CN117700490B - Four-branched-chain polypeptide and preparation method thereof - Google Patents
Four-branched-chain polypeptide and preparation method thereof Download PDFInfo
- Publication number
- CN117700490B CN117700490B CN202310023681.2A CN202310023681A CN117700490B CN 117700490 B CN117700490 B CN 117700490B CN 202310023681 A CN202310023681 A CN 202310023681A CN 117700490 B CN117700490 B CN 117700490B
- Authority
- CN
- China
- Prior art keywords
- kkk
- polypeptide
- conh
- fmoc
- branched
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 77
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 77
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 77
- 238000002360 preparation method Methods 0.000 title abstract description 13
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 claims abstract description 16
- 230000002194 synthesizing effect Effects 0.000 claims abstract description 10
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 66
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 50
- 238000000034 method Methods 0.000 claims description 34
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 claims description 26
- 150000001413 amino acids Chemical class 0.000 claims description 24
- 238000006243 chemical reaction Methods 0.000 claims description 24
- 238000001514 detection method Methods 0.000 claims description 24
- 238000005406 washing Methods 0.000 claims description 24
- 239000002244 precipitate Substances 0.000 claims description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 18
- 238000001556 precipitation Methods 0.000 claims description 17
- 239000011347 resin Substances 0.000 claims description 17
- 229920005989 resin Polymers 0.000 claims description 17
- 238000004140 cleaning Methods 0.000 claims description 15
- 238000005336 cracking Methods 0.000 claims description 15
- 239000013049 sediment Substances 0.000 claims description 15
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 14
- 238000009833 condensation Methods 0.000 claims description 12
- 230000005494 condensation Effects 0.000 claims description 12
- UMRUUWFGLGNQLI-QFIPXVFZSA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-6-[(2-methylpropan-2-yl)oxycarbonylamino]hexanoic acid Chemical compound C1=CC=C2C(COC(=O)N[C@@H](CCCCNC(=O)OC(C)(C)C)C(O)=O)C3=CC=CC=C3C2=C1 UMRUUWFGLGNQLI-QFIPXVFZSA-N 0.000 claims description 10
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 10
- 125000003963 dichloro group Chemical group Cl* 0.000 claims description 10
- 230000001376 precipitating effect Effects 0.000 claims description 10
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 claims description 9
- 239000000047 product Substances 0.000 claims description 8
- 230000008961 swelling Effects 0.000 claims description 8
- WMSUFWLPZLCIHP-UHFFFAOYSA-N (2,5-dioxopyrrolidin-1-yl) 9h-fluoren-9-ylmethyl carbonate Chemical compound C12=CC=CC=C2C2=CC=CC=C2C1COC(=O)ON1C(=O)CCC1=O WMSUFWLPZLCIHP-UHFFFAOYSA-N 0.000 claims description 7
- 239000007788 liquid Substances 0.000 claims description 7
- 239000000243 solution Substances 0.000 claims description 7
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 6
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 5
- 239000007864 aqueous solution Substances 0.000 claims description 5
- 238000001035 drying Methods 0.000 claims description 5
- 150000003839 salts Chemical class 0.000 claims description 5
- 238000004090 dissolution Methods 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- 239000004472 Lysine Substances 0.000 claims description 3
- 230000000903 blocking effect Effects 0.000 claims description 3
- 238000003776 cleavage reaction Methods 0.000 claims description 3
- 125000003588 lysine group Chemical group [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 claims description 3
- 238000001226 reprecipitation Methods 0.000 claims description 3
- 238000002390 rotary evaporation Methods 0.000 claims description 3
- 239000000779 smoke Substances 0.000 claims description 3
- 239000007787 solid Substances 0.000 claims description 3
- 239000002904 solvent Substances 0.000 claims description 3
- 230000008602 contraction Effects 0.000 claims description 2
- 238000005119 centrifugation Methods 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 claims 1
- 230000015572 biosynthetic process Effects 0.000 abstract description 20
- 238000003786 synthesis reaction Methods 0.000 abstract description 20
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical group OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 16
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 10
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 8
- 239000003814 drug Substances 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 5
- 238000004108 freeze drying Methods 0.000 description 5
- 238000002514 liquid chromatography mass spectrum Methods 0.000 description 5
- 108020004707 nucleic acids Proteins 0.000 description 5
- 102000039446 nucleic acids Human genes 0.000 description 5
- 150000007523 nucleic acids Chemical class 0.000 description 5
- 239000012071 phase Substances 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 108020004459 Small interfering RNA Proteins 0.000 description 3
- 239000006227 byproduct Substances 0.000 description 3
- 238000009472 formulation Methods 0.000 description 3
- 230000014759 maintenance of location Effects 0.000 description 3
- 239000003960 organic solvent Substances 0.000 description 3
- 238000010532 solid phase synthesis reaction Methods 0.000 description 3
- 238000001308 synthesis method Methods 0.000 description 3
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 229920002521 macromolecule Polymers 0.000 description 2
- 239000002105 nanoparticle Substances 0.000 description 2
