CN116196432A - 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 - Google Patents
一种多级环境响应性控释外泌体复合材料及其制备方法与应用 Download PDFInfo
- Publication number
- CN116196432A CN116196432A CN202211649228.4A CN202211649228A CN116196432A CN 116196432 A CN116196432 A CN 116196432A CN 202211649228 A CN202211649228 A CN 202211649228A CN 116196432 A CN116196432 A CN 116196432A
- Authority
- CN
- China
- Prior art keywords
- exosome
- environment
- sensitive
- exosomes
- environmentally
- 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.)
- Pending
Links
- 210000001808 exosome Anatomy 0.000 title claims abstract description 89
- 239000002131 composite material Substances 0.000 title claims abstract description 32
- 238000013270 controlled release Methods 0.000 title claims abstract description 26
- 238000002360 preparation method Methods 0.000 title claims abstract description 15
- 239000000463 material Substances 0.000 claims abstract description 33
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 claims abstract description 24
- 125000003396 thiol group Chemical group [H]S* 0.000 claims abstract description 16
- -1 methacrylic acid modified hyaluronic acid Chemical class 0.000 claims abstract description 13
- 235000012000 cholesterol Nutrition 0.000 claims abstract description 11
- 230000015556 catabolic process Effects 0.000 claims abstract description 8
- 238000006731 degradation reaction Methods 0.000 claims abstract description 8
- 238000012650 click reaction Methods 0.000 claims abstract description 7
- 230000005284 excitation Effects 0.000 claims abstract description 7
- 239000002052 molecular layer Substances 0.000 claims abstract description 5
- 150000003904 phospholipids Chemical class 0.000 claims abstract description 5
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 claims abstract description 3
- 201000008482 osteoarthritis Diseases 0.000 claims description 6
- 229920001184 polypeptide Polymers 0.000 claims description 6
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 6
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 6
- 102000004169 proteins and genes Human genes 0.000 claims description 6
- 108090000623 proteins and genes Proteins 0.000 claims description 6
- 238000000034 method Methods 0.000 claims description 5
- 102000029750 ADAMTS Human genes 0.000 claims description 2
- 108091022879 ADAMTS Proteins 0.000 claims description 2
- 102000015696 Interleukins Human genes 0.000 claims description 2
- 108010063738 Interleukins Proteins 0.000 claims description 2
- 102000002274 Matrix Metalloproteinases Human genes 0.000 claims description 2
- 108010000684 Matrix Metalloproteinases Proteins 0.000 claims description 2
- 102000005741 Metalloproteases Human genes 0.000 claims description 2
