CN105646675A - Coibamide A analog, synthesis method and applications thereof - Google Patents
Coibamide A analog, synthesis method and applications thereof Download PDFInfo
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- CN105646675A CN105646675A CN201510648096.7A CN201510648096A CN105646675A CN 105646675 A CN105646675 A CN 105646675A CN 201510648096 A CN201510648096 A CN 201510648096A CN 105646675 A CN105646675 A CN 105646675A
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Abstract
The invention provides a coibamide A analog, a synthesis method and applications thereof. The synthesis method comprises the following steps: providing hydrazine resin; adopting an activator, carrying out solid phase synthesis to obtain a cyclization precursor, which is represented by a formula (from the formula A to the formula F) and is loaded on hydrazine resin; carrying out cutting reactions in the presence of an oxidizing agent to separate polypeptide chains of the cyclization precursor from the resin so as to obtain a polypeptide chain cyclization precursor; removing the protective group P<NH> on the N terminals of the polypeptide chain cyclization precursor, then adopting a condensing agent to carry out ring-closure reactions so as to obtain a compound represented by the formula I; and the formula I and the formulas A to F are represented in the description. The invention also provides a coibamide A isomer represented by the formula I-1-1, which is represented in the description. The isomer has an excellent activity on inhibiting excessive proliferation of cells.
Description
Technical field
The present invention relates to Ke Yiba peptide A analogue and synthetic method thereof and application, belongs to Solid-phase organic synthesis field.
Background technology
US20120028905A1 discloses natural cyclic depsipeptide Ke Yiba peptide A (coibamideA) of a kind of height N-methyl that separation and purification obtains from Panama marine cyanobacterium Leptolyngbyasp. such as McPhail, lung cancer, mammary cancer, melanoma, leukemia and nervus centralis cancer etc. are all had the cytotoxicity of low nmole by it, mammary gland, nervus centralis and ovarian cancer cell are had good tissue selectivity simultaneously. Research shows that coibamideA carrys out anticancer propagation by a kind of new mechanism of action, therefore, and can using the primer of coibamideA as a kind of cancer therapy drug with good prospect. But, coibamideA ten points owing to existing in nature is limited, separation and purification coibamideA from algae is adopted to need complicated technique, a large amount of organic solvents, the longer time, and the receipts rate of coibamideA is very low, research and the demand of application cannot be met, such as, cannot meet the demand that coibamideA is carried out relevant pharmacodynamics assessment.
HeWei (TotalsynthesisofproposedstructureofcoibamideA in 2014, ahighlyN-andO-methylatedcytotoxicmarinecyclodepsipeptide .TetrahedronLett.2014,55,6109-12) the reported first total synthesis method of a kind of coibamideA, the method adopts the fragment assembly strategy of [(4+1)+3+3], by multistep liquid phase reaction, synthesize coibamideA. But this synthetic method needs multiple condensation reagent, length consuming time, and owing to there is amino acid whose racemization phenomenon in polypeptide condensation course, thus combined coefficient is very low.
Compared to time-consuming liquid-phase synthesis process, adopt solid-phase peptide synthesis (SPPS) that the synthesis of coibamideA can be made easy, quick. 2015, (Synthesisandbiologicalevaluationofthe [the D-MeAla such as Nabika11]-epimerofcoibamideA.Bio.Med.Chem.Lett.2015,25,302-6.) taking TCP resin as solid phase carrier, the cyclisation precursor comprising whole amino acid unit has been synthesized by Fmoc strategy, with TFA/DCM (5:95, v/v) after excising resin, finally by the isomer carrying out macrolide Reactive Synthesis 11 MeAla in the solution and occurring isomerized coibamideA, in cyclization, cyclisation receipts rate only 3.8%, calculate with the substitution value of first amino, synthesize the total recovery only 1% of the isomer of this coibamideA.
In summary, the synthetic method length consuming time of the coibamideA that prior art provides, receipts rate are low, and owing to can there is racemization in reaction process, also are difficult to guarantee that synthesized molecule is designed target molecule. Therefore develop a kind of method of efficient, Fast back-projection algorithm coibamideA or coibamideA isomer or its analogue, it is this area technical problem urgently to be resolved hurrily with the demand of satisfied research and application.
Summary of the invention
An object of the present invention is to provide the synthetic method of a kind of Ke Yiba peptide A (coibamideA) analogue, and this synthetic method can synthesize designed Ke Yiba peptide A analogue fast and efficiently.
Another object of the present invention is to provide the isomer of a kind of Ke Yiba peptide A, it is active that this isomer has good anti-cell hyper-proliferative.
It is still another object of the present invention to provide the application of the isomer of described Ke Yiba peptide A.
For achieving the above object, on the one hand, the present invention provides the synthetic method of a kind of Ke Yiba peptide A analogue, and this Ke Yiba peptide A analogue has the structural formula shown in formula I:
In formula I, R1��R2��R3��R5��R7��R8��R9��R11��R13��R14It is hydrogen, methyl or ethyl independently of one another;
R4For methoxy-benzyl, ethoxy benzyl, methylamino benzyl or ethylamino-benzyl;
R6��R10It is normal-butyl, isobutyl-or sec-butyl independently of one another;
R12For methoxymethyl, ethoxyl methyl, methoxy ethyl or ethoxyethyl group;
G be hydrogen, methyl,
Or amino protecting group; This amino protecting group is Boc, Fmoc, Cbz, Trt or Allyl;
R15��R21��R22It is n-propyl, sec.-propyl, isobutyl-or sec-butyl independently of one another;
R16��R17��R19��R20��R23��R24It is hydrogen, methyl, ethyl or amino protecting group independently of one another;
R18For being methoxymethyl, ethoxyl methyl, methoxy ethyl or ethoxyethyl group independently of one another;
Y is hydroxyl, methoxyl group, amino, methylamino-or two methylamino;
X is O, N or N (CH3);
Preferably, formula I has the structure shown in formula I-1:
More preferably, formula I-1 has the structure shown in formula I-1-1 or formula I-1-2:
Described method comprises the steps:
A., hydrazine resin is provided;
B. activator is adopted, the cyclisation precursor that in Solid-phase organic synthesis formula A��formula F, any one hydrazine is resin-carried:
The PNH of formula A��formula F in any one represents amino protecting group; This amino protecting group is Boc, Fmoc, Cbz, Trt or Allyl, it is preferable that Fmoc;
C. carrying out cleavage reaction under the existence of oxygenant, polypeptide chain in the cyclisation precursor making gained hydrazine in step b resin-carried and resin isolation, obtain polypeptide chain cyclisation precursor;
Preferably, the one of carbon tip of described polypeptide chain cyclisation precursor is carboxyl;
D. de-except, after the protecting group PNH on nitrogen end on gained polypeptide chain cyclisation precursor in step c, adopting condensing agent to carry out ring-closure reaction, obtain compound shown in formula I.