- 210000000056 organ Anatomy 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 108091030071 RNAI Proteins 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000006065 biodegradation reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000009194 climbing Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 238000009509 drug development Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000005611 electricity Effects 0.000 description 1
- 230000009881 electrostatic interaction Effects 0.000 description 1
- 238000005538 encapsulation Methods 0.000 description 1
- 230000009368 gene silencing by RNA Effects 0.000 description 1
- 230000037440 gene silencing effect Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- 108091070501 miRNA Proteins 0.000 description 1
- 239000002679 microRNA Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 238000010189 synthetic method Methods 0.000 description 1
- 238000007910 systemic administration Methods 0.000 description 1
- 238000012385 systemic delivery Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000012384 transportation and delivery Methods 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
- A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7088—Compounds having three or more nucleosides or nucleotides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7088—Compounds having three or more nucleosides or nucleotides
- A61K31/7105—Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
- A61K47/6921—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
- A61K47/6927—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores
- A61K47/6929—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K48/0008—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K48/005—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- General Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Genetics & Genomics (AREA)
- Hematology (AREA)
- Oncology (AREA)
- Communicable Diseases (AREA)
- Diabetes (AREA)
- Biochemistry (AREA)
- Immunology (AREA)
- Nanotechnology (AREA)
- Obesity (AREA)
- Peptides Or Proteins (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a four-branched-chain polypeptide and a preparation method thereof, wherein the preparation method of the four-branched-chain R 4-KKK-CONH2 comprises the following steps: step 1, synthesizing a polypeptide Fmoc-KKK-OH; step 2, synthesizing a main chain Fmoc 4 -KKK-OH; step 3, four branched polypeptide KW 4-KKK-CONH2 is synthesized. The preparation method of the four-branched-chain polypeptide R 4-KKK-CONH2 provided by the invention has the advantages that the main chain is separated from the side chain synthesis step, the main chain can be connected with linear polypeptides with any length to form branched polypeptides with different sequences, the applicability is wide, the purity is up to more than 94%, and a new thought is provided for the synthesis of the four-branched-chain polypeptide.
Description
Technical Field
The invention relates to the technical field of polypeptide synthesis, in particular to a four-branched-chain polypeptide and a preparation method thereof.
Background
The characteristics of the branched polypeptide macromolecule mainly comprise the following aspects: (1) Can be entangled with RNA through the interaction of static electricity and hydrogen bond, thereby assembling nano particles for wrapping RNA, and the encapsulation efficiency is high; (2) Can selectively target various tissues and cell types, and can be used for local delivery or systemic delivery; (3) Multiple siRNA with different targets can be carried at the same time, so that the siRNA can generate a synergistic gene silencing effect in the same target cell, and the curative effect of the RNAi is improved; (4) The application range is wider, and the preparation method can be used for wrapping various medicines such as siRNA, miRNA, mRNA and the like; (5) The natural amino acid composition can be used for biodegradation in cells, and has lower toxicity; (6) good water solubility. Because of these characteristics, the polypeptide macromolecule has wide potential application in the fields of biology and medicine.
However, the current branched polypeptide synthesis has the following problems:
(1) The reported synthesis methods of branched polypeptides are few in China.
(2) In the reported solid-phase synthesis method of the polypeptide, when the next amino acid is connected in the synthesis process, the amino acid and a condensing agent are directly added into a polypeptide reaction tube, and the reaction is insufficient to generate byproducts.
(3) The reported solid-phase synthesis method of the polypeptide has low purity and has the problem of trifluoroacetic acid residue, and if HPLC is used for purification, the purification efficiency is low and the yield loss is large.
(4) The reported solid-phase synthesis method of the polypeptide has the advantages that the finally obtained polypeptide contains water, and the water needs to be removed by a freeze-drying technology, so that the time is relatively consumed.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides four-branched-chain polypeptide R 4-KKK-CONH2 and a preparation method thereof, which mainly solve the problems of more byproducts, low purity of final products, trifluoroacetic acid residues, time consuming and complicated freeze-drying and water removal process and the like in the polypeptide synthesis process.
In order to solve the technical problems, the technical scheme of the invention is as follows:
In a first aspect of the present invention, there is provided a four-branched polypeptide having the structural formula:
Wherein K is lysine, R is KW sequence KWHHHKWHHHKWHHHKWHHHK, KHW sequence KHWHKHWHKHWWHKHWHK or JQ1 sequence GALMKFWKKYLRSSRKRSSRKRTQ.