- 108010006035 Metalloproteases Proteins 0.000 claims description 2
- 108060008682 Tumor Necrosis Factor Proteins 0.000 claims description 2
- 238000009792 diffusion process Methods 0.000 claims description 2
- 102000003390 tumor necrosis factor Human genes 0.000 claims description 2
- 229940079593 drug Drugs 0.000 claims 2
- 239000003814 drug Substances 0.000 claims 2
- 102000002938 Thrombospondin Human genes 0.000 claims 1
- 108060008245 Thrombospondin Proteins 0.000 claims 1
- 230000005855 radiation Effects 0.000 claims 1
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 abstract description 10
- 229920002674 hyaluronan Polymers 0.000 abstract description 10
- 229960003160 hyaluronic acid Drugs 0.000 abstract description 10
- 230000007613 environmental effect Effects 0.000 abstract description 5
- 238000006243 chemical reaction Methods 0.000 abstract description 4
- 230000000694 effects Effects 0.000 abstract description 3
- 239000007788 liquid Substances 0.000 abstract description 2
- 230000004043 responsiveness Effects 0.000 abstract 1
- 210000001519 tissue Anatomy 0.000 abstract 1
- 201000005488 Capillary Leak Syndrome Diseases 0.000 description 9
- 208000001353 Coffin-Lowry syndrome Diseases 0.000 description 9
- 239000000243 solution Substances 0.000 description 7
- PZBFGYYEXUXCOF-UHFFFAOYSA-N TCEP Chemical compound OC(=O)CCP(CCC(O)=O)CCC(O)=O PZBFGYYEXUXCOF-UHFFFAOYSA-N 0.000 description 6
- 230000009471 action Effects 0.000 description 5
- DCUFMVPCXCSVNP-UHFFFAOYSA-N methacrylic anhydride Chemical compound CC(=C)C(=O)OC(=O)C(C)=C DCUFMVPCXCSVNP-UHFFFAOYSA-N 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 238000003776 cleavage reaction Methods 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 235000018102 proteins Nutrition 0.000 description 3
- 230000007017 scission Effects 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- 125000004042 4-aminobutyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])N([H])[H] 0.000 description 2
- 102100027995 Collagenase 3 Human genes 0.000 description 2
- 108050005238 Collagenase 3 Proteins 0.000 description 2
- 108010003272 Hyaluronate lyase Proteins 0.000 description 2
- 102000001974 Hyaluronidases Human genes 0.000 description 2
- NWGKJDSIEKMTRX-AAZCQSIUSA-N Sorbitan monooleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OC[C@@H](O)[C@H]1OC[C@H](O)[C@H]1O NWGKJDSIEKMTRX-AAZCQSIUSA-N 0.000 description 2
- 150000001408 amides Chemical class 0.000 description 2
- 125000003277 amino group Chemical group 0.000 description 2
- 150000001840 cholesterol esters Chemical class 0.000 description 2
- 238000004132 cross linking Methods 0.000 description 2
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 2