The present invention study find step b Chinese style A��formula F the resin-carried cyclisation precursor of hydrazine after dicing, can effectively carry out ring-closure reaction, significantly improve the product rate of synthesis Ke Yiba peptide A analogue, when method of the present invention being applied to synthesis Ke Yiba peptide A or its steric isomer, calculate with the substitution value of first amino, its product rate up to 8%��20%, can be far superior to the Ke Yiba peptide A of prior art offer or the synthetic route of its steric isomer.
Preferably, in the step b of the method for the invention, adopt activator, the cyclisation precursor that the hydrazine shown in Solid-phase organic synthesis formula D is resin-carried;
More preferably, Solid-phase organic synthesis is such as formula the resin-carried cyclisation precursor of the hydrazine shown in D-1:
Further preferably, the cyclisation precursor that hydrazine shown in Solid-phase organic synthesis formula D-1-1 or formula D-1-2 is resin-carried:
According to specific embodiment of the invention scheme, in method of the present invention, the resin-carried cyclisation precursor of the hydrazine shown in formula D-1 prepares as follows:
B-1. activator is adopted, the main chain that the hydrazine shown in Solid-phase organic synthesis formula D-1-F1 is resin-carried:
Preferably, the main chain that hydrazine shown in Solid-phase organic synthesis formula D-1-1-F1 or D-1-2-F1 is resin-carried:
P described in formula D-1-F1, formula D-1-1-F1 or formula D-1-2-F1OHFor hydroxyl protecting group, it is preferable that the tertiary butyl, trityl, allyl group or benzyl;
B-2. adopt and go protecting group agent to take off except the hydroxyl protecting group P on the resin-carried main chain of step b-1 gained hydrazineOH;
B-3. the hydroxyl exposed in step b-2 carries out esterification with the structure shown in obtained formula D-1-F2:
De-except the amino protecting group P in formula D-1-F2NHAfter, continue coupling and form the resin-carried cyclisation precursor of the hydrazine shown in formula D-1;
Preferably, the hydroxyl exposed in step b-2 carries out esterification with the structure shown in obtained formula D-1-1-F2 or formula D-1-2-F2:
The de-amino protecting group P except formula D-1-1-F2 or formula D-1-2-F2NHAfter, continue coupling to form the resin-carried cyclisation precursor of the hydrazine shown in formula D-1-1 or formula D-1-2.
By selecting, specific resin and protecting group strategy make step b-2 de-except P in the present inventionOHWhen do not make the connecting key of resin and peptide chain rupture, effectively synthesized the cyclisation precursor that the hydrazine shown in formula D-1 is resin-carried.
Those skilled in the art should know, and when the amino acid adopting a series of configuration to determine, the synthetic method of application the present invention can prepare Ke Yiba peptide A, or a certain Ke Yiba peptide A isomer or its analogue determining configuration. In step b-1, such as select the amino acid that a series of configuration is determined, adopt activator, Solid-phase organic synthesis formula D-1-1-F2, and in step b-3, form formula D-1-2, cutting can prepare the Ke Yiba peptide A shown in the formula I-1-2 determining configuration disclosed in prior art after closing ring; In step b-1, such as select again the amino acid that a series of configuration is determined, adopt promoting agent, Solid-phase organic synthesis main chain formula D-1-1-F1, and form formula D-1-1 in step b-3, cut and can the isomer of Ke Yiba peptide A of the determination configuration shown in preparation formula I-1-1 after closing ring.
According to specific embodiment of the invention scheme, in step b-1, the reagent of employing is Fmoc-MeIle7-OH��Fmoc-MeSer(Me)6-OH��Fmoc-MeThr(POH)5-OH��Fmoc-MeLeu4-OH��Fmoc-MeSer(Me)3-OH and Me2Val1-Hiv2-OH; Or
The part or all of fragment being selected from following sequence protected or do not protect is adopted to be reagent: Me2Val1-Hiv2-MeSer (Me)3-MeLeu4-MeThr (POH)5-MeSer (Me)6-MeIle7��
Those skilled in the art should know, when the reagent adopted is Fmoc-MeIle7-OH��Fmoc-MeSer(Me)6-OH��Fmoc-MeThr(POH)5-OH��Fmoc-MeLeu4-OH��Fmoc-MeSer(Me)3-OH and Me2Val1-Hiv2During-OH, be in fact on hydrazine resin successively one by one coupling to form the resin-carried main chain of described hydrazine. When the reagent adopted is protection or the part or all of fragment being selected from backbone sequence do not protected is reagent; in fact it is that the fragment meeting the resin-carried main chain peptide sequence of hydrazine is coupled on resin successively; array configuration and the number of above-mentioned fragment are unrestricted; as long as the resin-carried main chain of the hydrazine shown in described formula D-1-F1 can be formed, such as coupling Fmoc-MeIle on the de-hydrazine resin except Fmoc group7After-OH, coupling Fmoc-MeThr (P successivelyOH)5-MeSer (Me)6-OH��Fmoc-MeSer(Me)3-MeLeu4-OH and Me2Val1-Hiv2-OH; Such as coupling Fmoc-MeIle on the de-hydrazine resin except Fmoc group again7After-OH, coupling Fmoc-MeLeu successively4-MeThr (POH)5-MeSer (Me)6-OH and Me2Val1-Hiv2-MeSer (Me)3-OH��
According to specific embodiment of the invention scheme, take off except the amino protecting group P in formula D-1-F2 in step b-3NHAfterwards, the reagent of employing is the Tyr (Me) of Fmoc or Boc radical protection10��MeLeu9Or Ala8; Or
Take off except the amino protecting group P in formula D-1-F2 in step b-3NHAfterwards, the part or all of fragment being selected from following sequence adopting Fmoc or Boc radical protection is reagent: Tyr (Me)10-MeLeu9-Ala8��
Those skilled in the art should know, when the reagent adopted is the Tyr (Me) of Fmoc or Boc radical protection10��MeLeu9Or Ala8Time, in fact it is except amino protecting group P de-NHFormula D-1-1-F2 on successively one by one coupling to form formula D-1.When adopt reagent be Fmoc or Boc radical protection be selected from Tyr (Me)10-MeLeu9-Ala8When part or all of fragment in sequence is reagent, in fact it is be coupled to the fragment meeting this sequence successively to take off except amino protecting group PNHFormula D-1-1-F2 on, array configuration and the number of above-mentioned fragment are unrestricted, as long as formula D-1 can be formed.