In a second aspect of the present invention, there is provided a method for preparing the four-branched polypeptide, comprising the steps of:
Step 1: swelling the dichloro resin, and then washing the dichloro resin by DMF (dimethyl formamide), and accessing the first Fmoc-Lys (Boc) -OH; after the reaction is finished, cleaning, end capping and cleaning are sequentially carried out; then deprotecting and cleaning again, preparing Lys condensation liquid, connecting a second Fmoc-Lys (Boc) -OH, repeating the second Lys step to connect a3 rd Fmoc-Lys (Boc) -OH, cleaning, then cracking, and precipitating after cracking reaction; removing TFA after the precipitation is dissolved, and then precipitating and removing water to obtain polypeptide Fmoc-KKK-OH;
Step 2: fmoc-KKK-OH and Fmoc-OSu are reacted under alkaline conditions to prepare Fmoc 4 -KKK-OH;
Step 3: swelling AM resin, washing DMF, deprotecting, washing, preparing a condensation solution, inoculating Fmoc 4 -KKK-OH, washing, blocking and washing sequentially after the reaction is finished, deprotecting, washing, inoculating a first amino acid condensation solution of a prepared side chain into a first side chain amino acid, washing, repeating the steps, synthesizing 4R side chains at the same time, and cracking after the reaction is finished; precipitation is carried out after the cracking reaction; removing TFA after the precipitation is dissolved, and then precipitating and removing water to obtain four-branched-chain polypeptide R 4-KKK-CONH2;
The R side chain is KW sequence KWHHHKWHHHKWHHHKWHHHK, KHW sequence KHWHKHWHKHWWHKHWHK or JQ1 sequence GALMKFWKKYLRSSRKRSSRKRTQ.
Preferably, the amount of each amino acid added in the step 1 is 3-5eq of the amount of the dichloro resin;
The end capping method involved in the step 1 is as follows: adding AcOH in an amount of 3-5eq of dichloro resin, and then adding TEA in an amount of 7-10eq; the end capping method involved in the step 3 is as follows: the amount of AcOH added is 3-5eq of the amount of AM resin, acOH is reacted with HBTU for 8-15min before DIPEA is added. The DIPEA formulation procedure was as follows: DIPEA: DMF volume ratio 1:4-5, DIPEA is added in an amount of 0.4-0.8ml per 0.1mmol of amino acid.
Preferably, the cleaning method in step 1 and step 3 is as follows: 2 times of DMF washing, 2 times of DCM swelling, 2 times of EtOH shrinkage, DCM swelling and DMF washing, and the reagent is used in an amount which is less than that of the resin.
Preferably, when the next amino acid is linked in the step 1 and the step 3, an amino acid condensation solution needs to be prepared in advance, and the amino acid and the HBTU are mixed according to the mass ratio of 1:0.8-1.2 is dissolved in DMF for 8-15min, then poured into a polypeptide reaction tube, and DIPEA is added. The DIPEA formulation procedure was as follows: DIPEA: DMF volume ratio 1:4-5; the amount of DIPEA added is 0.4-0.8ml per 0.1mmol amino acid.
Preferably, the precipitation is carried out after the cleavage reaction in step 1 and step 3; TFA was removed after dissolution of the precipitate, followed by reprecipitation, as follows: precipitating with glacial ethyl ether after cracking, separating out precipitate at 1-5deg.C, and centrifuging to obtain precipitate;
Precipitate removal TFA: dissolving the centrifuged sediment by adopting MeOH, dropwise adding (a plurality of drops of) TEA and TFA to react into salt, then dropwise adding into EA with the volume of 20-40 times of methanol to separate out sediment, centrifuging for 1-3min at 10000-15000r/min, dissolving the centrifuged sediment by using MeOH, dropwise adding into EA with the volume of 20-40 times of methanol to separate out sediment, centrifuging, repeating for 2-3 times until the dropwise adding of TEA does not generate white smoke any more, completely removing TFA residues in the polypeptide at the moment, and collecting the centrifuged sediment.
As a further preference, the precipitate collected after TFA removal is sonicated with MeOH, then rotary evaporated to a transparent film, and then added in a volume ratio of 3-20:1, separating out white turbidity from PE and EA, and spin-drying to obtain white powdery polypeptide. The water in the product can be carried out by dissolving the precipitate with the organic solvent for multiple times and spin-drying, so that the purpose of removing water is achieved, and the subsequent freeze-drying treatment of the product is not needed.
Preferably, step 2 consists in dissolving Fmoc-KKKK-OH in THF+MeOH (1:0.8-1.2) and adding TEA 7-10eq; fmoc-OSu was dissolved in THF and then slowly added dropwise to Fmoc-KKK-OH, and the reaction was stirred at room temperature for 1-3 hours after the completion of the addition. Each 100mg of Fmoc-KKK-OH was dissolved in 10-30ml of THF+MeOH (1:0.8-1.2); fmoc-OSu was dissolved in 1-10mL of THF per 100 mg. The mass ratio of Fmoc-KKK-OH, fmoc-OSu and TEA was 1:3-7:7-10.