- 235000018417 cysteine Nutrition 0.000 description 2
- 238000000502 dialysis Methods 0.000 description 2
- 238000013467 fragmentation Methods 0.000 description 2
- 238000006062 fragmentation reaction Methods 0.000 description 2
- 229960002773 hyaluronidase Drugs 0.000 description 2
- 229940057995 liquid paraffin Drugs 0.000 description 2
- 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 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 238000013112 stability test Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 229910021642 ultra pure water Inorganic materials 0.000 description 2
- 239000012498 ultrapure water Substances 0.000 description 2
- OOIBFPKQHULHSQ-UHFFFAOYSA-N (3-hydroxy-1-adamantyl) 2-methylprop-2-enoate Chemical compound C1C(C2)CC3CC2(O)CC1(OC(=O)C(=C)C)C3 OOIBFPKQHULHSQ-UHFFFAOYSA-N 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- PEEHTFAAVSWFBL-UHFFFAOYSA-N Maleimide Chemical compound O=C1NC(=O)C=C1 PEEHTFAAVSWFBL-UHFFFAOYSA-N 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 150000001841 cholesterols Chemical class 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000013268 sustained release Methods 0.000 description 1
- 239000012730 sustained-release form Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 230000017423 tissue regeneration Effects 0.000 description 1
- 238000009281 ultraviolet germicidal irradiation Methods 0.000 description 1
Images
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
- A61K47/54—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 an organic compound
- A61K47/554—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 an organic compound the modifying agent being a steroid plant sterol, glycyrrhetic acid, enoxolone or bile acid
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
-
- 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
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/16—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing nitrogen, e.g. nitro-, nitroso-, azo-compounds, nitriles, cyanates
- A61K47/18—Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
- A61K47/183—Amino acids, e.g. glycine, EDTA or aspartame
-
- 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
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/28—Steroids, e.g. cholesterol, bile acids or glycyrrhetinic acid
-
- 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
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
-
- 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
- A61K47/56—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 an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
- A61K47/61—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 an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule the organic macromolecular compound being a polysaccharide or a derivative thereof
-
- 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
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Veterinary Medicine (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Engineering & Computer Science (AREA)
- Epidemiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Rheumatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Physical Education & Sports Medicine (AREA)
- Immunology (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Dermatology (AREA)
- Molecular Biology (AREA)
- Virology (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Developmental Biology & Embryology (AREA)
- Biomedical Technology (AREA)
- Inorganic Chemistry (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Pain & Pain Management (AREA)
- Medicinal Preparation (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
Abstract
本发明公开了一种多级环境响应性控释外泌体复合材料及其制备方法与应用，属于生物医用材料领域。所述外泌体复合材料包括甲基丙烯酸改性的透明质酸、环境敏感连接分子和外泌体，所述环境敏感连接分子一端的胆固醇能够插入到外泌体的磷脂分子层从而与外泌体实现物理结合，另一端的巯基能够在紫外光照下与甲基丙烯酸改性的透明质酸发生点击反应而相互结合，从而实现外泌体与透明质酸的响应性结合。部分外泌体表面暴露有巯基，可以在紫外的激发下与甲基丙烯酸中的双键发生点击反应，从而直接与材料结合，这部分外泌体随材料降解释放，避免进入体内后被快速清除。所述复合材料进入体内后首先通过液体交换作用释放少量未与材料结合的游离的外泌体，随后在特定组织环境下发生环境敏感键断裂，从而在目标区域“精准释放”外泌体，最后其余与材料结合的外泌体能随着材料的降解随之释放，起到“长效缓释”的作用，从而实现多级环境响应性控释和缓释效果。
Description
技术领域
本发明属于生物医用材料领域，特别涉及到一种多级环境响应性控释外泌体复合材料及其制备方法与应用。
背景技术
外泌体在注射入体内后会被快速清除，因此构建外泌体的缓释体系显得极为重要。直接将外泌体与材料共混虽然较为方便简单，但是往往由于材料的孔径大以及与材料结合不紧密等问题，会造成外泌体的突释；将外泌体通过化学方法修饰与材料结合则有破坏外泌体的结构，影响外泌体功能的风险。因此，设计环境响应性的多级控释外泌体的材料则备受期待。
发明内容
针对现有技术中存在的上述问题，本发明提供了一种多级环境响应性控释外泌体复合材料及其制备方法与应用，所述外泌体复合材料可以根据不同应用环境进行定制设计，可以构建出三级环境响应性控释外泌体的缓释体系。
本发明通过以下技术方案来实现：
一种环境敏感连接分子，其特征在于，所述环境敏感连接分子一端为胆固醇，另一端修饰巯基。所述环境敏感连接分子一端的胆固醇能够插入到外泌体的磷脂分子层从而与外泌体实现物理结合，另一端的巯基能够在紫外光激发下与甲基丙烯酸酐改性的透明质酸中的双键发生点击反应，从而实现外泌体与透明质酸的响应性结合。作为可选方式，在上述环境敏感连接分子中，所述巯基与胆固醇之间还连接有环境敏感键。
作为可选方式，在上述环境敏感连接分子中，所述环境敏感键可选择的范围较多，包括pH响应性断裂的键(如弱酸下断裂的酰腙键)和紫外照射断裂的磷酸酯键，以及白介素家族蛋白(IL)、基质金属蛋白酶家族蛋白(MMP)、血小板反应蛋白解整合素金属肽酶家族蛋白(ADAMTS)、肿瘤坏死因子家族蛋白(TNF)等的响应性断裂键。进一步的，所述环境敏感键为骨关节炎环境感键，具体为MMP响应性断裂多肽(如RVGLP)，上述MMP响应性断裂多肽(RVGLP)能够在MMP-13存在的情况下发生断裂。
作为可选方式，在上述环境敏感连接分子具体为：CLS-RVGLP-SH。
本发明还提供了一种多级环境响应性控释外泌体复合材料，包括甲基丙烯酸酐改性的透明质酸、本发明所述的环境敏感连接分子和外泌体，所述甲基丙烯酸酐改性的透明质酸通过巯基与环境敏感连接分子端部的马来酰亚胺相联，所述环境敏感连接分子端部的胆固醇能够插入到所述外泌体的磷脂分子层，从而使外泌体与巯基化透明质酸通过环境敏感连接分子形成交联稳定的结构。
作为可选方式，在上述多级环境响应性控释外泌体复合材料中，所述环境敏感连接分子为CLS-MMP响应性断裂多肽-SH。
作为可选方式，在上述多级环境响应性控释外泌体复合材料中，部分所述外泌体表面暴露有巯基(外泌体在TCEP的作用下表面蛋白的二硫键被还原可暴露巯基)，能够在紫外光的激发下与甲基丙烯酸中的双键发生点击反应，从而直接与材料结合，这部分外泌体能够随材料降解释放，所述的外泌体也有部分直接与材料共混，能够通过扩散作用释放。
作为可选方式，在上述多级环境响应性控释外泌体复合材料中，所述复合材料为可注射凝胶、微凝胶或支架材料。
本发明还提供了一种上述的环境敏感连接分子的制备方法，其特征在于，包括以下步骤：
1)合成RVGLP多肽时，两端分别接C(半胱氨酸)和K(赖氨酸)序列；利用赖氨酸侧链的氨基与胆固醇酯琥珀酸单酯通过酰胺反应接上胆固醇基团，即形成SH-RVGLP-CLS(巯基-MMP响应性断裂多肽-胆固醇)。
