment group knurl weight average significantly is lower than control group; Compare with control group; Difference all has statistical significance, and experimental result shows that various formulas (1) compound has obvious restraining effect to the S180 transplanted tumor, share with DDP to have synergism; And the formula of various dose (1) compound also has the positively related restraining effect of dosage to the S180 transplanted tumor, and particular case is seen table 2.

Table 2 formula (1) compound is to the restraining effect

of S180 transplanted tumor

Group	Mouse number (only)	Knurl heavy (gram)	Tumour inhibiting rate (%)	The P value
					Control group	10	2.53±0.47
The DDP group	10	1.18±0.11	53.4	＜0.01
					1	10	1.35±0.41	46.6	＜0.01
2	10	1.81±0.68	28.5	＜0.01
					3	10	1.74±0.71	31.2	＜0.01
4	10	1.37±0.45	45.8	＜0.01
					5	10	1.23±0.39	51.4	＜0.01
6	10	1.26±0.34	50.2	＜0.01
					7	10	1.27±0.41	49.8	＜0.01
8	10	1.38±0.51	45.5	＜0.01
					9	10	1.68±0.60	33.6	＜0.01
10	10	2.12±0.73	16.2	＜0.05
					11	10	1.80±0.57	28.9	＜0.05
12	10	1.15±0.20	54.5	＜0.01
					13	10	1.06±0.16	58.1	＜0.01
The DDP combined group	10	1.07±0.18	60.1	＜0.01

4.2 formula (1) compound is to the influence of S180 knurl body microvessel density

Each is organized the tumour Pathologic specimen and dyes through CD31; The matter blood vessel all has painted between tumor tissues; Dyed the positive expression of tawny with endotheliocyte, microvascular size, morphological differences are bigger, and what have is merely single endotheliocyte and endotheliocyte family; The tube chamber that has is not obvious or form is irregular, and the MVD of borderline tumor tissue is higher than central authorities.See a large amount of new trichoblast blood vessels in the control group tumour, positive target be positioned at tumor tissues and between the tawny particle that distributes of matter slabbing or lumps, each medication group positive expression cell granulations reduces, and sees table 3.

The table average microvessel density of 3S180 knurl body

Group	Mouse number (only)	Average microvessel density	P value (each group and control group)
				Control group	10	127.4±38.4
The DDP group	10	79.9±21.5	＜0.01
				1	10	76.6±23.1	＜0.01
2	10	87.1±23.4	＜0.01
				3	10	82.4±27.7	＜0.01
4	10	73.9±29.5	＜0.01
				5	10	70.4±25.9	＜0.01
6	10	71.8±20.7	＜0.01
				7	10	72.1±21.0	＜0.01
8	10	77.3±25.6	＜0.01
				9	10	80.8±29.2	＜0.01
10	10	98.3±34.8	＜0.05
				11	10	78.9±28.6	＜0.05
12	10	65.7.1±10.9	＜0.01
				13	10	55.8±12.9	＜0.01
The DDP combined group	10	56.5±13.6	＜0.01

4.3 formula (1) compound is organized the influence of vegf expression to mouse S180 sarcoma

Showed by immune group result, opticmicroscope observe down the control group tumor tissues and therebetween matter see a large amount of in the form of sheets or the brown yellow granule that distributes of lumps, each medication group positive expression cell obviously reduces.Vegf expression average area density and average gray result are seen table 4.

Table 4S180 sarcoma is organized vegf expression data sheet

Group	Mouse number (only)	Centre plane density	P (each group and control group)	Average gray	P (each group and control group)
						Control group	10	39.2±3.39		157.3±32.8
The DDP group	10	18.6±3.74	＜0.01	81.4±24.6	＜0.01
						1	10	9.7±3.25	＜0.01	84.1±28.9	＜0.01
2	10	10.8±4.12	＜0.01	112.4±31.5	＜0.01
						3	10	9.8±4.23	＜0.01	101.5±35.8	＜0.01
4	10	9.6±3.58	＜0.01	88.3±28.5	＜0.01
						5	10	7.8±2.67	＜0.01	76.2±25.3	＜0.01