Concentrating the solvent by rotary evaporation after the reaction in the step 2 is finished, adding 1-3 times of EA and 2-6 times of PE, uniformly mixing, placing in a refrigerator at-20 to-25 ℃ for overnight, centrifuging for 1-3min to remove supernatant after 10000-15000r/min of precipitation is separated out, and using the volume ratio of lower layer solids to be 2-4:1, dropwise adding the solution of DCM and MeOH into EA with 20-40 times (the total volume of the dissolved polypeptide of DCM and MeOH) volume, uniformly mixing, centrifuging, removing supernatant, and repeating the step for 3-4 times to obtain a purer product.
Preferably, the HPLC detection conditions of R 4-KKK-CONH2 are: a C8 column; column temperature: 30-40 ℃; flow rate: 1ml/min; detection wavelength: 220nm; mobile phase: a0.05wt% phosphoric acid aqueous solution, B: HPLC methanol; the method comprises the following steps: 0 to 20min,65% A to 40% A; 20- > 30min,40% A- > 0% A.
According to the preparation method of the four-branched-chain polypeptide R 4-KKK-CONH2, the main chain and the side chain synthesis steps are separated, the main chain can be connected with linear polypeptides with any length to form branched polypeptides with different sequences, the applicability is wide, the purity is up to more than 94%, and a new thought is provided for the synthesis of the four-branched-chain polypeptide.
In a third aspect of the invention, there is provided the use of the four-branched polypeptide. By 2028, the market value of nucleic acid pharmaceuticals is expected to reach $100 billion. The four-branched-chain polypeptide can overcome biological barriers, realize high-efficiency transmembrane and effective in-vivo transportation, deliver nucleic acid drugs to other organs except liver in a local or systemic administration mode, has the potential of targeting other organs and tissues, is a bottleneck problem to be solved urgently in nucleic acid drug development, can be combined with nucleic acid through electrostatic interaction to form nano particles, and is an effective nucleic acid drug delivery system.
Compared with the prior art, the invention has the following beneficial effects:
(1) The invention discloses a method for synthesizing four-branched-chain polypeptides, which can be connected with R side chain polypeptides with any length after the synthesis of main chain Fmoc 4 -KKK-OH to obtain a plurality of four-branched-chain polypeptides R 4-KKK-CONH2 and provides a new idea for synthesizing branched-chain polypeptides.
(2) The cleaning and end capping steps of the invention can reduce the generation of byproducts in the polypeptide synthesis process, thereby improving the purity of the polypeptide.
(4) The ethyl acetate precipitation method adopted by the invention can efficiently remove the residual trifluoroacetic acid in the polypeptide, and has simple and rapid operation and the purity of more than 94%.
(5) According to the invention, the water in the product can be taken out to achieve the purpose of removing water through dissolving the precipitate with the organic solvent for multiple times and spin-drying, so that the powdery polypeptide can be quickly and efficiently obtained without freeze-drying and water removal.
Drawings
FIG. 1R4-KKK-CONH 2 synthetic route;
FIG. 2Fmoc-KKK-OH synthesis steps;
FIG. 3 LC-MS spectrum of Fmoc-KKK-OH;
FIG. 4Fmoc 4-KKK-OH;
FIG. 5 LC-MS spectrum of Fmoc 4-KKK-OH;
FIG. 6 is a diagram of the synthetic steps of R4-KKK-CONH 2;
FIG. 7 LC-MS spectrum of KW4-KKK-CONH 2;
FIG. 8 HPLC detection pattern of KW4-KKK-CONH 2;
FIG. 9 LC-MS spectrum of KHW4-KKK-CONH 2;
FIG. 10 HPLC detection pattern of KHW4-KKK-CONH 2;
FIG. 11 LC-MS spectrum of JQ14-KKK-CONH 2;
FIG. 12 HPLC detection pattern of JQ14-KKK-CONH 2.
Detailed Description
The following describes the technical scheme of the present invention in further detail with reference to the accompanying drawings and specific examples, but the present invention is not limited to the following technical scheme.
The following examples refer to the english abbreviations as shown in table 1.
TABLE 1
Example 1
The invention aims to provide a preparation method of four-branched-chain polypeptide R 4-KKK-CONH2, which has the structure shown as follows:
Wherein K is lysine, R is any linear polypeptide sequence of 1-20 amino acids in different permutation and combination, including but not limited to KW sequence (KWHHHKWHHHKWHHHKWHHHK) and KHW sequence (KHWHKHWHKHWWHKHWHK) and JQ1 sequence (GALMKFWKKYLRSSRKRSSRKRTQ).
The synthesis steps of the four-branched polypeptide are shown in figure 1.
Step 1: synthetic polypeptide Fmoc-KKK-OH
3 Fmoc-Lys (Boc) -OH were attached to dichloro resin: swelling the dichloro resin, washing with DMF and then accessing the first Fmoc-Lys (Boc) -OH; after the reaction is finished, cleaning, end capping and cleaning are sequentially carried out; then deprotecting and cleaning again, preparing Lys condensation liquid, connecting Fmoc-Lys (Boc) -OH, repeating the second Lys step to connect 3 rd Fmoc-Lys (Boc) -OH, cleaning and then cracking, and precipitating after cracking reaction; after dissolution of the precipitate, TFA is removed, followed by precipitation and removal of water to obtain the polypeptide Fmoc-KKK-OH.