本发明还提供了一种上述的多级环境响应性控释外泌体复合材料的制备方法，其特征在于，包括以下步骤：
(1)制备甲基丙烯酸酐改性的透明质酸；
(2)制备环境敏感连接分子；
(3)制备缓释复合材料。
作为可选方式，在上述多级环境响应性控释外泌体复合材料的制备方法中，所述步骤(1)具体为：将透明质酸(HA)粉末溶入超纯水中，搅拌均匀后调至弱碱性，滴加甲基丙烯酸酐，冰浴下反应4h，随后室温搅拌过夜。透析后冷冻干燥得到海绵状固体的甲基丙烯酸改性的透明质酸(HA-MA)。
作为可选方式，在上述多级环境响应性控释外泌体复合材料的制备方法中，所述步骤(3)具体为：将SH-RVGLP-CLS用TCEP处理后，4℃外泌体共混过夜，与溶解的HA-MA混合(调pH至中性)。司班80混于液态石蜡中搅拌均匀，将混合的溶液滴加至油相中，改变转速得到不同粒径，经紫外光照后交联形成的载外泌体的微凝胶体系。
作为可选方式，在上述多级环境响应性控释外泌体复合材料的制备方法中，所述步骤(3)可以进一步根据病症调控3级释放外泌体的配比，如在SH-RVGLP-CLS用TCEP处理后，4℃下添加1份外泌体共混过夜，另外2份外泌体分别用或不用TCEP处理(得到EXO-SH和EXO)，将随后将响应性释放分子修饰的外泌体，暴露巯基的外泌体和纯外泌体与溶解的HAMA共混，在紫外光照下制备微凝胶。
本发明还提供了一种上述的环境敏感连接分子的应用，其特征在于，将其用于制备骨关节炎治疗体系。
本发明还提供了一种上述的多级环境响应性控释外泌体复合材料的应用，其特征在于，将其用于制备骨关节炎治疗材料体系。
本说明书中公开的所有特征，或公开的所有方法或过程中的步骤，除了互相排斥的特征和/或步骤以外，均可以以任何方式组合。
本发明的有益效果：
(1)本发明所述环境敏感连接分子一端的胆固醇能够插入到外泌体的磷脂分子层从而与外泌体实现物理结合，另一端的巯基能够在紫外光激发下与甲基丙烯酸酐改性的透明质酸中的双键发生点击反应而相互结合，从而实现外泌体与透明质酸的响应性结合。另一方面暴露巯基的外泌体也可以在紫外光激发下与甲基丙烯酸酐改性的透明质酸中的双键发生点击反应而相互结合从而实现外泌体与材料的稳定结合，随材料降解而释放，避免进入体内后被快速清除。
(2)本发明所述外泌体复合材料具备多级环境响应的控释和缓释效果，进入体内后，由于符合材料液体交换作用下，材料中存在的少量未与材料结合的游离的外泌体会逐步释放，成为组织修复的“先头兵”。其次，在骨关节炎中MMP-13的表达大大加大，其中一种改性的胆固醇分子，即CLS-MMP特异性断裂多肽-SH，MMP酶特异性断裂多肽使随着环境中MMP的增加而断裂，使缓释体系在到达骨关节炎症部位时，能在患区“精准释放”外泌体。最后，其余与材料直接结合的外泌体能随着材料的降解随之释放，起到“长效缓释”的作用。
附图说明：
图1为本发明所述复合材料的合成示意图。
图2为本发明所述复合材料的分级释放示意图。
图3为实施例2中所述稳定性试验结果。
图4为实施例3中所述分级释放试验结果。
具体实施方式：
为了使本发明的目的、技术方案及优点更加清楚明白，下面结合具体实施方式对本发明作进一步的详细描述，但不应将此理解为本发明上述主题的范围仅限于下述实施例。
实施例1
(1)制备甲基丙烯酸酐改性的透明质酸：
将25g透明质酸(HA)粉末溶入125mL超纯水中，搅拌均匀后调至pH＝8.5，滴加5.625mL的甲基丙烯酸酐(MA)，保持pH冰浴下反应4h，随后室温搅拌过夜。透析后冷冻干燥得到海绵状固体的甲基丙烯酸改性的透明质酸。
(2)制备胆固醇高分子链
设计合成CLS-RVGLP-SH的胆固醇修饰的多肽。合成多肽时，两端分别接C(半胱氨酸)，K(赖氨酸)序列；利用赖氨酸侧链的氨基与胆固醇酯琥珀酸单酯通过酰胺反应接上胆固醇基团，即形成SH-RVGLP-CLS(巯基-MMP响应性断裂多肽-胆固醇)
(3)乳化法制备缓释微凝胶
将SH-RVGLP-CLS用1mM的TCEP处理后，加入过量外泌体在4℃下共混过夜(部分外泌体可以在TCEP的作用下暴露巯基)，以质量比1：50(与HA-MA相比)的比例与10mg(-20mg/mL)的HA-MA混合(调pH至中性)。司班80混于液态石蜡中搅拌均匀，将混合的溶液滴加至油相中，改变转速得到不同粒径，经紫外光照后交联形成的载外泌体的微凝胶体系。
实施例2稳定性试验
取实施例1制备的微凝胶材料，在37摄氏度条件下分别置于5U/mL的透明质酸酶及PBS溶液中，隔一段时间测微凝胶体积变化。
结果如图3所示显示：在透明质酸酶的作用下降解较快，3天左右基本降解。PBS溶液中微凝胶发生溶胀。
实施例3分级释放试验
取实施例1制备的微凝胶材料，通过NTA法分别在PBS、MMP溶液下、及HA酶溶液的作用下探究外泌体的释放情况。
结果如图4所示：多级缓释体系在MMP处理下先响应性释放了一部分外泌体随后缓释速率平缓逐步释放。在HA酶中随材料降解，外泌体释放较快，PBS中外泌体释放较慢。
最后说明的是，以上实施例仅用以说明本发明的技术方案而非限制，尽管参照较佳实施例对本发明进行了详细说明，本领域的普通技术人员应当理解，可以对本发明的技术方案进行修改或者等同替换，而不脱离本发明技术方案的宗旨和范围，其均应涵盖在本发明的权利要求范围当中。
Claims (10)
1.一种环境敏感连接分子，其特征在于，一端有胆固醇，另一端修饰巯基。
2.根据权利要求1所述的环境敏感连接分子，其特征在于，所述巯基与胆固醇之间还连接有环境敏感键。
3.根据权利要求2所述的环境敏感连接分子，其特征在于，所述环境敏感键为pH响应性断裂的键和紫外照射断裂的磷酸酯键，以及白介素家族蛋白(IL)、基质金属蛋白酶家族蛋白(MMP)、血小板反应蛋白解整合素金属肽酶家族蛋白(ADAMTS)、肿瘤坏死因子家族蛋白(TNF)中的至少一种。
4.一种多级环境响应性控释外泌体复合材料，其特征在于，包括甲基丙烯酸酐改性的透明质酸、权利要求1所述的环境敏感连接分子和外泌体，所述甲基丙烯酸酐改性的透明质酸中的双键能够在紫外光的激发下与环境敏感连接分子端部的巯基通过点击反应相连接，所述环境敏感连接分子端部的胆固醇能够插入到所述外泌体的磷脂分子层，从而使外泌体与巯基化透明质酸通过环境敏感连接分子形成交联稳定的结构。
5.根据权利要求4所述的多级环境响应性控释外泌体复合材料，其特征在于，所述环境敏感连接分子包括CLS-MMP响应性断裂多肽(RVGLP)-SH。