According to specific embodiment of the invention scheme, in step b or step b-1, described activator is one or more the combination in DCC, DIC, HATU, HBTU, HCTU, HOAt, HOBt, PyBOP, BOP-Cl, phosgene, trichloromethylchloroformate and triphosgene, it is preferable that phosgene, trichloromethylchloroformate or triphosgene;
More preferably, in step b-1, phosgene, trichloromethylchloroformate or triphosgene is adopted to activate the single amino acid of Fmoc radical protection to be connected or amino acid fragment and Me independently of one another2Val1-Hiv2-OH, activation step comprises: by the single amino acid of Fmoc radical protection to be connected or amino acid fragment, or Me2Val1-Hiv2-OH photoreactive gas, trichloromethylchloroformate or triphosgene are dissolved in inert solvent, add organic bases, are obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, or Me2Val1-Hiv2-Cl, wherein, is converted into the mole number after phosgene calculates with phosgene or by trichloromethylchloroformate, triphosgene, and the mole number of phosgene is single amino acid or the amino acid fragment of described Fmoc radical protection, or Me2Val1-Hiv21.0��2.0 times of the mole number of-OH;
More preferably, in step b-3, adopt phosgene, trichloromethylchloroformate or triphosgene activate Fmoc or Boc radical protection single amino acid to be connected or amino acid fragment independently of one another, activation step comprises: independently of one another by Fmoc or Boc radical protection single amino acid or amino acid fragment photoreactive gas, trichloromethylchloroformate or triphosgene are dissolved in inert solvent, add organic bases, it is obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, wherein, with phosgene or by trichloromethylchloroformate, triphosgene is converted into the mole number after phosgene and calculates, the mole number of phosgene is 1.0��2.0 times of the mole number of Fmoc or Boc radical protection single amino acid or amino acid fragment, further preferably, in step b-3, after being obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, add HOAt immediately and make single amino acid acyl chlorides or amino acid fragment acyl chlorides change into corresponding ester, wherein, the mole number of HOAt is 0.5��1.0 times of the mole number of Fmoc or Boc radical protection single amino acid or amino acid fragment,
Wherein, in step b-1 or step b-3, described organic bases is one or more the combination in trimethylpyridine, diisopropylethylamine, pyridine, lutidine and 2-toluquinoline;
Described inert solvent is one or more the combination in tetrahydrofuran (THF), two alkane, diglyme and 1,3-propylene dichloride.
The present invention studies and finds when single amino acid is Argine Monohydrochloride (non-N-methyl amino acid), multiple side reaction can be there is in the acyl chlorides using BTC activation to produce, reduce condensation efficiency, by adding HOAt, acyl chlorides is converted into corresponding-OAt ester, can be significantly improved it and the solid phase condensation efficiency of space steric hindrance is big, nucleophilie nucleus ability is poor amino acid (with the amino acid of the substituting groups such as methyl on such as nitrogen-atoms), condensation product rate can be improved within the relatively short time.
According to specific embodiment of the invention scheme, in step a, described hydrazine resin is phenylhydrazine resin;More preferably, the substitution degree of described hydrazine resin is 0.1��1.0mmol/g.
According to specific embodiment of the invention scheme, in step c, described oxygenant is that neutralized verdigris, N-bromine are for one or more the combination in succimide and oxygen;
In steps d, described condensing agent is EDC HCl, DCC, DIC, DMTMM+BF4 -��DMTMM+Cl-, one or more combination in HATU, HBTU, HCTU, HOAt, HOBt, BOP, BOP-Cl, PyBOP, PyAOP, DPPA, FDP and FDPP.
In the present invention, hydrazine key is oxidized to diazene key by acting as of the oxygenant in step c, subsequently under the effect of cutting agent, polypeptide chain in the cyclisation precursor such as, making step b gained hydrazine resin-carried under the effect of water and resin isolation, obtain polypeptide chain cyclisation precursor, those skilled in the art should know this step and can first carry out being oxidized cutting resin again, it is also possible in the cutting carrying out resin of oxidation simultaneously.
On the other hand, the present invention provides the isomer of a kind of Ke Yiba peptide A, and the isomer of this Ke Yiba peptide A has the structure shown in formula I-1-1:
The present invention studies the activity finding that the isomer of this Ke Yiba peptide A has good anti-cell hyper-proliferative.
The structure of the isomer of the structure of the Ke Yiba peptide A shown in formula I-1-2 disclosed in contrast prior art and the Ke Yiba peptide A shown in formula I-1-1 can be found out, the configuration of the isomer of this Ke Yiba peptide A 2 carboxylic acids and 11 amino acids there occurs change, and brings unexpected antiproliferation due to the change of configuration.
Another further aspect, the present invention provides the application of the isomer of the Ke Yiba peptide A shown in formula I-1-1 in the medicine for the preparation of overmedication proliferative disease;
Preferably, wherein said excess proliferative disease comprises one or more in colon and rectum carcinoma, brain tumor (being preferably glioblastoma), lung cancer (being preferably nonsmall-cell lung cancer), epidermis squama cancer, bladder cancer, carcinoma of the pancreas, mammary cancer, ovarian cancer, cervical cancer, carcinoma of endometrium, colorectal cancer, renal cell carcinoma, adenocarcinoma of esophagus, esophageal squamous cell carcinoma, non-Hodgkin lymphoma, liver cancer, skin carcinoma, thyroid carcinoma, head and neck cancer, prostate cancer, neurospongioma and nasopharyngeal carcinoma; More preferably, described excess proliferative disease is mammary cancer, nonsmall-cell lung cancer.
In summary, the present invention provides the method for the Ke Yiba peptide A analogue designed by a kind of synthesis quick, efficient, and utilizes the method to obtain the isomer of a kind of Ke Yiba peptide A with good anti-cell hyper-proliferative.
Abbreviation in the present invention has as given a definition:
BTC: triphosgene; EDC HCl:1-ethyl-(3-dimethylaminopropyl) phosphinylidyne diimine hydrochloride; DCC:N, N'-dicyclohexyl carbimide; DIC:N, N'-DIC; DMTMM+BF4 -: 4-(4,6-dimethoxy-triazine-2-base)-4-methylmorpholine a tetrafluoro borate; DMTMM+Cl-: 4-(4,6-dimethoxy-triazine-2-base)-4-methylmorpholine villaumite; HATU:O-(7-nitrogen benzotriazole)-N, N, N', N'-tetramethyl-urea phosphofluoric acid ester; HBTU: benzotriazole-N, N, N', N'-tetramethyl-urea hexafluorophosphate; HCTU:6-chlorine benzotriazole-1,1,3,3-tetramethyl-urea phosphofluoric acid ester; HOAt:1-hydroxyl-7-azo benzotriazole; HOBt:1-hydroxybenzotriazole; BOP: benzotriazole-1-base oxygen base three (dimethylamino) phosphorus hexafluorophosphate;BOP-Cl: two (2-oxo-3-dislikes oxazolidinyl) secondary phosphoryl chloride; PyBOP: phosphofluoric acid benzotriazole-1-base-oxygen base tripyrrole alkyl phosphorus; PyAOP:(3H-1,2,3-triazolo [4,5-b] pyridine-3-oxygen base) three-1-pyrrolidyl hexafluorophosphates; DPPA: diphenyl phosphate azide; FDP:pentafluorophenyldiphenylphosphate; FDPP: pentafluorophenyl group diphenyl phosphoester; DMAP:4-Dimethylamino pyridine; TFA: trifluoroacetic acid; TIS: tri isopropyl silane; Fmoc:9-fluorenylmethyloxycarbonyl; Boc: tertiary butoxy carbonyl; Cbz (Z): carbobenzoxy-(Cbz); Trt: trityl; Allyl: allyloxycarbonyl;
Fmoc-MeIle7-OH:Fmoc-(L)-MeIle7-OH:
Fmoc-MeSer(Me)6-OH:Fmoc-(L)-MeSer(Me)6-OH:
Fmoc-MeThr(POH)5-OH:Fmoc-(L)-MeThr(POH)5-OH:
Fmoc-MeLeu4-OH:Fmoc-(L)-MeLeu4-OH:
Fmoc-MeSer(Me)3-OH:Fmoc-(L)-MeSer(Me)3-OH:
Me2Val1��Hiv2-OH:Me2Val1-Hiv2-Cl:
(L)-Me2Val1��(D)-Hiv2-OH:
(L)-Me2Val1��(L)-Hiv2-OH:
Tyr(Me)10:MeLeu9:Ala8:
Tyr(Me)10-MeLeu9-Ala8:
Fmoc-(D)-MeAla11-OH:Fmoc-(L)-MeAla11-OH:
Boc-(L)-Tyr(Me)10-OH:Fmoc-(L)-MeLeu9-OH:
Fmoc-(L)-Ala8-OH:
Me2Val1-Hiv2-MeSer (Me)3-MeLeu4-MeThr (POH)5-MeSer (Me)6-MeIle7:
Accompanying drawing explanation
Fig. 1 is the basic flow sheet of the isomer of the Ke Yiba peptide A shown in embodiment 1 preparation formula I-1-1;
Fig. 2 is the basic line figure that embodiment 1 prepares the resin-carried cyclisation precursor formula 4 of phenylhydrazine;
Fig. 3 is embodiment 1 by the basic line figure of the isomer of the Ke Yiba peptide A shown in the resin-carried cyclisation precursor formula 4 preparation formula I-1-1 of phenylhydrazine;
Fig. 4 is the cyclisation precursor 6 of embodiment 1 preparation1HNMR schemes;
Fig. 5 is the cyclisation precursor 6 of embodiment 1 preparation13CNMR schemes;
Fig. 6 is the HRMS figure of cyclisation precursor 6 prepared by embodiment 1;
Fig. 7 is the isomer of the Ke Yiba peptide A shown in formula I-1-1 prepared by embodiment 11HNMR schemes;
Fig. 8 is the isomer of the Ke Yiba peptide A shown in formula I-1-1 prepared by embodiment 113CNMR schemes;
Fig. 9 is the HRMS figure of the isomer of the Ke Yiba peptide A shown in formula I-1-1.
Embodiment
In order to the technology feature to the present invention, object and useful effect have understanding clearly, now in conjunction with specific embodiments and accompanying drawing the technical scheme of the present invention is carried out following detailed explanation, it should be appreciated that these examples are only not used in for illustration of the present invention and limit the scope of the invention.
Embodiment 1
The present embodiment provides isomer and its preparation method of the Ke Yiba peptide A shown in formula I-1-1.
Fig. 1 show generally the preparation method of the isomer of the Ke Yiba peptide A shown in formula I-1-1: adopts and provides the Fmoc phenylhydrazine resin of protection; De-except, after the Fmoc protecting group on phenylhydrazine resin, synthesizing the main chain that phenylhydrazine is resin-carried; De-except (L)-MeThr5Tert. butyl protection group on hydroxyl; First amino acid (the D)-MeAla being connected on side chain by esterification11; Synthesis side chain; The resin-carried cyclisation precursor of oxygen cutting hydrazine prepares the polypeptide chain cyclisation precursor that one of carbon tip is carboxyl; Last cyclisation obtains the isomer of the Ke Yiba peptide A shown in formula I-1-1.
Specifically, the cyclisation precursor 4 that the synthesis phenylhydrazine of the route shown in Fig. 2 is resin-carried is first pressed.
(1) preparation of the main chain 1 that phenylhydrazine is resin-carried:
The solid phase reactor of 10mL adds the phenylhydrazine resin (400mg, 0.66mmol/g, 0.26mmol are purchased from Novabiochem) with Fmoc protection and 3mLCH2Cl2, by swelling for phenylhydrazine resin 30min, extract CH2Cl2Take off except the Fmoc protecting group on phenylhydrazine resin with 3mL20% piperidines/DMF solution, after 10min, extract reaction solution, again take off except the Fmoc protecting group on phenylhydrazine resin with 3mL20% piperidines/DMF solution, after 10min, extract reaction solution, adopt DMF (4 �� 3mL) washing resin, then with anhydrous THF (2 �� 3mL) washing resin, for subsequent use;Simultaneously by 97mgFmoc-(L)-MeIle7-OH (0.26mmol) and 32mg triphosgene (BTC, 0.11mmol) it is dissolved in the anhydrous THF of 2mL, the �� L trimethylpyridine (collidine that adds 420 is slowly dripped in this solution, 3.17mmol), reaction produces a large amount of white precipitate immediately, reacts 3min, this reaction solution is transferred to above-mentioned gained and takes off in the phenylhydrazine resin except Fmoc after adding, add 870 �� LDIEA (5.28mmol) again, N2Bubbling mixes even, after condensation reaction 30min, extracts reaction solution, with DMF (4 �� 3mL) washing resin, then with 3mL dry DMF washing resin; The DMF solution that 125 �� L diacetyl oxide (1.32mmol) and 2mL contain 200 �� LDIEA (1.20mmol) is added in resin, reaction 10min, extract reaction solution, with DMF (4 �� 3mL) washing resin, obtained Fmoc-(L)-MeIle7-phenylhydrazine resin, its substitution value is 0.30mmol/g (400mg, 0.12mmol, 1eq);