The amount of each amino acid added in the step 1 is 4eq of the amount of the dichloro resin;
The end capping method involved in step 1 is: the amount of AcOH added is 4eq of the amount of dichloro resin, then TEA 8eq; the end capping method involved in the step 3 is as follows: the amount of AcOH added was 4eq of AM resin, which was reacted with HBTU for 10min before DIPEA was added.
LC-MS detection of Fmoc-KKK-OH is shown in FIG. 3: positive ion mode is selected, m/z range: 200-800, flag=15.0v.
LC-MS results analysis: the precise molecular weight of Fmoc-KKK-OH is: 624.4. the detection molecular weight is as follows: 625.2[ M+H ] +,313.2[M+2H]2+,209.0[M+3H]3+, the synthesized Fmoc-KKK-OH structure was correct.
Step 2: synthesis of backbone Fmoc 4 -KKK-OH
Fmoc-KKK-OH was reacted with Fmoc-OSu under alkaline conditions to produce Fmoc 4 -KKK-OH of the formula shown in FIG. 4.
(1) Fmoc-KKK-OH was dissolved in 10mLTHF+10mLMeOH, added 455. Mu. lTEA, and placed in a round bottom flask; fmoc-OSu was added to 10mL of HF and placed in a constant pressure dropping funnel, placed on a round bottom flask, slowly added dropwise for 20min, and after the addition was completed, the reaction was stirred at room temperature for 2h.
(2) The solvent was concentrated to 5ml by rotary evaporation, 30mLEA and 10mLPE were added and mixed well, and the mixture was placed in a refrigerator at-20℃overnight, and the precipitation was waited for.
(3) 12000R/min, centrifuging for 2min, removing supernatant, dissolving solid with 3mLDCM+1mLMeOH ultrasound, adding 30mLEA, mixing, centrifuging, removing supernatant, repeating the steps for 3-4 times, and climbing up the plate to see whether impurity is removed completely.
(4) LC-MS detection of Fmoc 4 -KKK-OH is shown in FIG. 5: negative positive ion mode is selected, m/z range: 900-1400, flag=15.0v.
LC-MS results analysis: the precise molecular weight of Fmoc 4 -KKK-OH is: 1290.57. the detection molecular weight is as follows: 1289.4[ M-H ] -,1325.4[M+Cl]-, the synthesized Fmoc 4 -KKK-OH was structurally correct.
Step 3: synthesis of four branched polypeptide R 4-KKK-CONH2
Swelling AM resin, washing DMF, deprotecting, washing, preparing condensation liquid, inoculating Fmoc 4 -KKK-OH, washing, blocking and washing after the reaction, deprotecting, washing, inoculating the first amino acid condensation liquid of the prepared side chain into the first side chain amino acid, washing, repeating the steps, synthesizing 4R side chains at the same time, and cracking after the reaction is finished; precipitation is carried out after the cracking reaction; removing TFA after the precipitation is dissolved, and then precipitating and removing water to obtain four-branched-chain polypeptide R 4-KKK-CONH2;
the cleaning method involved in the step 1 and the step 3 is as follows: 2 DMF washes, 2 DCM swells, 2 EtOH contractions, DCM swells and DMF washes; the usage amount is not limited by the resin.
When the next amino acid is connected in the step 1 and the step 3, an amino acid condensation liquid needs to be prepared in advance, and the amino acid and the HBTU are mixed according to the mass ratio of 1:1 in DMF for 10min, then poured into a polypeptide reaction tube, and DIPEA was added. The DIPEA formulation procedure was as follows: DIPEA: DMF volume ratio 174:826 was added in an amount of 0.6ml per 0.1mmol of amino acid.
Precipitation is carried out after the cleavage reaction in the step 1 and the step 3; TFA is removed after dissolution of the precipitate, followed by reprecipitation and water removal, as follows: precipitating with glacial ethyl ether after cracking, separating out precipitate at 1-5deg.C, and centrifuging to obtain precipitate;
Precipitate removal TFA: dissolving the centrifuged sediment by adopting a small amount of MeOH, dropwise adding a trace amount of TEA to react with TFA to form salt, then dropwise adding the salt into 30 times of volume EA to separate out sediment, centrifuging for 2min at 12000r/min, dissolving the centrifuged sediment by using MeOH, dropwise adding TEA into the EA to separate out sediment, centrifuging, repeating for 2-3 times until the TEA is dropwise added to prevent white smoke, completely removing TFA residues in the polypeptide, and collecting the centrifuged sediment.