6.根据权利要求4所述的多级环境响应性控释外泌体复合材料，其特征在于，部分所述外泌体表面暴露有巯基，能够在紫外光的激发下与甲基丙烯酸中的双键发生点击反应，从而直接与材料结合，这部分外泌体能够随材料降解释放，所述的外泌体也有部分直接与材料共混，能够通过扩散作用释放。
7.根据权利要求4所述的多级环境响应性控释外泌体复合材料，其特征在于，所述复合材料为可注射凝胶、微凝胶或支架材料。
8.一种如权利要求4所述的多级环境响应性控释外泌体复合材料的制备方法，其特征在于，包括以下步骤：
(1)制备甲基丙烯酸酐改性的透明质酸；
(2)制备环境敏感连接分子；
(3)制备响应性控释微凝胶体系。
9.一种如权利要求1所述的环境敏感连接分子的应用，其特征在于，将其用于制备骨关节炎治疗药物或材料。
10.一种如权利要求4所述的多级环境响应性控释外泌体复合材料的应用，其特征在于，将其用于制备骨关节炎治疗药物或材料。
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211649228.4A CN116196432A (zh) | 2022-12-21 | 2022-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
| CN202311777828.3A CN117752804A (zh) | 2022-12-21 | 2023-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211649228.4A CN116196432A (zh) | 2022-12-21 | 2022-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116196432A true CN116196432A (zh) | 2023-06-02 |
Family
ID=86510353
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211649228.4A Pending CN116196432A (zh) | 2022-12-21 | 2022-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
| CN202311777828.3A Pending CN117752804A (zh) | 2022-12-21 | 2023-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311777828.3A Pending CN117752804A (zh) | 2022-12-21 | 2023-12-21 | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN116196432A (zh) |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102266288A (zh) * | 2011-07-14 | 2011-12-07 | 四川大学 | 一种基于胆固醇修饰的还原敏感性肿瘤靶向脂质体 |
| CN103110954A (zh) * | 2013-01-31 | 2013-05-22 | 北京大学 | 胆固醇修饰的生物可降解聚阳离子载体及制备方法和用途 |
| CN104892962A (zh) * | 2015-06-05 | 2015-09-09 | 四川大学 | 一种巯基/二硫键可控自交联透明质酸水凝胶的制备方法及其应用 |
| CN107596379A (zh) * | 2017-10-25 | 2018-01-19 | 武汉大学 | 一种mmp‑8响应性智能牙周药物控释水凝胶材料及其用途 |
| WO2019169380A1 (en) * | 2018-03-02 | 2019-09-06 | The Regents Of The University Of California | Stem cell-derived exosomes for the treatment of corneal scarring |
| KR102053065B1 (ko) * | 2018-06-27 | 2019-12-06 | 가톨릭대학교 산학협력단 | 히알루론산 및 독소루비신을 이용한 pH 감응성 항암 엑소좀 조성물 |
| WO2020198888A1 (en) * | 2019-04-02 | 2020-10-08 | Volumina Medical Sa | Composition comprising a cross-linked polyol |
| KR20200145115A (ko) * | 2019-06-20 | 2020-12-30 | 영남대학교 산학협력단 | 양친매성 독소루비신 및 엑소좀을 이용한 항암 하이브리드 엑소좀 조성물 |
-
2022
- 2022-12-21 CN CN202211649228.4A patent/CN116196432A/zh active Pending
-
2023
- 2023-12-21 CN CN202311777828.3A patent/CN117752804A/zh active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102266288A (zh) * | 2011-07-14 | 2011-12-07 | 四川大学 | 一种基于胆固醇修饰的还原敏感性肿瘤靶向脂质体 |
| CN103110954A (zh) * | 2013-01-31 | 2013-05-22 | 北京大学 | 胆固醇修饰的生物可降解聚阳离子载体及制备方法和用途 |
| CN104892962A (zh) * | 2015-06-05 | 2015-09-09 | 四川大学 | 一种巯基/二硫键可控自交联透明质酸水凝胶的制备方法及其应用 |
| CN107596379A (zh) * | 2017-10-25 | 2018-01-19 | 武汉大学 | 一种mmp‑8响应性智能牙周药物控释水凝胶材料及其用途 |