6	?10	?8.1±2.41	?＜0.01	79.1±27.0	?＜0.01
						7	?10	?8.2±2.16	?＜0.01	78.5±29.2	?＜0.01
8	?10	?9.6±3.76	?＜0.01	82.7±28.7	?＜0.01
						9	?10	?10.0±2.86	?＜0.01	94.1±25.6	?＜0.01
10	?10	?13.84±4.19	?＜0.01	121.5±34.7	?＜0.05
						11	?10	?9.0±3.37	?＜0.01	90.4±30.2	?＜0.05
12	?10	?6.9±2.56	?＜0.01	64.8±23.6	?＜0.01
						13	?10	?6.8±2.59	?＜0.01	57.7±20.3	?＜0.01
The DDP combined group	?10	?7.78±2.69	?＜0.01	50.7±6.84	?＜0.01

5 conclusions

Invasive growth and metastatic potential are the essential characteristics of malignant tumour; It also is the lethal major cause of malignant tumor patient; There is conclusive evidence to show; Malignant tumor patient more than 90% is finally died from metastases and recurrence, and shifts usually early stage the generation, and about 50% patient has produced metastasis when clinical diagnosis goes out primary tumor.For the metastatic tumor of fast breeding, few apoptosis, the distribution of many kitchen ranges property, heterogeneous growth, conventional treatment means such as present operation, radiotherapy, chemotherapy are often failed, final death.Shifting real is malignant tumour intractable and intractable basic reason.Research shows: in growth, infiltration and the transfer of solid tumor, the vasculogenesis that continues is a key factor., oncocyte division (generally is no more than 1～2mm when rising in value to certain volume ²), if still there is not the grow into nutritive substance and the oxygen that provide necessary and breed the needed various factor of blood vessel, cell count that it is dead and outgrowth cell count about equally, oncocyte group stops expansion and reaches metastable state.When the tumor promotion host is a vascular proliferation, some new vesseles are grown into behind the tumor tissues, and tumor cell group increases sharply, and volume increases, tissue infiltration towards periphery, and have metastasis tendency.Experimental result shows in pharmacology embodiment 2 bodies; Formula (1) compound group all obviously reduces with MVD, the VEGF ratio of control group; And formula (1) compound and cisplatin combined group of MVD, VEGF value reduce more obviously, and this shows formula (1) compound and cisplatin combined treatment tumour, has synergy; Reduce angiogenesis, improved anticancer effect.

Can prove that through the data in pharmacology embodiment 1 and 2 formula (1) compound is inhibited to mammiferous angiogenesis and VEGF, can treat or prevent diseases associated therewith.

Claims (13)

1. compound is:

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-nitroxide) propionic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 β-(3-methyl-4-nitroxide)-2-ester thiohenic acid

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17-ketone-3-(2-nitroxide) acetic ester

3-methoxyl group female steroid-1,3,5 (10)-triolefins-17 β-(4-nitroxymethyl) benzoic ether

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester-17-(3-nitroxide) propionic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxyl-17s-acetic ester-3-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-methoxyl group-17 beta-hydroxies-3-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10), 9 (11)-tetraenes-2-oxyethyl group-17 β hydroxyl-17-(2-nitroxide) acetic ester

3-hydroxy-estra-1,3,5 (10)-triolefins-2-oxyethyl group-17-ketone-3-(2-nitroxide) acetic ester.

2. acceptable salt suppresses the application of the medicine of mammal VEGF on the described compound 1 of claim 1 or its physiology in preparation.

On the described compound 1 of claim 1 or its physiology acceptable salt the preparation angiogenesis inhibitor application.

4. the said compound 1 of claim 1 is in the application of preparation treatment tumour class disease medicament.

The said compound 1 of claim 1 in the medicine of preparation treatment ophthalmology eye neovascular disease application.

6. the application of the said compound 1 of claim 1 in the medicine of preparation treatment malignant hematologic disease.

7. the application of the said compound 1 of claim 1 in the medicine of preparation treatment bronchial asthma.

8. the application of the said compound 1 of claim 1 in the medicine of the loose disease of preparation treatment alba.

9. a medicinal compsns is characterized in that said composition comprises acceptable salt on the said compound 1 of claim 1 or its physiology.

10. medicinal compsns as claimed in claim 9 is characterized in that being mixed by acceptable drug auxiliary material on acceptable salt on the said compound 1 of claim 1 or its physiology and one or more physiology.

11. medicinal compsns as claimed in claim 9 is characterized in that being mixed with liquid preparation, sterilization preparation and sterile preparation, solid preparation, semi-solid preparation, aerosol.

12. an ophthalmic preparation is characterized in that said preparation contains acceptable salt on the said compound 1 of claim 1 or its physiology, and as the pharmaceutical excipient of carrier.

13. an oral prepns is characterized in that said preparation contains acceptable salt on the said compound 1 of claim 1 or its physiology, and as the pharmaceutical excipient of carrier.