The precipitate collected after TFA removal was sonicated with a small amount of MeOH, then rotary evaporated to a clear film, then PE and EA were added to precipitate a white turbidity, and dried to give the polypeptide as a white powder. The water in the product can be carried out by dissolving the precipitate with the organic solvent for multiple times and spin-drying, so that the purpose of removing water is achieved, and the subsequent freeze-drying treatment of the product is not needed.
The Fmoc-KKK-OH synthesis procedure is shown in FIG. 2 and the R 4-KKK-CONH2 synthesis procedure is shown in FIG. 6.
Example 2
The structure of the synthesis method of the four-branched-chain polypeptide KW 4-KKK-CONH2 is shown as follows, and the preparation method comprises the following steps:
Step 1: synthesizing a polypeptide Fmoc-KKK-OH;
Step 2: synthesizing a main chain Fmoc 4 -KKK-OH;
Step 3: synthesizing four branched chain polypeptide KW 4-KKK-CONH2;
LCMS detection results are shown in fig. 7: positive ion mode is selected, m/z range: 100-2200, flag=100.0v.
LC-MS results analysis: the precise molecular weight of KW 4-KKK-CONH2 is: 12524.32. the detection molecular weight is as follows: 118.1[ M+107H ] 107+,235.3[M+59Na]59+, the resultant KW 4-KKK-CONH2 was structurally correct.
HPLC detection conditions and results:
Column: c8 (10 x 250mm,5 μm); column temperature: 35 ℃; flow rate: 1ml/min; detection wavelength: 220nm; mobile phase: a0.05% phosphoric acid aqueous solution, BHPLC methanol; the method comprises the following steps: 0 to 20min,65% A to 40% A,20 to 30min,40% A to 0% A; retention time: 7.328min; purity: 99.1% and the HPLC chromatogram is shown in FIG. 8.
Example 3
The structure of the synthetic method of the four-branched polypeptide KHW 4-KKK-CONH2 is shown as follows, and the preparation method comprises the following steps:
step 1: synthetic polypeptide Fmoc-KKK-OH
Step 2: synthesis of backbone Fmoc 4 -KKK-OH
Step 3: synthesis of four branched polypeptide KHW 4-KKK-CONH2
The LC-MS detection results are shown in fig. 9: positive ion mode is selected, m/z range: 200-1400, flag=50.0v.
LC-MS results analysis: the exact molecular weight of KHW 4-KKK-CONH2 is: 11963.97. KHW 4-KKK-CONH2 synthesized with molecular weight ：227.2[M+53H]53+,268.1[M+52K]52+,317.1[M+43K]43+,380.8[M+35K]35+,449.3[M+28Na]28+,542.4[M+23Na]23+,704.2[M+17H]17+, was detected to have the correct structure.
HPLC detection conditions and results:
Column: c8 (10 x 250mm,5 μm); column temperature: 35 ℃; flow rate: 1ml/min; detection wavelength: 220nm; mobile phase: a0.05% phosphoric acid aqueous solution, BHPLC methanol; the method comprises the following steps: 0 to 20min,65% A to 40% A,20 to 30min,40% A to 0% A; retention time: 7.548min; purity: 99.1%, and the HPLC detection result is shown in FIG. 10.
Example 4
The synthesis method of the four-branched-chain polypeptide JQ1 4-KKK-CONH2 has the structure shown as follows, and the preparation method comprises the following steps:
step 1: synthetic polypeptide Fmoc-KKK-OH
Step 2: synthesis of backbone Fmoc 4 -KKK-OH
Step 3: synthesis of four branched polypeptide JQ1 4-KKK-CONH2
The LC-MS detection results are shown in fig. 11: positive ion mode is selected, m/z range: 600-900, flag=135.0v.
LC-MS results analysis: the exact molecular weight of JQ1 4-KKK-CONH2 is: 12325.1. the JQ1 4-KKK-CONH2 synthesized by the detection molecular weight ：638.7[M+20Na]20+,648.1[M+19H]19+,689.0[M+19K]19+,725.7[M+17H]17+,763.7[M+17K]17+,771.2[M+16H]16+,808.4[M+16K]16+,821.5[M+15H]15+,843.6[M+15Na]15+, has the correct structure.
HPLC detection conditions and results:
Column: c8 (10 x 250mm,5 μm); column temperature: 35 ℃; flow rate: 1ml/min; detection wavelength: 220nm; mobile phase: a0.05% phosphoric acid aqueous solution, BHPLC methanol; the method comprises the following steps: 0 to 20min,65% A to 40% A,20 to 30min,40% A to 0% A; retention time: 7.477min; purity: 93.9% and the HPLC detection result is shown in FIG. 12.
It should be noted that the above-mentioned embodiments are only preferred embodiments of the present invention, and are not intended to limit the scope of the present invention, and the equivalent substitutions or alternatives made on the basis of the above description are all included in the scope of the present invention.
