CN1609118A - Cell wall acyl-alanyl-D-isoglutamine derivative and its prepn, medicine composition and use - Google Patents
Cell wall acyl-alanyl-D-isoglutamine derivative and its prepn, medicine composition and use Download PDFInfo
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Abstract
The present invention discloses compound Nalpha-(1-alpha-benzyl-N-acetyl-cell wall acyl-L-alanyl-D-isoglutamine aminoacyl)-Nbeta-(3-nitro-trans-cinnamenyl)-L-lysyl amine and its preparation process. It has the function of treating tumor and may be used as adjuvant for the treatment or assisting treatment of late stage tumor, AIDS, hemorrhagic shock, lymphoma, viral disease and autoimmune disease.
Description
Technical field
The present invention relates to the compound N shown in general formula (1)
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-L-lysyl amine and preparation method thereof, it has the effect of treatment tumour, and can be as the purposes that treatment or assisting therapy arranged of adjuvant to late tumor, acquired immune deficiency syndrome (AIDS), hemorrhagic shock, lymphoma, virus disease, autoimmune disease.
Background technology
After the branch bacillus heat-killed, be suspended in the Fu Shi Freund's complete adjuvant of making in the mineral oil (FCA), can increase body effectively antigenic humoral immune reaction and cell immune response.N-ethanoyl Muramyl dipeptide (MDP; N-Acetylmuramyl Dipeptide) is the minimum resulting structure unit of FCA; has wide biological activity, as non-specific anti-infective (pneumobacillus, intestinal bacteria, Pseudomonas aeruginosa, listerisa monocytogenes in mjme, Candida albicans etc.), non-specific antitumor (fibrosarcoma, hepatoma etc.), immunomodulatory, short dormancy etc.But its toxicity (immunogenicity causes allergic reaction, pyrogenicity, cause inflammation, pain) and metabolism limit rapidly etc. its application clinically.The Muramyl dipeptide compound is to activate specifically or each cell of inhibition human immune system by non-, generates biological activity as scavenger cell, myelomonocyte, neutrophil leucocyte, T and bone-marrow-derived lymphocyte etc.People have dropped into very big enthusiasm to its structure of modification, have synthesized hundreds of MDP derivatives, have found some activity higher, the compound that side effect is lower, and its structure activity relationship carried out deep discussion.Insert the N-muramyl-tripeptide-O-phosphatidylethanolamine (MTP-PE) of fat-soluble group at the peptide chain carbon teminal, have very strong antitumor action, finished the second stage of clinical study in 1991 in the U.S., its major side effects is heating and shiver with cold.N
α-MDP-N
ε-stearyl Methionin [MDP-Lys (L18)] has very strong anti-infective, antitumor action, has kept the immunocompetence of MDP, and pyrogenicity reduces, and obtains approval in June, 1991 in Japan, as put, immunostimulant listing after the chemotherapy.
Summary of the invention
In order to overcome the deficiencies in the prior art, the invention provides a kind of brand-new compound N
α-(1-α-benzyl-N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-lysyl amine, code name is " MDP-C ".
Another object of the present invention is to provide multiple preparation N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
εThe method of-(3-nitro-trans styryl carbonyl)-L-lysyl amine.
Another object of the present invention is to provide a kind of pharmaceutical composition, and it comprises medicine effective dose, as carrier commonly used in the compound of the formula (I) of active ingredient and isomer and the pharmacy field.
A further object of the present invention is to provide the antitumor in preparation of a kind of new compound and composition thereof, or as adjuvant to the application in the medicine of the purposes that treatment or assisting therapy are arranged of late tumor, acquired immune deficiency syndrome (AIDS), hemorrhagic shock, lymphoma, virus disease, autoimmune disease.
In order to finish the present invention's purpose, the present invention takes following technical scheme:
Specifically, one aspect of the present invention relates to suc as formula the compound N shown in (I)
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-L-lysyl amine:
According to the invention still further relates to preparation The compounds of this invention N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
εThe method of-(3-nitro-trans styryl carbonyl)-L-lysyl amine specifically, comprises the steps:
(A) Fmoc protection D-isoGln's is synthetic, sees route 1.
Route 1
Change the hydroxyl of the carboxylic acid of the amido end of D-L-glutamic acid into amido, simultaneously with original amido in the Fmoc protection D-L-glutamic acid.
When the hydroxyl of the carboxyl of D-glu famine cardinal extremity changes amido into, at first the amido of D-L-glutamic acid is protected, preferred protecting group is a carbobenzoxy-(Cbz).
When D-L-glutamic acid carries out the carbobenzoxy-(Cbz) protection, preferably D-L-glutamic acid and benzyloxy dicarbonyl chloride reaction.
The reaction of D-L-glutamic acid and benzyloxy dicarbonyl chloride is preferably under the condition that alkali exists.Preferred alkali is NaOH.
Reaction preferably 5～30 minutes.
Temperature of reaction, preferably 30～80 ℃, be more preferably 40-70 ℃, most preferably be 55-65 ℃.
Use hcl acidifying pH=1～5 again.
The Glu of carbobenzoxy-(Cbz) protection is under the condition that DCC exists and feed ammonia gas react.
Temperature of reaction: cryosel is bathed cooling
The time of reaction is: 30 minutes
After the hydroxyl of carboxylic acid is replaced as amido, slough the protecting group that the first step connects, slough carbobenzoxy-(Cbz) and preferably use the hydrogenant method; The hydrogenant method is preferably used palladium carbon;
The temperature of hydrogenation: 0-40
Hydrogenant pressure: normal pressure-3 normal atmosphere
The hydrogenant time: 1 ~ 3 day.
Fmoc protecting group on the amino acid whose amido;
Temperature of reaction: cool off under the condition of ice bath
Reaction times: 24 hours
With acidifying pH to 2 ~ 4.
Solvent uses saturated 100mL NaHCO
3With 200mL acetone.
(B) the protection teichoic acid is synthetic, sees route 2.Document: Liebigs Ann.Chem.1986,35-47
Synthesizing of circuit 2 protection protection teichoic acids
1-benzyl-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-4,6-O-benzal-3-O-[(R, S)-(2 '-methoxycarbonyl) ethyl]-α-D-glucoside (4r, s) split: the silica gel consumption is 1: 40-60; Preferred silica gel consumption is 1: 45-55.With the ascending elutriant of polarity wash-out successively, preferred elutriant is ethyl acetate/CH successively
2Cl
2=0%, 15%, 20%, 25%, 40%, ethyl acetate.Detect with high performance liquid phase, merge respectively and contain 4r, 4s flows part, concentrates and gets teichoic acid methyl esters (4r) and different teichoic acid methyl esters (4s) respectively.