As shown in Figure 2, repeat above-mentioned deprotection and condensation step, adopt the mode of coupling one by one, continue coupling Fmoc-(L)-MeSer (Me) successively6-OH��Fmoc-(L)-MeThr(t-Bu)5-OH��Fmoc-(L)-MeLeu4-OH��Fmoc-(L)-MeSer(Me)3-OH and (L)-Me2Val1��(D)-Hiv2-OH, until completing the resin-carried main chain of phenylhydrazine 1 ((L)-Me2Val1-(D)-Hiv2-(L)-MeSer (Me)3-(L)-MeLeu4-(L)-MeThr (t-Bu)5-(L)-MeSer (Me)6-(L)-MeIle7-phenylhydrazine resin) synthesis, wherein the step of deprotection and condensation is substantially identical, such as, at obtained Fmoc-(L)-MeIle7After-phenylhydrazine resin, de-except Fmoc-(L)-MeIle with 3mL20% piperidines/DMF solution7Fmoc protecting group on-resin; after 10min; extract reaction solution; again take off except the Fmoc protecting group on phenylhydrazine resin with 3mL20% piperidines/DMF solution; after 10min, extract reaction solution, with DMF (4 �� 3mL) washing resin; again with anhydrous THF (2 �� 3mL) washing resin, for subsequent use. Activate each Fmoc-to be connected (L)-MeSer (Me) simultaneously6-OH, is specifically operating as: by Fmoc-to be connected (the L)-MeSer (Me) of 4.0eq6The triphosgene (BTC) of-OH and 1.64eq is dissolved in the anhydrous THF of 2mL; slowly drip in this solution and add 12eq trimethylpyridine (collidine); reaction produces a large amount of white precipitate immediately; add reaction 3min, this reaction solution is transferred to de-(L)-NHMeIle except Fmoc protecting group7In-phenylhydrazine resin, then add 20eqDIEA, N2Bubbling mixes even, condensation reaction 0.5��1h, until the detection of tetrachlorobenzoquinone reagent to after reacting completely, extracting reaction solution, with DMF (4 �� 3mL) washing resin;
(2) preparation of the cyclisation precursor 4 that phenylhydrazine is resin-carried:
Main chain 1 resin-carried for phenylhydrazine obtained above is used CH2Cl2(2 �� 3ml) washs, and extracts CH2Cl2, add 3.0mLTFA/TIS/H2O (v/v/v=95:2.5:2.5) mixing solutions is de-except tert. butyl protection group t-Bu, extracts solvent, again add TFA/TIS/H after 2min2O (volume ratio is 95:2.5:2.5) mixing solutions (3.0mL) reacts 5min, uses CH2Cl2(2 �� 3ml) and DMF (4 �� 3mL) washing resin, then with 3mL dry DMF washing resin, obtain the de-main chain 2 resin-carried except the phenylhydrazine of t-Bu, for subsequent use. Simultaneously by Fmoc-(the D)-MeAla of 4.0eq11-OH and 2eqDCC is dissolved in 2mLCH2Cl2/ DMF (v/v=1:1), stirring reaction 2h at 0 DEG C, centrifugation white precipitate, transfers to reaction solution in the de-main chain 2 resin-carried except the phenylhydrazine of t-Bu, adds the DMAP of catalytic amount, N2Bubbling mixes even, and condensation reaction 2.5h, extracts reaction solution, with DMF (4 �� 3mL) washing resin, obtains product 3;
Fmoc protecting group in 3 is removed with 3mL20% piperidines/DMF solution is de-, after 10min, with DMF (4 �� 3mL) washing resin, then with dry DMF (2 �� 3mL) washing resin, for subsequent use. Simultaneously by 4.0eqBoc-(L)-Tyr (Me)10-OH and 1.64eq triphosgene (BTC) are dissolved in the anhydrous THF of 1mL, slowly drip in this solution and add 12eq trimethylpyridine (collidine), reaction produces a large amount of white precipitate immediately, 4.0eqHOAt and 1.5mLDIEA/DMF (15% is added in reaction system, v/v), after reaction 3min, this reaction solution is transferred in the de-product 3 except Fmoc, N2Bubbling mixes even, condensation reaction 0.5��1h, until the detection of tetrachlorobenzoquinone reagent to after reacting completely, extracting reaction solution, with DMF (4 �� 3mL) washing resin;
Use CH2Cl2(2 �� 3ml) washing resin, extracts CH2Cl2, add 3.0mLTFA/TIS/H2O (v/v/v=95:2.5:2.5) mixing solutions, de-except Boc protecting group, extract solvent after 2min, again add 3.0mLTFA/TIS/H2O (v/v/v=95:2.5:2.5) mixing solutions, reaction 5min, uses CH2Cl2(2 �� 3mL) and DMF (4 �� 3mL) washing resin, then with the anhydrous THF washing resin of 3mL, for subsequent use. Simultaneously by 4.0eqFmoc-(L)-MeLeu9-OH and 1.64eq triphosgene (BTC) are dissolved in the anhydrous THF of 2mL, slowly drip in this solution and add 12eq trimethylpyridine (collidine), reaction produces a large amount of white precipitate immediately, add reaction 3min, this reaction solution is transferred in the de-phenylhydrazine resin except Boc obtained above, add 20eqDIEA, N again2Bubbling mixes even, condensation reaction 0.5��1h, until the detection of tetrachlorobenzoquinone reagent to after reacting completely, extracting reaction solution, with DMF (4 �� 3mL) washing resin;
3mL20% piperidines/DMF solution is adopted to take off except the Fmoc protecting group in above-mentioned gained phenylhydrazine resin; after 10min; extract reaction solution; again take off except the Fmoc protecting group on phenylhydrazine resin with 3mL20% piperidines/DMF solution; after 10min, extract reaction solution, with DMF (4 �� 3mL) washing resin; again with dry DMF (2 �� 3mL) washing resin, for subsequent use. Simultaneously by 4.0eqFmoc-(L)-Ala8-OH and 1.64eq triphosgene (BTC) are dissolved in the anhydrous THF of 1mL, slowly drip in this solution and add 12eq trimethylpyridine (collidine), reaction produces a large amount of white precipitate immediately, 4.0eqHOAt and 1.5mLDIEA/DMF (15% is added in reaction system, v/v), after reaction 3min, this reaction solution is transferred in the de-phenylhydrazine resin except Fmoc, N2Bubbling mixes even, condensation reaction 0.5��1h, until the detection of tetrachlorobenzoquinone reagent to after reacting completely, extracting reaction solution, with DMF (4 �� 3mL) washing resin, complete the synthesis of the resin-carried cyclisation precursor 4 of phenylhydrazine.
(3) synthesis of the isomer of the Ke Yiba peptide A shown in formula I-1-1
By the isomer of the Ke Yiba peptide A shown in the route synthesis type I-1-1 shown in Fig. 3:
Take out the cyclisation precursor 4 that in step (2), gained phenylhydrazine is resin-carried, add 4mLCH2Cl2��10mgCu(OAc)2, 200 �� L pyridines and 200 �� L water, under room temperature after oscillatory reaction 24h, by resin filtering, and use 20mLCH2Cl2Washing resin. Organic phase being concentrated, residue preparation HPLC purifies (10% acetonitrile H2O (TFA containing 1%) Gradient elution 5min, the acetonitrile H of 10% to 100%2O (TFA containing 1%) gradient elution 25min, retention time TR=27min), collect product and solvent under reduced pressure is steamed dry, obtain product 5 (27mg, comes in first amino acid whose substitution value, and receipts rate is 20%).HRMS (ESI) m/z: Theoretical Calculation C80H123N10O19[M+H]+Accurate molecular weight be 1527.8960, actual measurement is 1527.8963.
15.3mg product 5 (0.01mmol) adds 1.5mLEt2NH and 1.5mLCH3CN, stirred at ambient temperature reaction 15min, evaporated under reduced pressure solvent, residue preparation HPLC purifies (10% acetonitrile H2O (TFA containing 1%) Gradient elution 5min, the acetonitrile H of 10% to 100%2O (TFA containing 1%) gradient elution 25min, retention time TR=20min), collect product, obtain white solid 6 after lyophilize for subsequent use, its1HNMR��13CNMR and HRMS is respectively as shown in Fig. 4, Fig. 5 and Fig. 6.
19.2mgEDC HCl (0.10mmol), 13.6mgHOAt (0.10mmol), 70 �� LDIEA (0.40mmol) are dissolved in 30mLCH2Cl2In, under ice bath, it is cooled to 0 DEG C. Above-mentioned white solid 6 is dissolved in CH2Cl2(2mL) in, slowly drop in the reaction solution of EDC HCl/HOAt/DIEA at 0 DEG C, remove ice bath after reaction 1h, reaction mixture is continued at room temperature stirring reaction 17h, removes solvent under reduced pressure, purify (10% acetonitrile H with preparation HPLC2O (TFA containing 1%) Gradient elution 5min, the acetonitrile H of 10% to 100%2O (TFA containing 1%) gradient elution 25min, retention time TR=25min), collect product, lyophilize, obtains the isomer of the Ke Yiba peptide A shown in 7.2mg formula I-1-1, and receipts rate is 60%, comes in first amino acid whose substitution value, and total recovery is 12%, its1HNMR��13CNMR and HRMS is respectively as shown in Fig. 7, Fig. 8 and Fig. 9.
[��]D 23�C58.4��(c0.10,CHCl3);
IR:3376,2957,1734,1644,1512,1468,1404,1248,1097,757cm-1;
1HNMR(600MHz,CDCl3) �� 0.83 (d, J=6.6Hz, 3H), 0.88-0.99 (m, 18H), 1.00 (d, J=4.2Hz, 3H), 1.05-1.08 (m, 9H), 1.11 (m, 1H), 1.11 (d, J=7.2Hz, 3H), 1.28 (d, J=6.6Hz, 3H), 1.31 (m, 1H), 1.36 (m, 2H), 1.50 (m, 2H), 1.60 (m, 1H), 1.68 (m, 1H), 2.02 (m, 1H), 2.05 (m, 1H), 2.21 (m, 1H), 2.34 (brs, 9H), 2.75 (s, 3H), 2.84 (m, 2H), 2.86 (s, 3H), 2.89 (m, 3H), 2.99 (m, 1H), 3.04 (s, 3H), 3.12 (s, 3H), 3.15 (s, 3H), 3.30 (s, 3H), 3.35 (s, 3H), 3.53 (m, 1H), 3.61 (m, 1H), 3.65 (dd, J=11.4, 4.2Hz, 1H), 3.75 (m, 1H), 3.77 (s, 3H), 3.83 (m, 1H), 3.90 (m, 1H), 4.73 (m, 1H), 5.00 (d, J=5.5Hz, 1H), 5.11 (m, 1H), 5.32-5.38 (m, 2H), 5.50 (brs, 1H), 5.69 (m, 1H), 6.00 (brs, 1H), 6.35 (brs, 1H), 6.63 (brs, 1H), 6.68 (brs, 1H), 6.77 (d, J=9.0Hz, 2H), 7.09 (d, J=8.4Hz, 2H), ,
13CNMR(100MHz,CDCl3) �� 11.6,12.9,15.7,17.9,18.0,18.4,18.6,19.5,19.6,21.2,21.4,23.2,23.3,24.2,24.9,25.3,27.6,28.9,29.7,29.9,30.1,30.2,31.3,32.0,36.2,37.9,38.9,39.4,41.3,47.0,49.9,51.0,51.1,52.5,52.9,55.3,58.6,58.8,63.5,64.7,68.6,69.3,73.8,74.8,113.7,128.4,130.4,158.6,167.2,168.7,169.7,170.0,170.5,171.2,171.3,171.5,172.4ppm;
HRMS (ESI-TOF) m/z: Theoretical Calculation C65H111N10O16[M+H]+Accurate molecular weight is 1287.8174, and actual measurement is 1287.8176.
Embodiment 2
The present embodiment provides the Ke Yiba peptide A shown in formula I-1-2 and its preparation method.
Method and the embodiment 1 preparation formula I-1-2 of preparation formula I-1-2 are substantially similar, only with (L)-Me2Val1-(L)-Hiv2(L)-Me in-OH alternative embodiment 12Val1-(D)-Hiv2-OH, and with Fmoc-(L)-MeAla11Fmoc-(D)-MeAla in-OH alternative embodiment 111-OH, all the other steps are all identical, come in first amino acid whose substitution value in experimentation, and total recovery is 8%.
[��]D 23�C128.9��(c0.7,CHCl3);
IR:2960,2923,1732,1645,1508,1459,1405,1248,1093,1012,756cm-1;
1HNMR(CDCl3, 400MHz): �� 0.62-1.15 (m, 36H), 1.15-1.38 (m, 7H), 1.40-1.51 (m, 2H), 1.60-1.76 (m, 2H), 2.00-2.10 (m, 2H), 2.12-2.20 (m, 1H), 2.30-2.35 (m, 6H), 2.65-3.40 (m, 28H), 2.45 (brs, 1H), 3.45-3.60 (m, 2H), 3.60-3.70 (m, 1H), 3.70-3.86 (m, 6H), 4.52-5.24 (m, 5H), 5.34-5.60 (m, 2H), 5.60-5.75 (m, 2H), 5.77-5.80 (m, 0.7H), 5.89 (d, 0.3H, J=4.2Hz), 6.57 (brs, 1H), 6.80-6.85 (m, 2H), 6.97 (brs, 1H), 7.10-7.20 (m, 2H) ppm,
13CNMR(CDCl3, 100MHz): �� 11.6, 13.5, 14.1, 15.9, 18.0, 18.1, 18.5, 18.6, 18.7, 19.1, 19.3, 19.6, 21.2, 21.5, 22.7, 23.5, 23.6, 24.2, 24.8, 25.4, 27.7, 27.8, 28.5, 28.8, 29.7, 29.9, 30.1, 30.8, 30.9, 31.0, 31.9, 32.0, 33.4, 36.4, 38.4, 40.0, 40.1, 41.3, 46.5, 50.1, 50.9, 52.1, 52.6, 55.0, 55.1, 55.2, 55.3, 58.7, 58.8, 58.9, 64.1, 64.7, 68.7, 68.9, 69.0, 74.0, 74.1, 74.5, 74.6, 113.9, 114.2, 114.4, 130.2, 130.4, 158.8, 169.6, 169.7, 170.0, 170.1, 170.3, 170.4, 171.6, 171.8ppm,
HRMS (ESI-TOF) m/z: Theoretical Calculation C65H111N10O16[M+H]+Accurate molecular weight be 1287.8174, actual measurement is 1287.8170.
The anti-cell transition proliferation activity of the isomer of the Ke Yiba peptide A shown in embodiment 3 formula I-1-1
1) medicine and preparation of reagents: 1L water is added in one bag of RPMI1640 substratum, add 2g sodium bicarbonate, 100,000 unit penicillin and 100mg Streptomycin sulphate, adjust ph to 7.4, degerming with 0.22 ��m of degerming membrane filtration. 95mL substratum adds deactivation new-born calf serum 5mL and is complete culture solution. Trypsinase D-hanks damping fluid is made into 0.25% solution, and after filtration sterilization, 4 DEG C save backup.