| WO2019169380A1 (en) * | 2018-03-02 | 2019-09-06 | The Regents Of The University Of California | Stem cell-derived exosomes for the treatment of corneal scarring |
| KR102053065B1 (ko) * | 2018-06-27 | 2019-12-06 | 가톨릭대학교 산학협력단 | 히알루론산 및 독소루비신을 이용한 pH 감응성 항암 엑소좀 조성물 |
| WO2020198888A1 (en) * | 2019-04-02 | 2020-10-08 | Volumina Medical Sa | Composition comprising a cross-linked polyol |
| KR20200145115A (ko) * | 2019-06-20 | 2020-12-30 | 영남대학교 산학협력단 | 양친매성 독소루비신 및 엑소좀을 이용한 항암 하이브리드 엑소좀 조성물 |
Non-Patent Citations (6)
Also Published As
| Publication number | Publication date |
|---|---|
| CN117752804A (zh) | 2024-03-26 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Deming | Functional modification of thioether groups in peptides, polypeptides, and proteins | |
| Caplan et al. | Control of self-assembling oligopeptide matrix formation through systematic variation of amino acid sequence | |
| Moatsou et al. | Self-assembly of temperature-responsive protein–polymer bioconjugates | |
| Kopeček | Smart and genetically engineered biomaterials and drug delivery systems | |
| US7417021B2 (en) | Biologically active conjugate of a biopolymer and therapeutic agent | |
| US8034618B2 (en) | PTH containing cell growth matrix | |
| US8524885B2 (en) | Hydrazido derivatives of hyaluronic acid | |
| JP2009102383A (ja) | 組織操作のための成長因子改変タンパク質マトリクス | |
| JP2002535108A (ja) | 共役不飽和基に対する求核付加反応によって作製される生体適合材料 | |
| EP0449592B1 (en) | Bioelastomeric drug delivery system | |
| US20140038826A1 (en) | Covalently cross linked hydrogels and methods of making and using same | |
| JP2008529972A (ja) | 酵素的に分解可能な連結を通じて生物活性因子が組み入れられている合成バイオマテリアル | |
| JP2007182407A (ja) | 徐放性ハイドロゲル製剤 | |
| WO2003087197A1 (en) | Hydrogel with controllable mechanical chemical and biological properties and method for making same | |
| WO2008083542A1 (fr) | Dérivés de gélatine multiples modifiés et leur matériau de réticulation | |
| JP2003522806A (ja) | 改良された薬物送達のための生体高分子の修飾 | |
| Gharios et al. | Chemical and biological engineering strategies to make and modify next-generation hydrogel biomaterials | |
| CN116196432A (zh) | 一种多级环境响应性控释外泌体复合材料及其制备方法与应用 | |
| Li et al. | Recent Advances in Post‐polymerization Modifications on Polypeptides: Synthesis and Applications | |
| EP0302284B1 (en) | Stabilized enzymes | |
| Fox et al. | Development and characterization of calmodulin-based copolymeric hydrogels | |
| US20030224052A1 (en) | Hydrogel with controllable merchanical, chemical, and biological properties and method for making same | |
| CN114891239B (zh) | 一种硼氮内配位硼酸酯水凝胶及其制备方法 | |
| CN115073552B (zh) | 一种多肽、水凝胶及其用途 | |
| Benavides et al. | Controlled Synthesis and Properties of Poly (l-homoserine) |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20230602 |