C) N
α-(1-α-benzyl-N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
εThe solid phase synthesis of-(3-nitro-trans styryl carbonyl)-lysyl amine is seen route 3.
The route of route 3 solid phase synthesis MDP-C
Resin: preferred Crown resin, Rink Amide resin, Rink MBHA Amide resin, Wang resin, and the resin that produces carboxyl or amide group after other cracking.
Resin removes protecting group.Deprotection: preferably use the solution of 20-50% piperidines/DMF, 20 minutes～3 hours reaction times.
Fmoc-Lys (Dde)-OH amino acid is connected resin.Condensing agent: preferred HBTU, TBTU, HATU, BOP, PyBrBOP, DCC, DIC, and other condensing agent for synthesizing polypeptide.Additive: preferred HOBt, HOAt, HOSu, and the synthetic additive that is used to form Acibenzolar of other polypeptide.Connect amino acid and preferably use TBTU and HOBt.Temperature of reaction: 10 ℃～80 ℃, preferably 25 ℃.Reaction times is 3～30 hours, preferably hour 5-24 hour.Take off the Fmoc protecting group after having reacted, preferably use 50% piperidines/DMF solution deprotection.
Making uses the same method connects Fmoc-D-isoGln-OH, Fmoc-Ala-OH and protection teichoic acid.
After using 30～60% anhydrous hydrazines/DMF solution to remove the Dde protecting group again, in polar solvent (for example water), react with 3-nitrocinnamic acid Acibenzolar.
With the 10ml 20%TFA-aqueous solution synthetic compound is cut from resin at last and fallen.
Nitrogen dries up, and removes the benzal protecting group with 10～30% HCl again and gets compound of the present invention.
For stable reaction is carried out fast, reactant is heated and stirring etc.; For the product purification that makes reaction adopts recrystallization and silica gel column chromatography.
The invention still further relates to a kind of pharmaceutical composition that contains medicine effective dose as described compound of general formula I and pharmaceutically acceptable carrier.
According to the present invention, the form that The compounds of this invention can isomer exists, and described usually " The compounds of this invention " comprises the isomer of this compound.
According to embodiment of the present invention, described The compounds of this invention also comprises hydrate, ester or the prodrug of acceptable salt, salt on its pharmacodynamics.
The invention still further relates to and contain as the The compounds of this invention of active ingredient and the pharmaceutical composition of conventional medicine vehicle or assistant agent.Usually pharmaceutical composition of the present invention contains the The compounds of this invention of 0.1-95 weight %.The general content of The compounds of this invention is 0.1-100mg in unit dosage form, and preferred unit dosage form contains 4-50mg.
The pharmaceutical composition of The compounds of this invention can be according to method preparation well known in the art.When being used for this purpose, if desired, The compounds of this invention The compounds of this invention and one or more solids or liquid medicine vehicle and/or assistant agent can be combined, make and can be used as suitable administration form or the dosage form that people's medicine or veterinary drug use.
The compounds of this invention or contain its pharmaceutical composition can the unit dosage form administration, route of administration can be enteron aisle or non-enteron aisle, as oral, muscle, subcutaneous, nasal cavity, oral mucosa, skin, peritonaeum or rectum etc.
The compounds of this invention or the route of administration that contains its pharmaceutical composition can be drug administration by injection.Injection comprises intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection and acupoint injection therapy etc.
Form of administration can be liquid dosage form, solid dosage.As liquid dosage form can be true solution class, colloidal type, particulate formulations, emulsion dosage form, mixed suspension form.Other formulations are tablet, capsule, dripping pill, aerosol, pill, pulvis, solution, suspensoid, emulsion, granule, suppository, lyophilized injectable powder etc. for example.
The compounds of this invention can be made ordinary preparation, also can be sustained release preparation, controlled release preparation, targeting preparation and various particulate delivery system.
For example, can be extensive use of various carrier well known in the art for the unit form of administration is made tablet.Example about carrier is, for example thinner and absorption agent are as starch, dextrin, calcium sulfate, lactose, N.F,USP MANNITOL, sucrose, sodium-chlor, glucose, urea, lime carbonate, white bole, Microcrystalline Cellulose, pure aluminium silicate etc.; Wetting agent and tackiness agent are as water, glycerine, polyoxyethylene glycol, ethanol, propyl alcohol, starch slurry, dextrin, syrup, honey, glucose solution, mucialga of arabic gummy, gelatine size, Xylo-Mucine, lac, methylcellulose gum, potassiumphosphate, polyvinylpyrrolidone etc.; Disintegrating agent, for example dry starch, alginates, agar powder, laminaran, sodium bicarbonate and Citric Acid, lime carbonate, polyoxyethylene sorbitol fatty acid ester, sodium laurylsulfonate, methylcellulose gum, ethyl cellulose etc.; Disintegration inhibitor, for example sucrose, Tristearoylglycerol, theobroma oil, hydrogenation wet goods; Absorption enhancer, for example quaternary ammonium salt, sodium lauryl sulphate etc.; Lubricant, for example talcum powder, silicon-dioxide, W-Gum, stearate, boric acid, whiteruss, polyoxyethylene glycol etc.Tablet further can also be made coating tablet, for example sugar coated tablet, thin membrane coated tablet, ECT, or double-layer tablets and multilayer tablet.
For example, can be extensive use of various carrier well known in the art for pill is made in the administration unit.Example about carrier is, for example thinner and absorption agent are as glucose, lactose, starch, theobroma oil, hydrogenated vegetable oil, polyvinylpyrrolidone, Gelucire, kaolin, talcum powder etc.; Tackiness agent is as gum arabic, tragacanth gum, gelatin, ethanol, honey, liquid sugar, rice paste or batter etc.; Disintegrating agent is as agar powder, dry starch, alginates, sodium laurylsulfonate, methylcellulose gum, ethyl cellulose etc.
For example for capsule is made in the administration unit, the effective constituent The compounds of this invention is mixed with above-mentioned various carriers, and the mixture that will obtain thus places hard gelatine capsule or soft capsule.Also the effective constituent The compounds of this invention can be made microcapsule, be suspended in and form suspensoid in the aqueous medium, in the hard capsule of also can packing into or make injection and use.