2) testing sample solution preparation: the isomer accurately taking in 1.29mg (1.0 ��m of ol) embodiment 1 coibamideA shown in formula I-1-1 prepared, it is added in the 0.5mL centrifuge tube of sterilizing, add 100 �� LDMSO, it is made into 10 ��Ms of stostes ,-40 DEG C of freezen protective. Get after melting before use and it is diluted to corresponding concentration with complete culture solution in right amount.
3) cell cultures and going down to posterity: the equal adherent culture of cell in containing in 10mL complete culture solution Tissue Culture Flask, in 37 DEG C, 5%CO2, cultivate under saturated humidity. Cell cover with bottle at the bottom of after wash twice with sterilizing D-hanks liquid, the trypsin digestion and cell that adds 0.25% 2 minutes, outwells trypsinase, after gently shaking cell and coming off completely, after adding complete culture solution 30ml, blow loose cell with transfer pipet, it is sub-packed in 3 new Tissue Culture Flasks, continues to cultivate.
4) drug treating: get one bottle, the cell just growing up to intact monolayer, collecting cell after tryptic digestion, even with transfer pipet piping and druming, get two cell suspension trypan blue (TrypanBlue) dyeing, in counted under microscope viable cell number (dead cell number must not more than 5%), adjust cell number to 1 �� 105 cells/ml with complete culture solution. In 96 porocyte culture plates, every hole adds 100 �� L cell suspensions, and culture plate is placed in CO2Incubator is cultivated 12 hours, in every hole, the complete culture solution of 100 �� L containing different concns testing sample is added after taking out culture plate, medicine final concentration is made to be respectively 1000,100.0,10.0,1.0,0.1 and 0.01nM, each concentration establishes 4 parallel holes, separately establishes 4 porocytes to add and does not do normal control hole containing medicine complete culture solution.After adding medicine, culture plate vibrates mixed even on microwell plate vibrator, is placed in CO2Incubator resume cultivates 72 hours. Taking out culture plate, every hole adds the MTT liquid 20 �� L of 5mg/mL, and vibration is mixed even, continues to cultivate 4 hours. Abandoning nutrient solution in every hole, the DMSO of the �� L that adds 100 dissolves formazan crystallization, and vibration 10min makes to dissolve completely.
5) determination of cytotoxic activity: 503nhibiting concentration measure the photoabsorption of each hole by microplate reader, measure wavelength 490nm. The inhibiting rate of medicine on cell proliferation is calculated: inhibiting rate (%)=[1-OD (dosing group)/OD (negative control group)] �� 100% according to each hole OD value. Straight-line regression done by the inhibiting rate MicrocalOrigin software that logarithm according to drug level is corresponding, obtains straight-line equation, and the drug level calculating inhibiting rate corresponding when 50% is testing sample to the 503nhibiting concentration (IC of tumour cell50), gained data are as shown in table 1:
The IC of the isomer of the Ke Yiba peptide A shown in table 1 formula I-1-150Value
The Ke Yiba peptide A shown in formula I-1-1 synthesized by the present invention has good anticancer cytoactive, especially anti-breast cancer cell and non-small cell cancer cells as can be seen from Table 1.
Claims (10)
1. the synthetic method of Yi Zhong Ke Yiba peptide A analogue, this Ke Yiba peptide A analogue has the structural formula shown in formula I:
In formula I, R1��R2��R3��R5��R7��R8��R9��R11��R13��R14It is hydrogen, methyl or ethyl independently of one another;
R4For methoxy-benzyl, ethoxy benzyl, methylamino benzyl or ethylamino-benzyl;
R6��R10It is normal-butyl, isobutyl-or sec-butyl independently of one another;
R12For methoxymethyl, ethoxyl methyl, methoxy ethyl or ethoxyethyl group;
G be hydrogen, methyl,
Or amino protecting group; This amino protecting group is Boc, Fmoc, Cbz, Trt or Allyl;
R15��R21��R22It is n-propyl, sec.-propyl, normal-butyl, isobutyl-or sec-butyl independently of one another;
R16��R17��R19��R20��R23��R24It is hydrogen, methyl, ethyl or amino protecting group independently of one another;
R18For methoxymethyl, ethoxyl methyl, methoxy ethyl or ethoxyethyl group;
Y is hydroxyl, methoxyl group, amino, methylamino-or two methylamino;
X is O, N or N (CH3);
Preferably, formula I has the structure shown in formula I-1:
More preferably, formula I-1 has the structure shown in formula I-1-1 or formula I-1-2:
Described method comprises the steps:
A., hydrazine resin is provided;
B. activator is adopted, the cyclisation precursor that in Solid-phase organic synthesis formula A��formula F, any one hydrazine is resin-carried:
The P of formula A��formula F in any oneNHRepresent amino protecting group; This amino protecting group is Boc, Fmoc, Cbz, Trt or Allyl, it is preferable that Fmoc;
C. carrying out cleavage reaction under the existence of oxygenant, polypeptide chain in the cyclisation precursor making gained hydrazine in step b resin-carried and resin isolation, obtain polypeptide chain cyclisation precursor;
Preferably, the one of carbon tip of described polypeptide chain cyclisation precursor is carboxyl;
D. de-except the protecting group P on nitrogen end on gained polypeptide chain cyclisation precursor in step cNHAfter, adopt condensing agent to carry out ring-closure reaction, obtain compound shown in formula I.
2. method according to claim 1, wherein, in stepb, adopts activator, and Solid-phase organic synthesis is such as formula the resin-carried cyclisation precursor of the hydrazine shown in D;
Preferably, Solid-phase organic synthesis is such as formula the resin-carried cyclisation precursor of the hydrazine shown in D-1:
More preferably, the cyclisation precursor that hydrazine shown in Solid-phase organic synthesis formula D-1-1 or formula D-1-2 is resin-carried:
3. method according to claim 2, wherein, the resin-carried cyclisation precursor of the hydrazine shown in formula D-1 prepares as follows:
B-1. activator is adopted, the main chain that the hydrazine shown in Solid-phase organic synthesis formula D-1-F1 is resin-carried:
Preferably, the main chain that hydrazine shown in Solid-phase organic synthesis formula D-1-1-F1 or D-1-2-F1 is resin-carried:
P described in formula D-1-F1, formula D-1-1-F1 or formula D-1-2-F1OHFor hydroxyl protecting group, it is preferable that the tertiary butyl, trityl, allyl group or benzyl;
B-2. adopt and go protecting group agent to take off except the hydroxyl protecting group P on the resin-carried main chain of step b-1 gained hydrazineOH;
B-3. the hydroxyl exposed in step b-2 carries out esterification with the structure shown in obtained formula D-1-F2:
De-except the amino protecting group P in formula D-1-F2NHAfter, continue coupling and form the resin-carried cyclisation precursor of the hydrazine shown in formula D-1;
Preferably, the hydroxyl exposed in step b-2 carries out esterification with the structure shown in obtained formula D-1-1-F2 or formula D-1-2-F2:
The de-amino protecting group P except formula D-1-1-F2 or formula D-1-2-F2NHAfter, continue coupling to form the resin-carried cyclisation precursor of the hydrazine shown in formula D-1-1 or formula D-1-2.
4. method according to claim 3, wherein, in step b-1, the reagent of employing is Fmoc-MeIle7-OH��Fmoc-MeSer(Me)6-OH��Fmoc-MeThr(POH)5-OH��Fmoc-MeLeu4-OH��Fmoc-MeSer(Me)3-OH and Me2Val1-Hiv2-OH; Or
The part or all of fragment being selected from following sequence protected or do not protect is adopted to be reagent: Me2Val1-Hiv2-MeSer (Me)3-MeLeu4-MeThr (POH)5-MeSer (Me)6-MeIle7��
5. method according to claim 3, wherein, takes off except the amino protecting group P in formula D-1-F2 in step b-3NHAfterwards, the reagent of employing is the Tyr (Me) of Fmoc or Boc radical protection10��MeLeu9Or Ala8; Or
Take off except the amino protecting group P in formula D-1-F2 in step b-3NHAfterwards, the part or all of fragment being selected from following sequence adopting Fmoc or Boc radical protection is reagent: Tyr (Me)10-MeLeu9-Ala8��
6. method according to any one of Claims 1 to 5, wherein, in step b or step b-1, described activator is one or more the combination in DCC, DIC, HATU, HBTU, HCTU, HOAt, HOBt, PyBOP, BOP-Cl, phosgene, trichloromethylchloroformate and triphosgene, it is preferable that phosgene, trichloromethylchloroformate or triphosgene;
More preferably, in step b-1, phosgene, trichloromethylchloroformate or triphosgene is adopted to activate the single amino acid of Fmoc radical protection to be connected or amino acid fragment and Me independently of one another2Val1-Hiv2-OH, activation step comprises: by the single amino acid of Fmoc radical protection to be connected or amino acid fragment, or Me2Val1-Hiv2-OH photoreactive gas, trichloromethylchloroformate or triphosgene are dissolved in inert solvent, add organic bases, are obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, or Me2Val1-Hiv2-Cl, wherein, is converted into the mole number after phosgene calculates with phosgene or by trichloromethylchloroformate, triphosgene, and the mole number of phosgene is single amino acid or the amino acid fragment of described Fmoc radical protection, or Me2Val1-Hiv21.0��2.0 times of the mole number of-OH;
More preferably, in step b-3, adopt phosgene, trichloromethylchloroformate or triphosgene activate Fmoc or Boc radical protection single amino acid to be connected or amino acid fragment independently of one another, activation step comprises: independently of one another by Fmoc or Boc radical protection single amino acid or amino acid fragment photoreactive gas, trichloromethylchloroformate or triphosgene are dissolved in inert solvent, add organic bases, it is obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, wherein, with phosgene or by trichloromethylchloroformate, triphosgene is converted into the mole number after phosgene and calculates, the mole number of phosgene is 1.0��2.0 times of the mole number of Fmoc or Boc radical protection single amino acid or amino acid fragment, further preferably, in step b-3, after being obtained by reacting corresponding single amino acid acyl chlorides or amino acid fragment acyl chlorides, add HOAt immediately and make single amino acid acyl chlorides or amino acid fragment acyl chlorides change into corresponding ester,Wherein, the mole number of HOAt is 0.5��1.0 times of the mole number of Fmoc or Boc radical protection single amino acid or amino acid fragment;
Wherein, in step b-1 or step b-3, described organic bases is one or more the combination in trimethylpyridine, diisopropylethylamine, pyridine, lutidine and 2-toluquinoline;
Described inert solvent is one or more the combination in tetrahydrofuran (THF), two alkane, diglyme and 1,3-propylene dichloride.
7. method according to any one of Claims 1 to 5, wherein, in step a, described hydrazine resin is phenylhydrazine resin; More preferably, the substitution degree of described phenylhydrazine resin is 0.1��1.0mmol/g.
8. method according to any one of Claims 1 to 5, wherein, in step c, described oxygenant is that neutralized verdigris, N-bromine are for one or more the combination in succimide and oxygen;
In steps d, described condensing agent is EDC HCl, DCC, DIC, DMTMM+BF4 -��DMTMM+Cl-, one or more combination in HATU, HBTU, HCTU, HOAt, HOBt, BOP, BOP-Cl, PyBOP, PyAOP, DPPA, FDP and FDPP.
9. the isomer of Ke Yiba peptide A of method synthesis according to any one of claim 1��8, the isomer of this Ke Yiba peptide A has the structure shown in formula I-1-1:
10. the application of the isomer of Ke Yiba peptide A according to claim 9 in the medicine for the preparation of overmedication proliferative disease;
Preferably, wherein said excess proliferative disease comprises one or more in colon and rectum carcinoma, brain tumor, lung cancer, epidermis squama cancer, bladder cancer, carcinoma of the pancreas, mammary cancer, ovarian cancer, cervical cancer, carcinoma of endometrium, colorectal cancer, renal cell carcinoma, adenocarcinoma of esophagus, esophageal squamous cell carcinoma, non-Hodgkin lymphoma, liver cancer, skin carcinoma, thyroid carcinoma, head and neck cancer, prostate cancer, neurospongioma and nasopharyngeal carcinoma; More preferably, described excess proliferative disease is mammary cancer, nonsmall-cell lung cancer.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106749545A (en) * | 2016-12-13 | 2017-05-31 | 深圳先进技术研究院 | The preparation method of GYMNOPEPTIDE A and GYMNOPEPTIDE B |
| CN109912697A (en) * | 2017-12-13 | 2019-06-21 | 深圳先进技术研究院 | Ke Yiba peptide A derivative and its preparation method and application |
| CN109908363A (en) * | 2017-12-12 | 2019-06-21 | 深圳先进技术研究院 | It is a kind of to target seamless release drug conjugate and the preparation method and application thereof |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106749545A (en) * | 2016-12-13 | 2017-05-31 | 深圳先进技术研究院 | The preparation method of GYMNOPEPTIDE A and GYMNOPEPTIDE B |
| CN109908363A (en) * | 2017-12-12 | 2019-06-21 | 深圳先进技术研究院 | It is a kind of to target seamless release drug conjugate and the preparation method and application thereof |
| CN109908363B (en) * | 2017-12-12 | 2022-03-08 | 深圳先进技术研究院 | Targeted traceless release drug conjugate and preparation method and application thereof |
| CN109912697A (en) * | 2017-12-13 | 2019-06-21 | 深圳先进技术研究院 | Ke Yiba peptide A derivative and its preparation method and application |
| CN109912697B (en) * | 2017-12-13 | 2020-12-18 | 深圳先进技术研究院 | Coibatide A derivative and preparation method and application thereof |
| CN112159462A (en) * | 2017-12-13 | 2021-01-01 | 深圳先进技术研究院 | Coibatide A derivative and application thereof |
| CN112159462B (en) * | 2017-12-13 | 2022-03-08 | 深圳先进技术研究院 | Coibatide A derivative and application thereof |
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| CN105646675B (en) | 2019-09-17 |
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