Claims (8)
1. A four-branched polypeptide R 4-KKK-CONH2, characterized in that: the structural general formula of the four-branched-chain polypeptide R 4-KKK-CONH2 is as follows:
wherein K is lysine, 4R are KW sequence KWHHHKWHHHKWHHHKWHHHK or 4R are KHW sequence KHWHKHWHKHWWHKHWHK.
2. The method for preparing the four-branched polypeptide R 4-KKK-CONH2 according to claim 1, wherein: the method comprises the following steps:
Step 1: swelling the dichloro resin, and then washing the dichloro resin by DMF (dimethyl formamide), and accessing the first Fmoc-Lys (Boc) -OH; after the reaction is finished, cleaning, end capping and cleaning are sequentially carried out; then deprotecting and cleaning again, preparing Lys condensation liquid, connecting a second Fmoc-Lys (Boc) -OH, repeating the second Lys step to connect a3 rd Fmoc-Lys (Boc) -OH, cleaning, then cracking, and precipitating after cracking reaction; removing TFA after the precipitation is dissolved, and then precipitating and removing water to obtain polypeptide Fmoc-KKK-OH;
Step 2: fmoc-KKK-OH and Fmoc-OSu are reacted under alkaline conditions to prepare Fmoc 4 -KKK-OH;
Step 3: swelling AM resin, washing DMF, deprotecting, washing, preparing a condensation solution, inoculating Fmoc 4 -KKK-OH, washing, blocking and washing sequentially after the reaction is finished, deprotecting, washing, inoculating a first amino acid condensation solution of a prepared side chain into a first side chain amino acid, washing, repeating the steps, synthesizing 4R side chains at the same time, and cracking after the reaction is finished; precipitation is carried out after the cracking reaction; after the precipitation is dissolved, TFA is removed, and then the precipitation and water removal are carried out, so that the four-branched-chain polypeptide R 4-KKK-CONH2 is obtained.
3. The method for preparing a four-branched polypeptide R 4-KKK-CONH2 according to claim 2, wherein the washing methods involved in step 1 and step 3 are: 2 DMF washes, 2 DCM swells, 2 EtOH contractions, DCM swells and DMF washes.
4. The method for preparing the four-branched polypeptide R 4-KKK-CONH2 according to claim 2, wherein: when the next amino acid is connected in the step 1 and the step 3, an amino acid condensation liquid needs to be prepared in advance, and the amino acid and the HBTU are mixed according to the mass ratio of 1:0.8-1.2 is dissolved in DMF for 8-15min, then poured into a polypeptide reaction tube, and DIPEA is added.
5. The method for preparing the four-branched polypeptide R 4-KKK-CONH2 according to claim 2, wherein:
Precipitation is carried out after the cleavage reaction in the step 1 and the step 3; TFA was removed after dissolution of the precipitate, followed by reprecipitation, as follows: precipitating with glacial ethyl ether after cracking, separating out precipitate at 1-5deg.C, and centrifuging to obtain precipitate;
Precipitate removal TFA: dissolving the centrifuged sediment by adopting MeOH, dropwise adding TEA and TFA to react to form salt, then dropwise adding the salt into EA with 20-40 times of the volume of the MeOH to separate out the sediment, centrifuging for 1-3min at 10000-15000r/min, dissolving the centrifuged sediment by using MeOH, dropwise adding TEA into EA with 20-40 times of the volume to separate out the sediment, centrifuging, repeating for 2-3 times until the TEA is dropwise added to prevent white smoke, completely removing TFA residues in the polypeptide at the moment, and collecting the centrifuged sediment.
6. The method for producing four-branched polypeptide R 4-KKK-CONH2 according to claim 5, wherein the precipitate collected after TFA removal is sonicated with MeOH, then rotary evaporated to a transparent film, and then added in a volume ratio of 3-20:1, separating out white turbidity from PE and EA, and spin-drying to obtain white powdery polypeptide.
7. The method for preparing the four-branched polypeptide R 4-KKK-CONH2 as claimed in claim 2, wherein after the reaction in the step 2 is completed, the solvent is concentrated by rotary evaporation, 1-3 times of volume EA and 2-6 times of volume PE are added and mixed uniformly, the mixture is placed in a refrigerator at-20 to-25 ℃ for overnight, 10000-15000R/min is carried out after precipitation, the supernatant is removed by centrifugation of 1-3 min, and the volume ratio of the lower layer solid is 2-4:1, dropwise adding the solution of DCM and MeOH into EA with 20-40 times of volume, uniformly mixing, centrifuging, removing supernatant, and repeating the steps for 3-4 times to obtain a purer product.