For example, The compounds of this invention is made injection preparation, as solution, suspensoid solution, emulsion, lyophilized injectable powder, this preparation can be moisture or non-water, can contain acceptable carrier, thinner, tackiness agent, lubricant, sanitas, tensio-active agent or dispersion agent on a kind of and/or multiple pharmacodynamics.Can be selected from water, ethanol, polyoxyethylene glycol, 1 as thinner, the isooctadecanol of ammediol, ethoxylation, the isooctadecanol of polyoxyization, Polyoxyethylene Sorbitol Fatty Acid Esters etc.In addition, ooze injection liquid, can in injection preparation, add proper amount of sodium chloride, glucose or glycerine, in addition, can also add conventional solubility promoter, buffer reagent, pH regulator agent etc. in order to prepare etc.These auxiliary materials are that this area is commonly used
In addition, as needs, also can in pharmaceutical preparation, add tinting material, sanitas, spices, correctives, sweeting agent or other material.
For reaching the medication purpose, strengthen result of treatment, medicine of the present invention or pharmaceutical composition can be with any known medication administrations.
The dosage of The compounds of this invention pharmaceutical composition depends on many factors, for example to prevent or treat the character and the severity of disease, the sex of patient or animal, age, body weight, personality and individual reaction, route of administration, administration number of times, therapeutic purpose, therefore therapeutic dose of the present invention can have large-scale variation.In general, the using dosage of Chinese materia medica composition of the present invention is well known to a person skilled in the art.Can be according to the actual drug quantity that is contained in the preparation last in the The compounds of this invention composition, in addition suitable adjustment to reach the requirement of its treatment significant quantity, is finished prevention of the present invention or therapeutic purpose.The consumption of the suitable dose scope compound of the present invention of the every day of The compounds of this invention is the 0.001-100mg/Kg body weight, is preferably the 0.1-60mg/Kg body weight, and more preferably the 1-30mg/Kg body weight most preferably is the 2-15mg/Kg body weight.Be 10-500mg The compounds of this invention every day that adult patient is taken, and is preferably 20-100mg, can once take or divide and take for 2-3 time; The dosage of children taking is preferably the 10-20mg/kg body weight according to every kg body weight 5-30mg.Above-mentioned dosage can the single dose form or be divided into several, for example two, three or four dosage form administrations, this is subject to administration doctor's the clinical experience and the dosage regimen of treatment means.Compound of the present invention or composition can be taken separately, or merge use with other treatment medicine or symptomatic drugs.
Pharmaceutical research shows, The compounds of this invention has the nonspecific immunity enhancement, inhibiting effect on tumor metastasis is arranged, share the synergistic antitumor effect with antitumour drug, subcutaneous injection does not have influence to rabbit body temperature, ConA inductive mice spleen lymphocytes proliferation there is significant promoter action, can significantly promotes DC inductive cytotoxic T cell, can significantly promote the secretion of Turnover of Mouse Peritoneal Macrophages TNF-α and the secretion of DC cell IL-2, IL-12 and TNF-α the P815 cell killing activity.
Therefore compound of the present invention can be used for the immunotherapy of tumour, as adjuvant late tumor, acquired immune deficiency syndrome (AIDS), hemorrhagic shock, lymphoma, virus disease, autoimmune disease etc. is had treatment or assisting therapy effect and does not have pyrogenicity.
English list of abbreviations
BOP: benzotriazole-1-oxygen base-three (dimethylin) phosphine hexafluorophosphate
Bz: benzyl
DCC: dicyclohexylcarbodiimide
DIC: DIC
DCM: methylene dichloride
Dde:4,4-dimethyl hexamethylene-2,6 diketone-1-subunit-ethyl
DEPT: undistorted polarization transfer gain
D-iso-Gln:D-configuration isoglutamine
DMAP: Dimethylamino pyridine
DMF: dimethyl formamide
DMSO: dimethyl sulfoxide (DMSO)
ESI: electron spray ionisation
FAB: fast atom bombardment(FAB)
FCA: Fu Shi Freund's complete adjuvant
The Fmoc:9-fluorenylmethyloxycarbonyl
Fmoc-OSu: nitrogen maloyl imines-9-fluorenes methoxy carbonyl acid activation ester
HATU:2-(1H-9-azepine benzotriazole-1-yl)-1,1,3,3-tetramethyl-urea hexafluorophosphate
HBTU:2-(1H-benzotriazole-1-yl)-1,1,3,3-tetramethyl-urea hexafluorophosphate
HOAt:1-hydroxyl-7-azepine benzotriazole
The HOBt:1-hydroxy benzo triazole
HOSu: nitrogen maloyl imines
L-Ala:L-configuration L-Ala
L-Lys:L-configuration Methionin
MDP:N-ethanoyl Muramyl dipeptide
MDP-C:N
α-(1-α-benzyl-N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
ε-(3-nitrostyrolene base carbonyl)-lysyl amine
MDP-Lys (L18): N
α-(N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
ε-stearyl Methionin
PyBOP: benzotriazole-1-oxygen base-three (1-pyrrolidyl) phosphine hexafluorophosphate
TBTU: benzotriazole-1-oxygen base-N, N, N ', N '-tetramethyl-urea a tetrafluoro borate
THF: tetrahydrofuran (THF)
TFA: trifluoroacetic acid
TMS: tetramethylsilane
NMM: n-formyl sarcolysine base morpholine
ConA: companion's cona protein A
LPS: phosphoric acid lipopolysaccharides
MTT: Thiazolyl blue
RmGM-CSF: mGM-CSF
RmIL-4: recombined small-mouse interleukin 4
IL-2: interleukin-22
IL-12: interleukin 12
TNF-α: tumour necrosis factor alpha.
Embodiment
Synthetic
The present invention is further illustrated for following examples.But the present invention is not limited to these embodiment.
Embodiment 1:Fmoc protection D-isoGln's is synthetic
Get 60g D-L-glutamic acid and place the 1000mL there-necked flask, add 200 ~ 300mL, 1 ~ 4NNaOH solution, induction stirring, Dropwise 50 ~ 100mL benzyloxy dicarbonyl chloride and 120mL1 ~ 4NNaOH solution last 30 minutes approximately simultaneously, are heated to 60 ℃, stirring reaction 5 ~ 30 minutes.Add 10 ~ 50mL 4N NaOH again, continued stirring reaction 1 hour.It is organic to add ether (350mL * 4) extraction.The aqueous solution cools off with ice bath, with hcl acidifying pH=1 ~ 5.After ethyl acetate extraction (500mL * 4), use anhydrous magnesium sulfate drying.Steaming desolventizes, and gets white solid 110g.
Get the Z-D-Glu that 12.46g makes above and be dissolved among the anhydrous THF of 20 ~ 50mL, stir and add 10.98g DCC down, continue to stir 7 hours, precipitation in a large number occurs.Remove by filter insolubles, with heavily steaming THF washing precipitation and merging filtrate on a small quantity.Filtrate is transferred in the 250mL there-necked flask, and cryosel is bathed cooling and under mechanical stirring, was fed ammonia gas react 30 minutes, and solid methanol recrystallization gets colourless product.Use ethyl acetate extraction, the organic layer anhydrous magnesium sulfate drying.Concentrating under reduced pressure, recrystallization gets colorless solid 6.2g, fusing point 174-176 ℃.
Get the there-necked flask that 14g Z-D-iso-Gln places a 500mL, add 1g 5%Pd/C, 100 ~ 300mL water again, induction stirring, normal pressure hydrogenation is 1 ~ 2 day under the room temperature.Elimination Pd/C, and wash with less water.
Add saturated 100mLNaHCO
3With 200mL acetone.The a small amount of for several times down Fmoc-0Su that adds 5 ~ 10 times of molar excess of ice bath cooling, stirring reaction is after 24 hours, with 2 ~ 4N HCl acidifying pH to 2 ~ 4.Rotation is steamed and is removed organic solvent, with raffinate ethyl acetate extraction (500mL * 5), organic phase anhydrous magnesium sulfate drying.After steaming desolventized, the resistates recrystallizing methanol got product 15.35g.Fusing point 204-205 ℃, productive rate 41.8%.Mass spectrum (FAB+): nucleo plasmic relation (%): 369.1[M+H
+] (60%), 179.1[C
14H
11 +] (100%).Infrared: 735,756,1279,1319,1543,1635,1685,1741,3182,3313,3406.Nucleus magnetic hydrogen spectrum (300MHz, [D
6] DMSO, 25 ℃, mark in the TMS) :=1.75-1.89 (m, 2H, CH
2), 2.24 (t, J=7.8Hz, 2H; CH
2), 3.92 (m, 1H, CH), 4.19-4.27 (m, 3H, CHCH
2), 7.02 (s, 1H, CONH), 7.28-7.88 (m, 10H, 8 ph-H+CONH
2), 12.07 (s, 1H, COOH).
Embodiment 2: protect the synthetic of teichoic acid
(1) 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-α, β-D-glucoside preparation:
Get a suitable reaction flask, about 200 ~ 500mL concentrated hydrochloric acid of packing into slowly splashes into the vitriol oil from dropping funnel, and the HCl gas that will produce feeds ice bath down in the benzylalcohol of the new 1500 ~ 3000mL of steaming, is dissolved with to the benzylalcohol till 40 ~ 80g HCl.Get 2-acetylaminohydroxyphenylarsonic acid 2-deoxy-D-glucose 72g ~ 144g, place suitable reaction flask, add 1500 ~ 3000mL again and contain HCl benzylalcohol.Be heated to 85 ~ 120 ℃, stirring reaction is after 1 hour, reduces temperature to 40 ~ 70 and ℃ reacts 5 ~ 10 hours again.Stir down, add and contain 200 ~ 400g K
2CO
3Be dissolved in the solution of 300mL water, tell organic phase, with anhydrous Na
2SO
4Dry.Filter, in 75-100 ℃ with oil pump pressure reducing and steaming phenylcarbinol, add 1: 1 normal hexane of 2000ml and ether mixture in the resistates ,-10 ℃ of crystallizatioies filter.Solid is washed with ether, use ethyl alcohol recrystallization, get product 76g, productive rate 75%.
(2) 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 4,6-O-benzal-2-deoxidation-α-D-glucoside (2a) preparation
Get 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-α, β-D-glucoside 93g places the reaction flask of 1000mL, adds 150 ~ 300mL dehydrated alcohol, adds 600mL toluene again, and rotary evaporation removes and desolvates.Add the DMF 300 ~ 600mL that heavily steamed, the anhydrous dioxane 300 ~ 600mL that heavily steamed, triethyl orthoformate 150 ~ 300mL, phenyl aldehyde 120 ~ 240mL, tosic acid 7.5 ~ 15g.Stirring reaction added the absolute anhydrous diethyl ether of 750mL after 8 ~ 16 hours under the room temperature, and 0 ℃ was stirred 1 hour, and suction filtration washs solid with ether, and vacuum-drying gets 2a product 91g, productive rate 76.2%, thin-layer chromatography Rf value Rf=0.37 (developping agent CHCl
3: EtOH=20: 1).
(3) 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 4,6-O-benzal-2-deoxidation-3-O-[(R, S)-(2 '-methoxycarbonyl) ethyl]-α-D-glucoside (4r, s) preparation:
In 5 liters of there-necked flasks, 80g exsiccant 2a is dissolved in the anhydrous dioxane that 3000ml heavily steamed, stir, make dissolving.Divide to add 30 ~ 67.2g 50%NaH for several times 50 ~ 80 ℃ of stirring reactions 1 ~ 10 hour.Cool the temperature to 40 ~ 65 ℃ then.Get 64g 2-chloropropionic acid, be dissolved in the anhydrous dioxane that about 150 ~ 300ml heavily steamed, slowly drop in the reaction solution at 40 ~ 65 ℃, be warming up to 50 ~ 80 ℃, drip 640mL water, rotation is steamed and is desolventized to doing.Add 200 ~ 400mL ethanol, again evaporate to dryness.Add the saturated NaCl aqueous solution of 800ml to solid, ice bath was placed 12 hours.Suction filtration, filter cake is dissolved in filter cake in the 2400ml hot water with a small amount of saturated NaCl solution washing again, with ether (400ml * 3) extracting impurity.The aqueous solution is heated to 80 ℃, uses decolorizing with activated carbon, be concentrated into about 400ml, add NaCl 140g, stir, NaCl is dissolved as far as possible.Put 12 hours for 4 ℃, separate out solid Na salt, filter drying.
Get above-mentioned solid and be dissolved in the 800ml dry DMF, add the 56g methyl iodide, stirring at room three days, oil pump removes DMF under reduced pressure, and with less water washing, suction filtration, drying gets solid 88g, productive rate 90.5%.
(4) 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-4,6-O-benzal-3-O-[(R, S)-(2 '-methoxycarbonyl) ethyl]-α-D-glucoside (4r s) splits:
Get 4r, s 8g dissolves with the 200ml methylene dichloride, and 200-300 order silica gel 40g mixes sample.Get the high 60cm of post, the glass chromatography column of internal diameter 5.5cm is mixed 400g (200-300 order) silica gel and 500ml methylene dichloride thoroughly, wet method dress post.After having adorned, standing over night.With following elutriant wash-out successively: ethyl acetate/CH
2Cl
2=0% (300ml), 15% (21), 20% (21), 25% (11), 40% (500ml), ethyl acetate (11).Collect the about 100ml/ bottle of drip, detect with HPLC, merge respectively and contain 4r, 4s flows part.Concentrate teichoic acid methyl esters 4.5g, yield 56.3%, different teichoic acid methyl esters 2.3g, yield 28.8%.
(5) 1-benzyl-2-acetylaminohydroxyphenylarsonic acid 2-deoxidation-4, the 6-O-benzal-3-O-[(R)-2 '-carboxy ethyl]-α-D-glucoside (5r) preparation:
4r 13.5g is dissolved in the 200ml dioxane at 40 ℃, stirs down and slowly drip 135ml 0.5 ~ 2N NaOH aqueous solution, TLC detection reaction situation was reacted after 12 hours, and saponification is thorough.The rotary evaporation solvent adds to solid in the 550ml water, and cryosel is bathed cooling, slowly drips about 750 ~ 1500ml, 1 ~ 2N HCl aqueous solution, o'clock stops to pH=3.0 ~ 40, and this moment, reaction solution became white gels shape solid.Suction filtration 12 hours, room temperature is dried naturally, and vacuum-drying with 400ml dehydrated alcohol recrystallization, gets 5r 12.7g, and fusing point 231-233 ℃, productive rate: 96.9%.
Embodiment 3: the solid phase synthesis compound N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-L-lysyl amine (X=NH
2)
The resin Crown that gets 5～8 μ mol takes off the Fmoc protection with 50% piperidines/DMF solution.With the Acibenzolar that Fmoc-Lys (Dde)-OH amino acid is made with TBTU and HOBt, reacted 24 hours down at 25 ℃ then with Crown.And then take off the Fmoc protecting group with 50% piperidines/DMF solution, connect D-Fmoc-isoGln-OH, Fmoc-Ala-OH and protection teichoic acid with active ester method equally.Use 30～60% anhydrous hydrazines/DMF solution to remove the Dde protecting group after 8 hours again, in polar solvent (as water), reacted 48 hours with 3-nitrocinnamic acid Acibenzolar.With the 10ml 20%TFA-aqueous solution synthetic compound is cut from resin at last and fallen.Nitrogen dries up, and removes the benzal protecting group with 10～30% HCl again, after nitrogen dries up, again with sherwood oil-ether-methylene dichloride-hexanaphthene mixed solvent washing three times, after the lyophilize N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-L-lysyl amine.The high-efficient liquid phase color spectral purity that obtains compound through C18 reversed phase column chromatography purifying is 95% (methyl alcohol: water=70: 30).Molecular composition is: C
41H
56N
8O
14, ESI mass spectrum: 885.2[M+H]
+Nucleus magnetic hydrogen spectrum (DMSO-d
6): 1.22 (d, J=8.0,3H), 1.23 (d, J=7.5,3H), 1.20--1.52 (m, 5H), 1.60--1.71 (m, 2H), 1.77 (s, 3H), 1.94 (hex, J=7.0,1H), 2.13 (t, J=7.0,2H), 3.15 (d, J=6.0,1H), 3.28 (t, J=9.0,1H), 3.47--3.51 (m, 3H), 3.63 (d, J=10.5,1H), 3.64 (d, J=10.5,1H), 3.81 (m, 1H), 4.12 (m, 2H), 4.25 (m, 2H), 4.43 (d, J=12.5,1H), 4.66 (d, J=12.5,1H), 4.72 (d, J=3.0,1H), 6.79 (d, J=16.0,1H), 6.94 (br, s, 1H, D
2O is commutative), 7.03 (br, s, 1H, D
2O is commutative), and 7.27--7.37 (m, 5H), 7.31 (br, s, 1H, D
2O is commutative), 7.52 (d, J=16.0,1H), 7.53 (br, s, 1H, D
2O is commutative), 7.69 (t, J=7.8,1H), 7.86 (d, 1H, D
2O is commutative), 8.00 (d, J=8.0,1H), 8.09 (d, 1H, D
2O is commutative), 8.13 (d, J=8.0,1H), 8.17 (d, 3H, D
2O is commutative), 8.37 (br, s, 1H).Nuclear-magnetism carbon spectrum and DEPT spectrum (125M, DMSO-d
6): 18.39 (CH
3), 19.00 (CH
3), 22.63 (CH
3), 23.02 (CH
2), 27.85 (CH
2), 28.82 (CH
2), 31.61 (CH
2), 31.73 (CH
2), 38.72 (CH
2), 48.20 (CH), 52.14 (CH), 52.40 (CH), 52.98 (CH), 60.53 (CH), 67.92 (CH
2), 69.52 (CH), 73.20 (CH), 76.50 (CH), 79.16 (CH), 95.98 (CH), 121.54 (CH), 123.68 (CH), 125.25 (CH), 127.54 (CH), 127.63 (2CH), 128.25 (2CH), 130.52 (CH), 133.86 (CH), 136.18 (CH), 136.88 (C), 137.74 (C), 148.33 (C), 164.34 (C=O), 169.67 (C=O), 171.61 (C=O), 172.10 (C=O), 172.71 (C=O), 173.30 (C=O), 173.98 (C=O).
Infrared (KBr, cm
-1): 698,735,808,976,1045,1122,1230,1352,1454,1529,1658,2870,2933,3288
Embodiment 4: the solid phase synthesis compound N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl) N
ε-(3-nitro-trans styryl carbonyl)-L-lysyl amine (X=NH
2)
The Fmoc:9-fluorenylmethyloxycarbonyl
Fmoc-OSu: nitrogen maloyl imines-9-fluorenes methoxy carbonyl acid activation ester
Take by weighing the Rink Amide resin (perhaps Rink MBHAAmide resin) of 200mg Fmoc protection; charge capacity is 0.58mmol/g; 20% piperidines/DMF with 5～10mL takes off Fmoc protection 2～3 hours; again with after the DMF washing 10 times; the DMF solution that adds 20～50 times of molar excess Fmoc-Lys (Dde)-Obt Acibenzolar, 25 ℃～80 ℃ were reacted 5～10 hours down.Repeat to take off the condensation course of Fmoc protecting group and Fmoc-D-isoGln-Obt, repeat to take off the condensation course of Fmoc protecting group and Fmoc-Ala-Obt again, until to the condensation course that repeats to take off Fmoc protecting group and protection protection cell wall sugar.Use 2%～20% NH at last
2NH
2After/DMF the solution-treated 2 minutes～30 minutes, the polar solvent (as water) that adds the 3-nitrocinnamic acid Acibenzolar of 1～5 times of molar excess reacted 48 hours.The compound cracking is identical with embodiment 3, the compd E SI mass spectrum that liquid phase-mass spectrometry system provides: 885.1[M+H]
+
Embodiment 5: the solid phase synthesis compound N
α-(1-α-benzyl-N-ethanoyl-muramyl-L-alanyl-D-isoglutamine acyl)-N
ε-(3-nitro-trans styryl carbonyl)-L-Methionin (X=OH)
Take by weighing 200mg Wang resin, charge capacity is 0.60mmol/g, in the DCM of the DMAP of 5 times of molar excess catalyzer and 100mL, reacted 3～5 days with Fmoc-Lys (Dde)-Obt Acibenzolar, use 20% piperidines/DMF of 5～10mL to handle then 2～3 hours, again with after the DMF washing 10 times, the DMF solution that adds 20～50 times of molar excess Fmoc-D-isoGln-Obt Acibenzolars, 25 ℃～80 ℃ were reacted 5～10 hours down.Repeat to take off the condensation course of Fmoc protecting group and Fmoc-Ala-Obt.Use 2%～20% NH at last
2NH
2After/DMF the solution-treated 2 minutes～30 minutes, the polar solvent (as water) that adds the 3-nitrocinnamic acid Acibenzolar of 1～5 times of molar excess reacted 48 hours.The compound cracking is identical with embodiment 3.Liquid phase-mass spectrometry system records the ESI mass spectrum of synthetic purpose compound: 886.3[M+H]
+
Pharmacological evaluation
The external biological screening experiment of experimental example 1:MDP-C immuno-potentiation and animal as a result thereof: male mice in kunming is provided by the court's animal center.
Experiment material: MTT is available from sigma company; P388 is a mouse lymph like cell knurl, is provided by the Shanghai cell bank.
Experimental technique: scavenger cell-mtt assay, action time: 72 hours, cell category: P388, sample concentration: 10
-5M
Get Kunming mouse number, every abdominal injection 1% starch fluid (physiological saline is joined) 2mL, in the 4th day sacrificed by exsanguination, sterilization skin, open the abdominal cavity, to every cold RPMI-1640 in the injection 5mL left and right sides, extract abdominal cavity water to aseptic silication test tube by rubbing the back gently, with asepsis injector to reduce sticking of scavenger cell.After cell washed 2 times with nutrient solution, it was 4 * 10 that counting is adjusted concentration
6/ mL.Get 96 well culture plates, every hole adds scavenger cell and tumour cell 50 a μ L, sample solution 10 μ L, and each concentration is done 3 multiple holes.At 5%CO
2, cultivated under 37 ℃ the condition 72 hours, add 20 μ L 5%MTT/ holes then, add 50 μ L 10%SDS-5% isopropylcarbinol-0.02mol HCl three liquid/holes after 3～5 hours again, 37 ℃ after night, surveys the O.D.570 value, the results are shown in Table 1.
Table 1 MDP derivative immunocompetence The selection result
Sample OD570X ± SD inhibiting rate (%) t check
Contrast 0.41 ± 0.02
MDP 0.25±0.03 56.2 P＜0.001
MDP-Lys 0.24±0.03 53.0 P＜0.001
(L18)
MDP-C 0.29±0.13 71.3 P＜0.001
Brief summary: the nonspecific immunity enhancement of this experiment detection compound, its result show that compound MDP-C may work in the immunotherapy of tumour.
Inhibiting effect on tumor metastasis in the animal body of experimental example 2 MDP-C
Animal: male C57BL/6 is provided by the court's animal center.
Experiment material: the preparation of sample and laboratory sample thereof: MDP-C, natural Japanese yew alcohol is all used the tween-80 hydrotropy, prepares suspension with physiological saline.
Experimental technique: get the strain of LLC knurl, add 2ml physiological saline by the 1g knurl and wear into homogenate, in mouse back leg intramuscular inoculation 0.1ml knurl liquid.Inoculate back 24 hours with mouse by the administration of body weight random packet.Experiment is established 4 groups altogether:
1. control group: the solvent of giving equal volume
2. taxol 3.5mg/kg organizes;
3.MDP-C 1.25mg/kg group;
4.MDP-C 1.25mg/kg+ taxol 3.5mg/kg group;
Be abdominal injection, the administration volume is 10mL/kg, once a day, and continuous 21 days.Inoculation back the 9th day, behind vetanarcol 35mg/kg anesthetized mice, with the amputation of lotus knurl leg, operation back intramuscular injection penicillin 100,000 units/kg.Put to death mouse on the 22nd day in the inoculation back, get lung, weigh, fixing, counting lung cancer metastasis kitchen range.(preparation of stationary liquid: glacial acetic acid: 2mL, picric acid 15mL, formalin 1mL) the results are shown in Table 2.
In the animal body of table 2 MDP-C the effect of anti-Lewis lung cancer metastasis and with the synergy of taxol
Metastasis (individual) SD (standard deviation) inhibiting rate (%)
Blank 12.0 3.9
MDP-C 5.2 6.5 56.6％
Taxol 3.8 3.1 67.3%
Taxol+MDP-C 3.2 3.4 73.3%
Brief summary: the result shows that MDP-C has inhibiting effect on tumor metastasis, has share the synergistic antitumor effect with taxol.
Embodiment 3:MDP-C is to the influence (the results are shown in Table 3) of rabbit body temperature
Animal: 20 of rabbit are provided by the court's animal center.
Experimental technique: be divided into two groups at random:
(1) .MDP-C 0.1mg/kg group;
(2) .MDP-C 0.2mg/kg group;
1. by body weight administration (seeing the following form)
Then subcutaneous injection skim-milk emulsion (skim-milk/water is 1/7, w/v) 2mL/kg.
3. measure anus temperature, per hour 1 time, continuous 4 times with thermometer after the pyrogenicity.
4. measuring 24 hours anus temperature after the pyrogenicity next day, is basal body temperature with the anus temperature before the administration, surveys the anus temperature with different time points, the results are shown in Table 3.
Table 3 compound MDP-C is to the influence of rabbit body temperature
Different time after the rabbit pyrogenicity of compound basis (hour) body temperature (℃)
MDP-C body temperature 1234 24
0.1 39.93 40.20 39.88 39.92 39.88 39.62
(mg/kg) ±0.24 ±0.97 ±0.41 ±0.52 ±0.34 ±0.31
0.2 39.80 39.65 39.62 39.92 39.93 39.57
(mg/kg) ±0.23 ±0.36 ±0.26 ±0.28 ±0.26 ±0.33
Brief summary: results suggest MDP-C 0.1, under the 0.2mg/kg dosage subcutaneous injection rabbit body temperature is not had influence, show its no pyrogenicity.
Embodiment 4:MDP-C is to the influence of ConA inductive mice spleen lymphocytes proliferation
Animal: male BALB/c mouse is provided by the court's animal center.
Experiment material: MTT, ConA and LPS are all available from sigma company.
Experimental technique:
1. the aseptic mouse spleen lymphocyte of getting, cultivating the keynote cell concn with complete RPM1-1640 is 5 * 10
6Cells/mL.
2. add ConA (2 μ g/mL) and different compounds respectively, at 5%CO
2, cultivated 72 hours in the humidity incubator fully.
3.MTT method is surveyed splenic lymphocytes, the results are shown in Table 4.
Table 4 compound MDP-C is to the influence of ConA inductive mice spleen lymphocytes proliferation
MDP-C concentration OD
570Rate of increase (%)
(mol/L)
1×10
-5 1.150±0.036 158.9
***
1×10
-6 1.292±0.032 191.1
***
1×10
-7 0.778±0.007 75.28
***
Blank group 0.444 ± 0.003
ConA(2μg/mL) 1.214±0.010
* *, expression P＜0.001
Brief summary: results suggest compound MDP-C has significant promoter action to ConA inductive mice spleen lymphocytes proliferation.
Embodiment 5:MDP-C to DC inductive cytotoxic T cell to the P815 cell killing
Active influence
Animal: male BALB/c mouse and C57BL/6 mouse are provided by the court's animal center.
Experiment material: rmGM-CSF, rmlL-4 and silk split the plain C of enzyme all available from sigma company.Non-radioactive Cytotoxicity 96 CTL test kits are purchased the company in Promega.The P815 cell is the loose oncocyte of mouse, and Nat'l Pharmaceutical ﹠ Biological Products Control Institute is so kind as to give.
Experimental technique:
1. the aseptic bone marrow cells in mice of getting, cultivating the keynote cell concn with complete RPM1-1640 is 5 * 10
6Cells/mL.
2. substratum is added rmGM-CSF (5ng/mL) and rmlL-4 (5ng/mL).Half amount is changed liquid every other day.
3. added P815 cell freeze thawing antigen and MDP-C on the 5th day in cultivation, and cultivated 18 hours.
4. split the plain C of enzyme with 25 μ g/mL silks and handled the DC cell 30 minutes, by 10
3Cells/well is connected to 24 orifice plates (Costr).
5. the aseptic mouse T lymphocyte of getting, cultivating the keynote cell concn with complete RPM1-1640 is 1 * 10
7Cells/mL is inoculated in above-mentioned orifice plate, 5%CO by 1mL/well
2, cultivated 5 days in the humidity incubator fully.
6. get supernatant,, the results are shown in Table 5 with Non-radioactive Cytotoxicity 96 CTL kit measurement CTL activity.
Table 5MDP-C is to the influence of DC inductive cytotoxic T cell to the P815 cell killing activity
MDP-C specificity release rate increment rate
Concentration (mol/L) is (%) (%)
1×10
-5 37.7 243.2
1×10
-6 45.2 311.1
1×10
-7 29.4 167.8
Blank group 11.0 ± 0.027
LPS(1.0μg/mL) 24.6±0.055
Brief summary: results suggest MDP-C can significantly promote DC inductive cytotoxic T cell to the P815 cell killing activity.Can example 4 and 5 all be to detect the important evidence that treat immune deficiency disorder, and its result shows that MDP-C may have treatment or assisting therapy effect to late tumor and acquired immune deficiency syndrome (AIDS) etc. as adjuvant.
Embodiment 6:MDP-C influences Turnover of Mouse Peritoneal Macrophages TNF-α excretory
Animal: male C57BL/6 mouse is provided by the court's animal center.
Experiment material: mouse TNF-α ELISA test kit is purchased the company in Diaclone.
Experimental technique:
1. the aseptic Turnover of Mouse Peritoneal Macrophages of getting, cultivating the keynote cell concn with complete RPM1-1640 is 5 * 10
6Cells/mL.
2. add LPS (1.0 μ g/mL) and different compounds respectively, at 5%CO
2, cultivated 24 hours in the humidity incubator fully, get supernatant.
3. detect the content of TNF-α in the supernatant with TNF-α E LlSA method, the results are shown in Table 6.
Table 6MDP-C influences Turnover of Mouse Peritoneal Macrophages TNF-α excretory
MDP-C mean value ± SD
Concentration (mol/L) is of TNF-α rate of increase (%) (pg/mL)
2.5×10
-6 26.341±0.003 482.4
***
1×10
-6 14.295±0.001 216.1
***
1×10
-7 11.114±0.001 145.7
**
Blank group 4.523 ± 0.001
LPS(1.0μg/mL) 31.568±0.003
Brief summary: results suggest MDP-C significantly promotes the secretion of Turnover of Mouse Peritoneal Macrophages TNF-α, may treatment or assisting therapy effect be arranged to diseases such as acquired immune deficiency syndrome (AIDS), hemorrhagic shock and lymphomas.
Embodiment 7:MDP-C is to the influence of mouse dcs (DC) cytokine secretion
Animal: male C57BL/6 mouse is provided by the court's animal center.
Experiment material: mouse TNF-α ELlSA test kit is purchased the company in Diaclone.IL-2 and IL-12 ELISA test kit are all purchased in Shenzhen U.S. biotechnology of crystalline substance company limited.
Experimental technique:
1. the aseptic bone marrow cells in mice of getting, cultivating the keynote cell concn with complete RPMI-1640 is 5 * 10
6Cells/mL.
2. substratum is added rmGM-CSF (5ng/mL) and rmIL4 (5ng/mL).Half amount is changed liquid every other day.
3. added the different compounds of LPS (1 μ g/mL) and different concns on the 5th day.
4. collected culture supernatant on the 7th day.
5. detect its secretion level with IL-2, IL-1 2 and TNF-α ELISA method respectively, the results are shown in Table 7.
Table 7 compound MDP-C is to the influence of mouse dcs (DC) cytokine secretion
MDP-C IL-2 IL-12 TNF-α
Concentration (mol/L)
Mean value ± SD rate of increase mean value ± SD rate of increase mean value ± SD rate of increase
(pg/mL) (％) (pg/mL) (％) (pg/mL) (％)
1×10
-5 102.5±0.036 142.2
*** 918.2±0.001 65.1
*** 14.636±0.001 163.9
**
1×10
-6 115.5±0.026 172.8
*** 1103.5±0.127 98.4
*** 16.341±0.006 194.7
**
1×10
-7 75.3±0.032 77.8
** 757.9±0.059 36.3
*** 12.023±0.007 116.8
*
Blank group 42.3 ± 0.025 556.3 ± 0.002 5.545 ± 0.002
LPS 95.9±0.055 1126.6±0.201 42.705±0.003
(1.0μg/mL)
Annotate:
*P＜0.05,
*P＜0.01,
* *P＜0.001 is with blank group ratio
Brief summary: results suggest MDP-C significantly promotes the secretion of mouse DC cell IL-2, IL-12 and TNF-α.May in the treatment of some virus disease, autoimmune disease and late tumor, have certain effect.
Claims (15)
1, the compound shown in the formula (I), and hydrate, ester or the prodrug of acceptable salt, salt on the pharmacodynamics.
X=NH
2, perhaps OH
(I)。
2, prepare the method for the described compound of claim 1, comprise the steps:
(A) Fmoc protection D-isoGln's is synthetic,
(B) the synthetic and fractionation of protection teichoic acid,
(C) N
α-(1-α-benzyl-N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
εThe solid phase synthesis of-(3-nitro-trans styryl carbonyl)-lysyl amine;
It is characterized in that:
Fmoc protection D-isoglutamic acid synthetic is that the hydroxyl with the carboxylic acid of the amido end of D-L-glutamic acid changes amido into; simultaneously with original amido in the Fmoc protection D-L-glutamic acid; and hydroxyl is protected the amido of D-L-glutamic acid before changing amido into, and hydroxyl removes protecting group after changing amido into.
According to the preparation method of claim 2, it is characterized in that 3, the hydroxyl of the carboxylic acid of the amido end of described D-L-glutamic acid is protected the amido of D-L-glutamic acid before changing amido into, used protecting group is a carbobenzoxy-(Cbz).
According to the preparation method of claim 3, it is characterized in that 4, described what remove that protecting group uses is the hydrogenant method.
According to the preparation method of claim 4, it is characterized in that 5, described hydrogenant method is a palladium carbon catalytic hydrogenation.
6, according to the preparation method of claim 2, it is characterized in that, the hydroxyl of the carboxylic acid of the amido end of described D-L-glutamic acid change into amido be by with the D-L-glutamic acid of carbobenzoxy-(Cbz) protection under the condition that DCC exists and ammonia gas react.
According to the preparation method of claim 2, it is characterized in that 7, the fractionation of described protection teichoic acid is to adopt the method for column chromatography to obtain the teichoic acid methyl esters.
8, according to the preparation method of claim 2, it is characterized in that described N
α-(1-α-benzyl-N-ethanoyl-muramyl-D-alanyl-L-isoglutamine acyl)-N
ε-(3-nitro-trans styryl the carbonyl)-solid phase synthesis of lysyl amine is to connect resin, Fmoc-Lys (Dde)-OH, Fmoc-D-isoGln-OH, Fmoc-Ala-OH and protection teichoic acid successively; again with 3-nitrocinnamic acid Acibenzolar reaction, again the synthetic compound is cut to fall from resin and remove the benzal protecting group at last.
9, preparation method according to Claim 8 is characterized in that, described resin is selected from the Crown resin, Rink Amide resin, Rink MBHA Amide resin, Wang resin, and the resin that produces carboxyl or amide group after other cracking.
10, preparation method according to Claim 8; it is characterized in that; the condensing agent that adopts when described connection resin, Fmoc-Lys (Dde)-OH, Fmoc-D-isoGln-OH, Fmoc-Ala-OH and protection teichoic acid is selected from HBTU; TBTU, HATU, BOP; PyBrBOP; DCC, DIC, and other condensing agent for synthesizing polypeptide.
11, according to the preparation method of claim 3; it is characterized in that; the additive that adopts when described connection resin, Fmoc-Lys (Dde)-OH, Fmoc-D-isoGln-OH, Fmoc-Ala-OH and protection teichoic acid is selected from HOBt; HOAt; HOSu, and the synthetic additive that is used to form Acibenzolar of other polypeptide.
12, a kind of pharmaceutical composition is characterized in that, contains the compound as claimed in claim 1 of effective dose, and acceptable carrier on the pharmacodynamics.
According to the pharmaceutical composition of claim 12, it is characterized in that 13, described pharmaceutical composition can be tablet, capsule, pill, injection, sustained release preparation, controlled release preparation and various particulate delivery system.
14, the application of claim 1 compound in preparation medicine for treating tumor thing.
15, the application in the adjuvant medicine of treatment of claim 1 compound or auxiliary treatment AIDS, hemorrhagic shock, lymphoma, virus disease, autoimmune disease.
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| WO2011147330A1 (en) | 2010-05-27 | 2011-12-01 | 中国医学科学院药物研究所 | Chemical synthesis and anti-tumor and anti-metastatic effects of dual functional conjugate |
| WO2017148416A1 (en) * | 2016-03-03 | 2017-09-08 | 凯惠科技发展(上海)有限公司 | Method for preparing maytansine esters and intermediate thereof |
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| EP3672939A1 (en) | 2017-08-21 | 2020-07-01 | Celgene Corporation | Processes for preparation of (s)-tert-butyl 4,5-diamino-5-oxopentanoate |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2011147330A1 (en) | 2010-05-27 | 2011-12-01 | 中国医学科学院药物研究所 | Chemical synthesis and anti-tumor and anti-metastatic effects of dual functional conjugate |
| US9085605B2 (en) | 2010-05-27 | 2015-07-21 | Shenzhen Salubris Pharmaceuticals Co., Ltd. | Chemical synthesis and anti-tumor and anti-metastatic effects of dual functional conjugate |
| EP3239139A1 (en) | 2010-05-27 | 2017-11-01 | Shenzhen Salubris Pharmaceuticals Co., Ltd | Chemical synthesis and anti-tumor and anti-metastatic effects of dual functional conjugate |
| WO2017148416A1 (en) * | 2016-03-03 | 2017-09-08 | 凯惠科技发展(上海)有限公司 | Method for preparing maytansine esters and intermediate thereof |
| CN107417701A (en) * | 2016-03-03 | 2017-12-01 | 凯惠科技发展(上海)有限公司 | A kind of Preparation Method And Their Intermediate of maytansine ester |
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