8. The method of preparing a four-branched polypeptide R 4-KKK-CONH2 according to claim 2, wherein the HPLC detection conditions of R 4-KKK-CONH2 are: a C8 column; column temperature: 30-40 ℃; flow rate: 1ml/min; detection wavelength: 220 nm; mobile phase: a0.05wt% phosphoric acid aqueous solution, B: HPLC methanol; the method comprises the following steps: 0 to 20 min,65% A to 40% A; 20- > 30 min,40% A- > 0% A.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2022117327373 | 2022-12-30 | ||
| CN202211732737 | 2022-12-30 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117700490A CN117700490A (en) | 2024-03-15 |
| CN117700490B true CN117700490B (en) | 2024-06-11 |
Family
ID=90157626
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310023681.2A Active CN117700490B (en) | 2022-12-30 | 2023-01-09 | Four-branched-chain polypeptide and preparation method thereof |
| CN202310632432.3A Pending CN117695405A (en) | 2022-12-30 | 2023-05-31 | Polypeptide delivery system for wrapping nucleic acid drug and application thereof |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310632432.3A Pending CN117695405A (en) | 2022-12-30 | 2023-05-31 | Polypeptide delivery system for wrapping nucleic acid drug and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN117700490B (en) |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2020134974A3 (en) * | 2020-10-26 | 2022-04-26 |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2171531C (en) * | 1993-09-13 | 2005-11-22 | Jean Marc Sabatier | Multiple branch peptide constructions for use against hiv |
| US7070807B2 (en) * | 1999-12-29 | 2006-07-04 | Mixson A James | Branched histidine copolymers and methods for using same |
| AU2005310131A1 (en) * | 2004-11-17 | 2006-06-08 | University Of Maryland, Baltimore | Highly branched HK peptides as effective carriers of siRNA |
| EP2782926B1 (en) * | 2011-11-24 | 2018-07-04 | Positec Power Tools (Suzhou) Co., Ltd | Peptide sequence design and use thereof for peptide-mediated sirna delivery |
| CN104725478B (en) * | 2013-12-18 | 2018-12-25 | 纳肽得有限公司 | Polypeptide compound, the assembly of polypeptide compound and siRNA and its application |
-
2023
- 2023-01-09 CN CN202310023681.2A patent/CN117700490B/en active Active
- 2023-05-31 CN CN202310632432.3A patent/CN117695405A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2020134974A3 (en) * | 2020-10-26 | 2022-04-26 |
Non-Patent Citations (2)
| Title |
|---|
| Highly branched HK peptides are effective carriers of siRNA;Qixin Leng等;《J Gene Med》;20050731;第7卷(第7期);第977-986页 * |
| 聚赖氨酸的生物医学领域应用现状;余波等;《现代药物与临床》;20221013;第37卷(第10期);第2377-2385页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117700490A (en) | 2024-03-15 |
| CN117695405A (en) | 2024-03-15 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| WO2007048190A1 (en) | Macromolecular compounds having controlled stoichiometry | |
| JP6947909B2 (en) | Multi-arm targeted anti-cancer conjugate | |
| WO2017198124A1 (en) | Multi-arm polymeric targeting anti-cancer conjugate | |
| CN113845571B (en) | Active polypeptide for inhibiting liver cancer cell growth and preparation method and application thereof | |
| US20090197315A1 (en) | Fullerene-based amino acids | |
| CN117700490B (en) | Four-branched-chain polypeptide and preparation method thereof | |
| CN108676074B (en) | Active polypeptide for promoting growth of liver cells | |
| CN1781933B (en) | Thymosin alpha 1 active segment cyclicpeptide analogue and its poly glycol derivative | |
| CN113845598B (en) | Protein targeting chimera degradation MDM2/MDMX protein staple peptide conjugate and application thereof | |
| CN113321706B (en) | Self-assembly polypeptide derivative and hydrogel prepared from same | |
| CN101302008B (en) | Solid phase synthesis method of single addition fullerene aminoacid derivate | |
| CN113801190B (en) | Preparation method of oligopeptide-1 hydrochloride | |
| CN109553658B (en) | Ang- (1-7) aza polypeptide analogue, preparation method and application | |
| US4058512A (en) | Synthetic peptides having growth promoting activity | |
| CN112279889B (en) | Polypeptide, derivative and application of polypeptide and derivative in preparation of antitumor drugs | |
| CN114796240B (en) | Self-assembled chiral short peptide drug and preparation method and application thereof | |
| CN117531021B (en) | Acanthopanax senticosus glycoside E-targeting peptide conjugate and application thereof | |
| CN117777245B (en) | Cordycepin-antibacterial peptide conjugate and synthetic method and application thereof | |
| CN116808231B (en) | Cell membrane penetrating peptide coupled sulpride prodrug system, preparation method and application | |
| CN111154002B (en) | Bifunctional peptide K14, gene vector and co-drug delivery system | |
| CN108912348B (en) | Short peptide hydrogel containing protein and method for improving storage stability of protein | |
| CN108295030A (en) | Bufalin micelle nano preparation and preparation method thereof | |
| CN117180446A (en) | Self-assembled polypeptide coupling medicine and application thereof | |
| CN116179609A (en) | Method for improving transfection efficiency of membrane-permeable polypeptide gene | |
| CN115317622A (en) | Preparation and application of anti-tumor polypeptide coupled drug |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |