CN103751795B - Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition - Google Patents
Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition Download PDFInfo
- Publication number
- CN103751795B CN103751795B CN201310186800.2A CN201310186800A CN103751795B CN 103751795 B CN103751795 B CN 103751795B CN 201310186800 A CN201310186800 A CN 201310186800A CN 103751795 B CN103751795 B CN 103751795B
- Authority
- CN
- China
- Prior art keywords
- hyaluronic acid
- acid
- conjugate
- dissolved
- granule
- 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.)
- Expired - Fee Related
Links
- 239000002246 antineoplastic agent Substances 0.000 title claims abstract description 62
- 239000002131 composite material Substances 0.000 title claims abstract description 47
- 238000002360 preparation method Methods 0.000 title claims abstract description 29
- 229940041181 antineoplastic drug Drugs 0.000 title claims abstract description 25
- 239000002105 nanoparticle Substances 0.000 title claims abstract description 16
- 239000000203 mixture Substances 0.000 title claims abstract description 9
- 229920002674 hyaluronan Polymers 0.000 claims abstract description 72
- 229960003160 hyaluronic acid Drugs 0.000 claims abstract description 68
- 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 claims abstract description 59
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 31
- 239000003814 drug Substances 0.000 claims abstract description 26
- 230000000694 effects Effects 0.000 claims abstract description 22
- 230000000259 anti-tumor effect Effects 0.000 claims abstract description 20
- 238000002347 injection Methods 0.000 claims abstract description 15
- 239000007924 injection Substances 0.000 claims abstract description 15
- 229920000728 polyester Polymers 0.000 claims abstract description 11
- 229920001400 block copolymer Polymers 0.000 claims abstract description 9
- 150000003863 ammonium salts Chemical class 0.000 claims abstract description 8
- 238000000034 method Methods 0.000 claims abstract description 8
- 230000008569 process Effects 0.000 claims abstract description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 53
- 239000012467 final product Substances 0.000 claims description 40
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 39
- 239000008187 granular material Substances 0.000 claims description 38
- 229940034982 antineoplastic agent Drugs 0.000 claims description 37
- 229960001592 paclitaxel Drugs 0.000 claims description 31
- 238000004108 freeze drying Methods 0.000 claims description 29
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 26
- 239000000243 solution Substances 0.000 claims description 23
- 239000003960 organic solvent Substances 0.000 claims description 22
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 21
- 229930012538 Paclitaxel Natural products 0.000 claims description 21
- -1 hyaluronic acid ammonium salt Chemical class 0.000 claims description 21
- VHYFNPMBLIVWCW-UHFFFAOYSA-N 4-Dimethylaminopyridine Chemical group CN(C)C1=CC=NC=C1 VHYFNPMBLIVWCW-UHFFFAOYSA-N 0.000 claims description 20
- 238000000502 dialysis Methods 0.000 claims description 20
- 239000002253 acid Substances 0.000 claims description 19
- 239000008367 deionised water Substances 0.000 claims description 19
- 229910021641 deionized water Inorganic materials 0.000 claims description 19
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 18
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 18
- RCINICONZNJXQF-MZXODVADSA-N taxol Chemical compound O([C@@H]1[C@@]2(C[C@@H](C(C)=C(C2(C)C)[C@H](C([C@]2(C)[C@@H](O)C[C@H]3OC[C@]3([C@H]21)OC(C)=O)=O)OC(=O)C)OC(=O)[C@H](O)[C@@H](NC(=O)C=1C=CC=CC=1)C=1C=CC=CC=1)O)C(=O)C1=CC=CC=C1 RCINICONZNJXQF-MZXODVADSA-N 0.000 claims description 17
- 239000012190 activator Substances 0.000 claims description 16
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 15
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 15
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 15
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 14
- 239000013067 intermediate product Substances 0.000 claims description 14
- 239000003054 catalyst Substances 0.000 claims description 13
- YWIVKILSMZOHHF-QJZPQSOGSA-N sodium;(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- Chemical compound [Na+].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 YWIVKILSMZOHHF-QJZPQSOGSA-N 0.000 claims description 13
- 238000013019 agitation Methods 0.000 claims description 12
- 239000000523 sample Substances 0.000 claims description 12
- 229940024606 amino acid Drugs 0.000 claims description 11
- 235000001014 amino acid Nutrition 0.000 claims description 11
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 11
- 238000001291 vacuum drying Methods 0.000 claims description 11
- 229960000549 4-dimethylaminophenol Drugs 0.000 claims description 10
- 150000002148 esters Chemical class 0.000 claims description 9
- 238000000746 purification Methods 0.000 claims description 9
- GVJRTUUUJYMTNQ-UHFFFAOYSA-N 2-(2,5-dioxofuran-3-yl)acetic acid Chemical compound OC(=O)CC1=CC(=O)OC1=O GVJRTUUUJYMTNQ-UHFFFAOYSA-N 0.000 claims description 8
- 229930195573 Amycin Natural products 0.000 claims description 8
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 claims description 8
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 8
- VSJKWCGYPAHWDS-FQEVSTJZSA-N camptothecin Chemical compound C1=CC=C2C=C(CN3C4=CC5=C(C3=O)COC(=O)[C@]5(O)CC)C4=NC2=C1 VSJKWCGYPAHWDS-FQEVSTJZSA-N 0.000 claims description 8
- 229960003668 docetaxel Drugs 0.000 claims description 8
- 229920000642 polymer Polymers 0.000 claims description 8
- 150000008064 anhydrides Chemical class 0.000 claims description 7
- 229920001577 copolymer Polymers 0.000 claims description 7
- 239000000047 product Substances 0.000 claims description 7
- 239000000376 reactant Substances 0.000 claims description 7
- 239000002904 solvent Substances 0.000 claims description 7
- VDZOOKBUILJEDG-UHFFFAOYSA-M tetrabutylammonium hydroxide Chemical compound [OH-].CCCC[N+](CCCC)(CCCC)CCCC VDZOOKBUILJEDG-UHFFFAOYSA-M 0.000 claims description 7
- NWXMGUDVXFXRIG-WESIUVDSSA-N (4s,4as,5as,6s,12ar)-4-(dimethylamino)-1,6,10,11,12a-pentahydroxy-6-methyl-3,12-dioxo-4,4a,5,5a-tetrahydrotetracene-2-carboxamide Chemical compound C1=CC=C2[C@](O)(C)[C@H]3C[C@H]4[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]4(O)C(=O)C3=C(O)C2=C1O NWXMGUDVXFXRIG-WESIUVDSSA-N 0.000 claims description 6
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 6
- ZDZOTLJHXYCWBA-VCVYQWHSSA-N N-debenzoyl-N-(tert-butoxycarbonyl)-10-deacetyltaxol Chemical compound O([C@H]1[C@H]2[C@@](C([C@H](O)C3=C(C)[C@@H](OC(=O)[C@H](O)[C@@H](NC(=O)OC(C)(C)C)C=4C=CC=CC=4)C[C@]1(O)C3(C)C)=O)(C)[C@@H](O)C[C@H]1OC[C@]12OC(=O)C)C(=O)C1=CC=CC=C1 ZDZOTLJHXYCWBA-VCVYQWHSSA-N 0.000 claims description 6
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 6
- 150000001413 amino acids Chemical class 0.000 claims description 6
- 230000008901 benefit Effects 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- 239000012535 impurity Substances 0.000 claims description 6
- 150000003053 piperidines Chemical class 0.000 claims description 6
- IBVAQQYNSHJXBV-UHFFFAOYSA-N adipic acid dihydrazide Chemical compound NNC(=O)CCCCC(=O)NN IBVAQQYNSHJXBV-UHFFFAOYSA-N 0.000 claims description 5
- 238000004440 column chromatography Methods 0.000 claims description 5
- 238000004132 cross linking Methods 0.000 claims description 5
- 238000002390 rotary evaporation Methods 0.000 claims description 5
- 229920002307 Dextran Polymers 0.000 claims description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 4
- 239000002585 base Substances 0.000 claims description 4
- 150000001718 carbodiimides Chemical class 0.000 claims description 4
- 230000007935 neutral effect Effects 0.000 claims description 4
- RINCXYDBBGOEEQ-UHFFFAOYSA-N succinic anhydride Chemical class O=C1CCC(=O)O1 RINCXYDBBGOEEQ-UHFFFAOYSA-N 0.000 claims description 4
- 229920000428 triblock copolymer Polymers 0.000 claims description 4
- QOSSAOTZNIDXMA-UHFFFAOYSA-N Dicylcohexylcarbodiimide Chemical compound C1CCCCC1N=C=NC1CCCCC1 QOSSAOTZNIDXMA-UHFFFAOYSA-N 0.000 claims description 3
- ZHNUHDYFZUAESO-UHFFFAOYSA-N Formamide Chemical group NC=O ZHNUHDYFZUAESO-UHFFFAOYSA-N 0.000 claims description 3
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 claims description 3
- 206010061535 Ovarian neoplasm Diseases 0.000 claims description 3
- 239000006185 dispersion Substances 0.000 claims description 3
- 229930182817 methionine Natural products 0.000 claims description 3
- 208000002154 non-small cell lung carcinoma Diseases 0.000 claims description 3
- 229920000382 poly(ethylene glycol) methyl ether-block-poly(L-lactide-co-glycolide) Polymers 0.000 claims description 3
- 239000002244 precipitate Substances 0.000 claims description 3
- 239000012264 purified product Substances 0.000 claims description 3
- 238000006467 substitution reaction Methods 0.000 claims description 3
- 239000000725 suspension Substances 0.000 claims description 3
- 208000029729 tumor suppressor gene on chromosome 11 Diseases 0.000 claims description 3
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims description 2
- 108010082627 (ethylene glycol)-poly(lactide)-poly(3-methyl-morpholine-2,5-dione) Proteins 0.000 claims description 2
- QDGAVODICPCDMU-UHFFFAOYSA-N 2-amino-3-[3-[bis(2-chloroethyl)amino]phenyl]propanoic acid Chemical compound OC(=O)C(N)CC1=CC=CC(N(CCCl)CCCl)=C1 QDGAVODICPCDMU-UHFFFAOYSA-N 0.000 claims description 2
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 claims description 2
- 206010006187 Breast cancer Diseases 0.000 claims description 2
- 208000026310 Breast neoplasm Diseases 0.000 claims description 2
- 206010008342 Cervix carcinoma Diseases 0.000 claims description 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 claims description 2
- QNAYBMKLOCPYGJ-UHFFFAOYSA-N D-alpha-Ala Natural products CC([NH3+])C([O-])=O QNAYBMKLOCPYGJ-UHFFFAOYSA-N 0.000 claims description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 2
- QNAYBMKLOCPYGJ-UWTATZPHSA-N L-Alanine Natural products C[C@@H](N)C(O)=O QNAYBMKLOCPYGJ-UWTATZPHSA-N 0.000 claims description 2
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 claims description 2
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 claims description 2
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 claims description 2
- 229930182844 L-isoleucine Natural products 0.000 claims description 2
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 claims description 2
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 claims description 2
- 229930195725 Mannitol Natural products 0.000 claims description 2
- 206010033128 Ovarian cancer Diseases 0.000 claims description 2
- DYUQAZSOFZSPHD-UHFFFAOYSA-N Phenylpropyl alcohol Natural products CCC(O)C1=CC=CC=C1 DYUQAZSOFZSPHD-UHFFFAOYSA-N 0.000 claims description 2
- 229920000436 Poly(lactide-co-glycolide)-block-poly(ethylene glycol)-block-poly(lactide-co-glycolide) Polymers 0.000 claims description 2
- 229920000432 Polylactide-block-poly(ethylene glycol)-block-polylactide Polymers 0.000 claims description 2
- 238000006788 Steglich reaction Methods 0.000 claims description 2
- 208000005718 Stomach Neoplasms Diseases 0.000 claims description 2
- 229930006000 Sucrose Natural products 0.000 claims description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 2
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 claims description 2
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims description 2
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 claims description 2
- 229960003767 alanine Drugs 0.000 claims description 2
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 claims description 2
- 201000008275 breast carcinoma Diseases 0.000 claims description 2
- 239000002775 capsule Substances 0.000 claims description 2
- 150000001732 carboxylic acid derivatives Chemical class 0.000 claims description 2
- 201000010881 cervical cancer Diseases 0.000 claims description 2
- 230000018044 dehydration Effects 0.000 claims description 2
- 238000006297 dehydration reaction Methods 0.000 claims description 2
- 239000002552 dosage form Substances 0.000 claims description 2
- 238000001647 drug administration Methods 0.000 claims description 2
- 239000000890 drug combination Substances 0.000 claims description 2
- IRXSLJNXXZKURP-UHFFFAOYSA-N fluorenylmethyloxycarbonyl chloride Chemical compound C1=CC=C2C(COC(=O)Cl)C3=CC=CC=C3C2=C1 IRXSLJNXXZKURP-UHFFFAOYSA-N 0.000 claims description 2
- 206010017758 gastric cancer Diseases 0.000 claims description 2
- 239000008103 glucose Substances 0.000 claims description 2
- 229960002989 glutamic acid Drugs 0.000 claims description 2
- KIUKXJAPPMFGSW-MNSSHETKSA-N hyaluronan Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)C1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H](C(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-MNSSHETKSA-N 0.000 claims description 2
- 229940099552 hyaluronan Drugs 0.000 claims description 2
- 229960000310 isoleucine Drugs 0.000 claims description 2
- 239000008101 lactose Substances 0.000 claims description 2
- 239000008176 lyophilized powder Substances 0.000 claims description 2
- 239000000594 mannitol Substances 0.000 claims description 2
- 235000010355 mannitol Nutrition 0.000 claims description 2
- 229960004452 methionine Drugs 0.000 claims description 2
- 239000012046 mixed solvent Substances 0.000 claims description 2
- 230000000144 pharmacologic effect Effects 0.000 claims description 2
- 229920000762 poly(caprolactone)-poly(ethylene glycol)-poly(caprolactone) Polymers 0.000 claims description 2
- 229940098458 powder spray Drugs 0.000 claims description 2
- 201000011549 stomach cancer Diseases 0.000 claims description 2
- 239000005720 sucrose Substances 0.000 claims description 2
- 239000003826 tablet Substances 0.000 claims description 2
- FDKXTQMXEQVLRF-ZHACJKMWSA-N (E)-dacarbazine Chemical group CN(C)\N=N\c1[nH]cnc1C(N)=O FDKXTQMXEQVLRF-ZHACJKMWSA-N 0.000 claims 1
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 claims 1
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 claims 1
- 108010020346 Polyglutamic Acid Proteins 0.000 claims 1
- 108010039918 Polylysine Proteins 0.000 claims 1
- 125000005211 alkyl trimethyl ammonium group Chemical group 0.000 claims 1
- 238000004458 analytical method Methods 0.000 claims 1
- 238000006555 catalytic reaction Methods 0.000 claims 1
- 201000007270 liver cancer Diseases 0.000 claims 1
- 208000014018 liver neoplasm Diseases 0.000 claims 1
- 210000004877 mucosa Anatomy 0.000 claims 1
- 108010054442 polyalanine Proteins 0.000 claims 1
- 229920002643 polyglutamic acid Polymers 0.000 claims 1
- 108010094020 polyglycine Proteins 0.000 claims 1
- 229920000232 polyglycine polymer Polymers 0.000 claims 1
- 229920000656 polylysine Polymers 0.000 claims 1
- 238000009738 saturating Methods 0.000 claims 1
- 230000008685 targeting Effects 0.000 abstract description 11
- 229940079593 drug Drugs 0.000 abstract description 10
- 238000001727 in vivo Methods 0.000 abstract description 4
- 239000003446 ligand Substances 0.000 abstract description 4
- 238000005516 engineering process Methods 0.000 abstract description 3
- 238000011068 loading method Methods 0.000 abstract description 3
- 125000006850 spacer group Chemical group 0.000 abstract description 3
- 230000009467 reduction Effects 0.000 abstract description 2
- 230000001268 conjugating effect Effects 0.000 abstract 2
- 238000009776 industrial production Methods 0.000 abstract 1
- 238000001308 synthesis method Methods 0.000 abstract 1
- 239000000562 conjugate Substances 0.000 description 57
- 238000003756 stirring Methods 0.000 description 19
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 18
- 210000004027 cell Anatomy 0.000 description 15
- 238000005160 1H NMR spectroscopy Methods 0.000 description 12
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 12
- 229940090044 injection Drugs 0.000 description 12
- FPQQSJJWHUJYPU-UHFFFAOYSA-N 3-(dimethylamino)propyliminomethylidene-ethylazanium;chloride Chemical compound Cl.CCN=C=NCCCN(C)C FPQQSJJWHUJYPU-UHFFFAOYSA-N 0.000 description 11
- DCPMPXBYPZGNDC-UHFFFAOYSA-N hydron;methanediimine;chloride Chemical compound Cl.N=C=N DCPMPXBYPZGNDC-UHFFFAOYSA-N 0.000 description 11
- HOGDNTQCSIKEEV-UHFFFAOYSA-N n'-hydroxybutanediamide Chemical compound NC(=O)CCC(=O)NO HOGDNTQCSIKEEV-UHFFFAOYSA-N 0.000 description 10
- 238000000425 proton nuclear magnetic resonance spectrum Methods 0.000 description 10
- 238000012377 drug delivery Methods 0.000 description 9
- 239000008213 purified water Substances 0.000 description 9
- UIIMBOGNXHQVGW-UHFFFAOYSA-M sodium bicarbonate Substances [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 8
- 229940127093 camptothecin Drugs 0.000 description 7
- 239000002245 particle Substances 0.000 description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 6
- 231100000135 cytotoxicity Toxicity 0.000 description 6
- 230000003013 cytotoxicity Effects 0.000 description 6
- 229960002918 doxorubicin hydrochloride Drugs 0.000 description 6
- 210000000865 mononuclear phagocyte system Anatomy 0.000 description 6
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 5
- MWWSFMDVAYGXBV-RUELKSSGSA-N Doxorubicin hydrochloride Chemical compound Cl.O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 MWWSFMDVAYGXBV-RUELKSSGSA-N 0.000 description 5
- 230000009471 action Effects 0.000 description 5
- 201000011510 cancer Diseases 0.000 description 5
- 235000013339 cereals Nutrition 0.000 description 5
- 230000003285 pharmacodynamic effect Effects 0.000 description 5
- 229920001223 polyethylene glycol Polymers 0.000 description 5
- 231100000331 toxic Toxicity 0.000 description 5
- 230000002588 toxic effect Effects 0.000 description 5
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 4
- DXRFZHILMCWCNG-UHFFFAOYSA-N N,N-dimethyl-1,8-naphthyridin-2-amine Chemical compound C1=CC=NC2=NC(N(C)C)=CC=C21 DXRFZHILMCWCNG-UHFFFAOYSA-N 0.000 description 4
- 230000004913 activation Effects 0.000 description 4
- 239000003005 anticarcinogenic agent Substances 0.000 description 4
- 230000000857 drug effect Effects 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 238000000338 in vitro Methods 0.000 description 4
- 239000010410 layer Substances 0.000 description 4
- 230000004048 modification Effects 0.000 description 4
- 238000012986 modification Methods 0.000 description 4
- 229920001610 polycaprolactone Polymers 0.000 description 4
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 4
- 210000001519 tissue Anatomy 0.000 description 4
- 231100000419 toxicity Toxicity 0.000 description 4
- 230000001988 toxicity Effects 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 108010003272 Hyaluronate lyase Proteins 0.000 description 3
- 102000001974 Hyaluronidases Human genes 0.000 description 3
- 241000699670 Mus sp. Species 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 229910021529 ammonia Inorganic materials 0.000 description 3
- 239000000460 chlorine Substances 0.000 description 3
- 229910052801 chlorine Inorganic materials 0.000 description 3
- 239000012043 crude product Substances 0.000 description 3
- 238000010586 diagram Methods 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 229960002773 hyaluronidase Drugs 0.000 description 3
- 229920002521 macromolecule Polymers 0.000 description 3
- 239000002609 medium Substances 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 239000004626 polylactic acid Substances 0.000 description 3
- 238000001556 precipitation Methods 0.000 description 3
- 238000001953 recrystallisation Methods 0.000 description 3
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 description 2
- VHSHLMUCYSAUQU-UHFFFAOYSA-N 2-hydroxypropyl methacrylate Chemical compound CC(O)COC(=O)C(C)=C VHSHLMUCYSAUQU-UHFFFAOYSA-N 0.000 description 2
- RGUIGXFOHAOFLG-UHFFFAOYSA-N 2-n,3-n-dimethylpyridine-2,3-diamine Chemical compound CNC1=CC=CN=C1NC RGUIGXFOHAOFLG-UHFFFAOYSA-N 0.000 description 2
- KLWPJMFMVPTNCC-UHFFFAOYSA-N Camptothecin Natural products CCC1(O)C(=O)OCC2=C1C=C3C4Nc5ccccc5C=C4CN3C2=O KLWPJMFMVPTNCC-UHFFFAOYSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 101000868273 Homo sapiens CD44 antigen Proteins 0.000 description 2
- 241000699666 Mus <mouse, genus> Species 0.000 description 2
- 241000209094 Oryza Species 0.000 description 2
- 235000007164 Oryza sativa Nutrition 0.000 description 2
- 206010057249 Phagocytosis Diseases 0.000 description 2
- 239000002202 Polyethylene glycol Substances 0.000 description 2
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 239000003729 cation exchange resin Substances 0.000 description 2
- 238000012412 chemical coupling Methods 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 239000012141 concentrate Substances 0.000 description 2
- 230000008878 coupling Effects 0.000 description 2
- 238000010168 coupling process Methods 0.000 description 2
- 238000005859 coupling reaction Methods 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- VSJKWCGYPAHWDS-UHFFFAOYSA-N dl-camptothecin Natural products C1=CC=C2C=C(CN3C4=CC5=C(C3=O)COC(=O)C5(O)CC)C4=NC2=C1 VSJKWCGYPAHWDS-UHFFFAOYSA-N 0.000 description 2
- 229960004756 ethanol Drugs 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 230000029142 excretion Effects 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- OVBPIULPVIDEAO-LBPRGKRZSA-N folic acid Chemical compound C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-LBPRGKRZSA-N 0.000 description 2
- 230000036571 hydration Effects 0.000 description 2
- 238000006703 hydration reaction Methods 0.000 description 2
- 230000007062 hydrolysis Effects 0.000 description 2
- 238000006460 hydrolysis reaction Methods 0.000 description 2
- 230000002209 hydrophobic effect Effects 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 239000000693 micelle Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 2
- 239000012044 organic layer Substances 0.000 description 2
- 230000008782 phagocytosis Effects 0.000 description 2
- 229920000747 poly(lactic acid) Polymers 0.000 description 2
- 239000004632 polycaprolactone Substances 0.000 description 2
- 239000000651 prodrug Substances 0.000 description 2
- 229940002612 prodrug Drugs 0.000 description 2
- 235000009566 rice Nutrition 0.000 description 2
- 239000000741 silica gel Substances 0.000 description 2
- 229910002027 silica gel Inorganic materials 0.000 description 2
- 229960001866 silicon dioxide Drugs 0.000 description 2
- RWRDLPDLKQPQOW-UHFFFAOYSA-N tetrahydropyrrole Natural products C1CCNC1 RWRDLPDLKQPQOW-UHFFFAOYSA-N 0.000 description 2
- 229960004295 valine Drugs 0.000 description 2
- 210000003462 vein Anatomy 0.000 description 2
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide Chemical class CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 description 1
- HAWSQZCWOQZXHI-FQEVSTJZSA-N 10-Hydroxycamptothecin Chemical compound C1=C(O)C=C2C=C(CN3C4=CC5=C(C3=O)COC(=O)[C@]5(O)CC)C4=NC2=C1 HAWSQZCWOQZXHI-FQEVSTJZSA-N 0.000 description 1
- ZFFMLCVRJBZUDZ-UHFFFAOYSA-N 2,3-dimethylbutane Chemical group CC(C)C(C)C ZFFMLCVRJBZUDZ-UHFFFAOYSA-N 0.000 description 1
- XHEYLBRFUKIQMM-UHFFFAOYSA-N 2-aminoethanol;dichloromethane Chemical compound ClCCl.NCCO XHEYLBRFUKIQMM-UHFFFAOYSA-N 0.000 description 1
- AOJJSUZBOXZQNB-VTZDEGQISA-N 4'-epidoxorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@@H](O)[C@H](C)O1 AOJJSUZBOXZQNB-VTZDEGQISA-N 0.000 description 1
- STQGQHZAVUOBTE-UHFFFAOYSA-N 7-Cyan-hept-2t-en-4,6-diinsaeure Natural products C1=2C(O)=C3C(=O)C=4C(OC)=CC=CC=4C(=O)C3=C(O)C=2CC(O)(C(C)=O)CC1OC1CC(N)C(O)C(C)O1 STQGQHZAVUOBTE-UHFFFAOYSA-N 0.000 description 1
- 241000227129 Aconitum Species 0.000 description 1
- 206010000830 Acute leukaemia Diseases 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 108700031361 Brachyury Proteins 0.000 description 1
- 102100032912 CD44 antigen Human genes 0.000 description 1
- 108010001857 Cell Surface Receptors Proteins 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- HTIJFSOGRVMCQR-UHFFFAOYSA-N Epirubicin Natural products COc1cccc2C(=O)c3c(O)c4CC(O)(CC(OC5CC(N)C(=O)C(C)O5)c4c(O)c3C(=O)c12)C(=O)CO HTIJFSOGRVMCQR-UHFFFAOYSA-N 0.000 description 1
- 108090000371 Esterases Proteins 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-Valine Natural products CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 description 1
- 125000003338 L-glutaminyl group Chemical group O=C([*])[C@](N([H])[H])([H])C([H])([H])C([H])([H])C(=O)N([H])[H] 0.000 description 1
- FBOZXECLQNJBKD-ZDUSSCGKSA-N L-methotrexate Chemical compound C=1N=C2N=C(N)N=C(N)C2=NC=1CN(C)C1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 FBOZXECLQNJBKD-ZDUSSCGKSA-N 0.000 description 1
- 125000003580 L-valyl group Chemical group [H]N([H])[C@]([H])(C(=O)[*])C(C([H])([H])[H])(C([H])([H])[H])[H] 0.000 description 1
- ROHFNLRQFUQHCH-UHFFFAOYSA-N Leucine Natural products CC(C)CC(N)C(O)=O ROHFNLRQFUQHCH-UHFFFAOYSA-N 0.000 description 1
- 206010058467 Lung neoplasm malignant Diseases 0.000 description 1
- 206010025323 Lymphomas Diseases 0.000 description 1
- OVBPIULPVIDEAO-UHFFFAOYSA-N N-Pteroyl-L-glutaminsaeure Natural products C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)NC(CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-UHFFFAOYSA-N 0.000 description 1
- 125000003047 N-acetyl group Chemical group 0.000 description 1
- 206010042971 T-cell lymphoma Diseases 0.000 description 1
- 208000027585 T-cell non-Hodgkin lymphoma Diseases 0.000 description 1
- 239000008186 active pharmaceutical agent Substances 0.000 description 1
- 150000001335 aliphatic alkanes Chemical class 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 235000011114 ammonium hydroxide Nutrition 0.000 description 1
- 230000001093 anti-cancer Effects 0.000 description 1
- 230000000118 anti-neoplastic effect Effects 0.000 description 1
- 239000012736 aqueous medium Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- RMGHCHRPOMLKEU-UHFFFAOYSA-M azanium tetrabutylazanium dihydroxide Chemical compound [NH4+].[OH-].[OH-].CCCC[N+](CCCC)(CCCC)CCCC RMGHCHRPOMLKEU-UHFFFAOYSA-M 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229920000249 biocompatible polymer Polymers 0.000 description 1
- 238000006065 biodegradation reaction Methods 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000012661 block copolymerization Methods 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000004700 cellular uptake Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000007385 chemical modification Methods 0.000 description 1
- 239000012295 chemical reaction liquid Substances 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- 230000004087 circulation Effects 0.000 description 1
- 208000029742 colonic neoplasm Diseases 0.000 description 1
- 238000013270 controlled release Methods 0.000 description 1
- 239000011258 core-shell material Substances 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 229960000975 daunorubicin Drugs 0.000 description 1
- STQGQHZAVUOBTE-VGBVRHCVSA-N daunorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(C)=O)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 STQGQHZAVUOBTE-VGBVRHCVSA-N 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 229960000935 dehydrated alcohol Drugs 0.000 description 1
- 238000002242 deionisation method Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 150000001991 dicarboxylic acids Chemical class 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229960004679 doxorubicin Drugs 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 230000005611 electricity Effects 0.000 description 1
- 230000009881 electrostatic interaction Effects 0.000 description 1
- 230000007515 enzymatic degradation Effects 0.000 description 1
- 229960001904 epirubicin Drugs 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 230000032050 esterification Effects 0.000 description 1
- 238000005886 esterification reaction Methods 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 229960000304 folic acid Drugs 0.000 description 1
- 235000019152 folic acid Nutrition 0.000 description 1
- 239000011724 folic acid Substances 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 125000000524 functional group Chemical group 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 125000003147 glycosyl group Chemical group 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 150000007857 hydrazones Chemical class 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 125000001165 hydrophobic group Chemical group 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 239000003978 infusion fluid Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- UWKQSNNFCGGAFS-XIFFEERXSA-N irinotecan Chemical compound C1=C2C(CC)=C3CN(C(C4=C([C@@](C(=O)OC4)(O)CC)C=4)=O)C=4C3=NC2=CC=C1OC(=O)N(CC1)CCC1N1CCCCC1 UWKQSNNFCGGAFS-XIFFEERXSA-N 0.000 description 1
- 229960004768 irinotecan Drugs 0.000 description 1
- 229950003188 isovaleryl diethylamide Drugs 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 201000011061 large intestine cancer Diseases 0.000 description 1
- 125000001909 leucine group Chemical group [H]N(*)C(C(*)=O)C([H])([H])C(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- 208000032839 leukemia Diseases 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 102000006240 membrane receptors Human genes 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 125000001360 methionine group Chemical group N[C@@H](CCSC)C(=O)* 0.000 description 1
- 229960000485 methotrexate Drugs 0.000 description 1
- OFXSXYCSPVKZPF-UHFFFAOYSA-N methoxyperoxymethane Chemical compound COOOC OFXSXYCSPVKZPF-UHFFFAOYSA-N 0.000 description 1
- 229960001156 mitoxantrone Drugs 0.000 description 1
- KKZJGLLVHKMTCM-UHFFFAOYSA-N mitoxantrone Chemical compound O=C1C2=C(O)C=CC(O)=C2C(=O)C2=C1C(NCCNCCO)=CC=C2NCCNCCO KKZJGLLVHKMTCM-UHFFFAOYSA-N 0.000 description 1
- 230000004899 motility Effects 0.000 description 1
- QKVIAQUPPCPXBX-UHFFFAOYSA-N n,n-dimethyl-1h-pyrrol-2-amine Chemical compound CN(C)C1=CC=CN1 QKVIAQUPPCPXBX-UHFFFAOYSA-N 0.000 description 1
- 238000000655 nuclear magnetic resonance spectrum Methods 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000002018 overexpression Effects 0.000 description 1
- 229940108949 paclitaxel injection Drugs 0.000 description 1
- 230000000242 pagocytic effect Effects 0.000 description 1
- 125000001151 peptidyl group Chemical group 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 210000002381 plasma Anatomy 0.000 description 1
- 239000002861 polymer material Substances 0.000 description 1
- 239000000580 polymer-drug conjugate Substances 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 235000018102 proteins Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 1
- 150000003222 pyridines Chemical class 0.000 description 1
- 230000010837 receptor-mediated endocytosis Effects 0.000 description 1
- 108020003175 receptors Proteins 0.000 description 1
- 102000005962 receptors Human genes 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- DCKVNWZUADLDEH-UHFFFAOYSA-N sec-butyl acetate Chemical group CCC(C)OC(C)=O DCKVNWZUADLDEH-UHFFFAOYSA-N 0.000 description 1
- 238000001338 self-assembly Methods 0.000 description 1
- 238000010898 silica gel chromatography Methods 0.000 description 1
- 229940126586 small molecule drug Drugs 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 235000017557 sodium bicarbonate Nutrition 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000012265 solid product Substances 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 150000003852 triazoles Chemical class 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 229920003169 water-soluble polymer Polymers 0.000 description 1
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicinal Preparation (AREA)
Abstract
The present invention relates to a preparation method and an application of a hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition with characteristics of active targeting antitumor effect and biodegradability. The preparation method is characterized by comprising: (1) a synthesis method for conjugating an antitumor drug and a spacer and conjugating a targeting ligand hyaluronic acid or an ammonium salt thereof and the antitumor drug-spacer; and (2) a new technology for assembling the hyaluronic acid-antitumor drug conjugate and amphiphilic polyester block copolymer composite nanoparticles. The composition has effects of substantially increased drug loading, efficacy improving, in vivo long-circulating effect achievement, active tumor targeting property and drug toxic-side effect reduction. The macromolecular conjugate and complex nanoparticle composition can be used for injection administration, oral administration or mucosal administration. In addition, the preparation method has characteristics of mature process and high yield, and is suitable for industrial production.
Description
Technical field
The present invention relates to chemical field, pharmaceutic adjuvant or field of pharmaceutical preparations.Specifically related to hyaluronic acid-anti-swollen
Tumor medicine conjugate and the preparation method of complex nanometer granule.It is coupled with linking arm including antitumor drug, target ligand is transparent
Matter acid or the preparation method of its ammonium salt and antitumor drug-linking arm；The invention still further relates to having active target head hyaluronic acid-anti-
Tumour medicine conjugate and the assembling new technique of amphipathic ester block copolymer composite nano-granule.
Background technology
Cancer has become one of serious principal disease threatening human life and health.Due to cancer early discovery and diagnosis relatively
For difficulty, chemotherapy clinically plays an important role, but still suffers from various problems and face the challenge, such as antitumor drug
Dissolubility is poor, toxic and side effects are big and the problems such as drug resistance.Rapid with oncology, molecular biology and high molecular polymer is sent out
Exhibition, the targeting conveying administration of antitumor drug becomes study hotspot.Polymer-drug conjugate (Polymer-drug
Delivery system, PDDS) there are many advantages, have evolved into the new field of drug delivery system.
Polymer-antineoplastic agent conjugate is that small-molecule drug is passed through covalent bond and is connected with water-soluble polymer, and medicine is even
Connection thing can significantly improve the dissolubility of insoluble drug, and polymer passes through simple hydrolysis or enzymatic degradation in body circulation, prolongs
Half-life in long internal blood plasma, reaching target site thus playing drug effect, the particularly important is antitumor conjugate and there is EPR effect
Should be able to accumulate in tumor locus.Main feature has：(1) improve the water solublity of insoluble drug；(2) improve medicine tumor-targeting
Prodrug composite nano-granule has nano-scale, is concentrated on tumor tissues by the EPR effect of tumor, has passive targeting；Multiple
The nucleocapsid structure closing nanoparticle is versatile and flexible, can resist as glycosyl (HA), peptidyl, Folic Acid in macromolecule surface connection-part
Body etc., is acted on target cell surface receptors molecular specific by ligand molecular, strengthens its specific recognition to tissue and organ,
Realize active targeting；(3) the stable transhipment realizing medicine avoids the physically trapping nano-micelle system initial stage to dash forward release problem, by medicine
Thing takes target site to and realizes intelligent release, thus reaching the effect strengthening drug effect, lowering toxic and side effects.
It is coupled with PTX using biocompatible polymer material, building targeted drug delivery system (DDS) has become current
Study hotspot.In conjugate, drug molecule is combined with main polymer chain by covalent bond, forms nucleocapsid in an aqueous medium
Type micelle, can increase drug solubility, improve bioavailability, the stability in blood circulation, it is to avoid medicine reaches target portion
By enzyme metabolism, inactivation before position, medicine is transported to greatest extent target site, reduces the toxic and side effects of medicine.
HA is water-soluble natural macromolecule polysaccharide, has multi-functional physical chemistry and biological property, such as bio-compatible
Property, biodegradability and immunogenicity, can be absorbed by specific cancer cell selectivity, have been widely used diversification carrier system
System.A large amount of active groups are contained, such as carboxyl, hydroxyl, N- acetyl group and reduction end all can be to its chemical modifications in strand.Main
Method of modifying is wanted to have esterification, crosslinking, grafting etc. to react, wherein hydroxyl and carboxyl are common covalent modification sites.Polymer-medicine
Thing conjugate has enough functional groups, so as to carry out chemical coupling.
Most of malignant tumor and its surrounding stromal tissue have the hyaluronic acid (HA) of high concentration, and overexpression HA receptor
CD44 or RAHMM, such as deformation galactophore epithelial cell, ovarian tumor cell, colon cancer, pulmonary carcinoma, acute leukemia cellses etc..With HA
Passive target can be worked in coordination with by the conjugate building for part with receptor-mediated active targeting, so that medicine is oriented and be gathered in tumor portion
Position, changes ingestion of medicines mechanism, reaches active targeting.
Chytil etc. passes through hydrazone key and amycin (DOX) and large hydrophilic molecular HPMA covalent coupling is formed macromole coupling
Thing (HPMA-DOX), vitro cytotoxicity experiment shows, HPMA-DOX is to mouse T cell lymphoma EL4 cell, mouse B cell
The all relatively free amycin of the toxicity of lymphoma 38C13 cell, human large intestine cancer SW620 cell etc. is strong, and internal pharmacodynamic study shows
Its antitumor curative effect increase [Chytil, P., Etrych, T., Konak, C., Sirova, M., Mrkvan, T., Rihova,
B., Ulbrich, K., Properties of HPMA copolymer-doxorubicin conjugates with pH-
controlled activation：effect of polymer chain modification.J.Controlled
Release.2006,115 (1), 26-36.].The design such as Norbedo Stefano has synthesized a kind of hyaluronic acid-hydroxy-camptothecin
Alkali (HA-CPT) conjugate, it is substantially strong compared with irinotecan to the toxicity of HT-29 cell, H460 cell and H460-M2 cell, adopts
Use hyaluronic acid decorated hydroxy camptothecin, by increasing capacitance it is possible to increase the combination of medicine and cell, thus improving cellular uptake
[WO2009074678A2].
In recent years, amphipathic polyester has been a great concern as pharmaceutical carrier, and the hydrophobic core of internal layer is as non-co-
Valency vesicle, the hydrophilic shell of outer layer prevents protein adsorption, and it can be self-assembled into core-shell structure copolymer nanoparticle, thus avoiding non-specific
Reticuloendothelial system (RES) absorbs.By by hydrophilic PEG and hydrophobic group synthetic copolymer, such as with polylactic acid (PLA),
Polycaprolactone (PCL), polylactic-co-glycolic acid (PLGA) chemical coupling block copolymer.PEGization strategy can reduce liver picked-up
Rate, improves tumor-targeting.PEGization HA nanoparticle can shield HA surface in cyclic process, exposes HA target by before picked-up
Point, is more beneficial for improving the therapeutical effect of tumour medicine.Based on the drug delivery system of HA, the treatment for some tumors has
Great potential using value, novel Drug Delivery Systems provide theoretical foundation to the clinical practice of anticarcinogen.
Prodrug design strategy with paclitaxel as parent, is that the wide clinical application of paclitaxel provides new opportunity.This
Invention with targeting ligand hyaluronic acid for water-soluble backbone material, by different linking arms by antitumor drug such as paclitaxel,
Docetaxel, amycin, camptothecine are coupled into amphipathic polymer-anticarcinogen conjugate with hyaluronic acid, using from group
Macromole conjugate and amphipathic nature block polymer are prepared the targeting drug delivery system of novel composite nano grain so as to energy by dress technology
Enough selective target tumor positions, improve the Internal pharmacokinetics feature of chemotherapeutics, reduce toxic and side effects, extend internal delay
Time, improve antitumor curative effect.
Content of the invention
It is a further object to provide a kind of compound receiving of being combined with block copolymer of above-mentioned macromole conjugate
Grain of rice novel Drug Delivery Systems, it is to avoid the phagocytosis of reticuloendothelial system and renal excretion, extend the internal holdup time.
For reaching above-mentioned purpose, the present invention is realized by the following technical programs.Using amphipathic PEG block copolymerization
Thing and tumor targeting macromolecule conjugate carry out being self-assembly of new composite Nano kernel structure.Hydrophilic segment PEG is led to
Cross modification nanoparticle system, there is long circulating action and the function of avoiding reticuloendothelial system phagocytic, hydrophobic chain segment is permissible
Insertion nanoparticle core, forms more stable nanoparticle drug-supplying system.This Multifunction composite nano-granule drug-supplying system, can
Improve blood stability, improve tumor-targeting, reduce toxic and side effects, extend the internal holdup time, strengthen antitumor and treat
Effect, is more suitable for clinic and quotes, be that anticancer strategy provides new Research Thinking.
The present invention provides a kind of or series to have the macromole hyaluronic acid-antitumor of tumor-targeting and multi-function action
Medicine conjugate and composite nano-granule compositionss it is characterised in that：This conjugate be using antitumor drug active group hydroxyl,
Amino and hydroxyl, amino or carboxyl (- OH ,-NH2With-OH ,-NH2,-COOH), by linking arm formed antineoplastic agent-linking arm
The hydroxyl of intermediate product, hyaluronic acid or its ammonium salt or carboxyl are reacted with intermediate product, form hyalomitome by Covalent bonding together
Acid-antineoplastic agent conjugate.
Hyaluronic acid-antineoplastic agent Conjugate Molecules structural representation such as Fig. 1；
The purpose of the present invention is that hyaluronic acid-antineoplastic agent conjugate is assembled into nanoparticle, and prior purpose is logical
Cross self-assembling technique and hyaluronic acid-antineoplastic agent conjugate and amphipathic nature block polymer are assembled into many work(by electrostatic interaction
Energy composite nano-granule, described Multifunctional composite nanometer kernel structure schematic diagram such as Fig. 2；
Wherein：Spacer is linking arm, and AB, ABA, ABC are amphipathic-polyester block copolymer；Amphipathic group is poly-
Ethylene glycol, single methoxy ether Polyethylene Glycol, polyamino acid；
Hyaluronic acid-antineoplastic agent conjugate and the combination weight of amphipathic ester copolymer are：1: 0.2～5, hyalomitome
The weight that acid is coupled with antineoplastic agent ratio is for 1: 0.05～1；
Hyaluronic acid is 1: 20～1: 1 with the mol ratio of antitumor drug, and hyaluronic acid substitution value is 3%～48%；Excellent
Select mol ratio 1: 5～1: 2, preferably clear matter acid substitution value is 12%～35%.
The linking arm (Spacer) of described hyaluronic acid and antineoplastic agent is dicarboxylic anhydride, dicarboxylic acids, end have amino or
The carboxylic acid of hydroxyl；For the antitumor drug containing amino or carboxyl, select end have the linking arm of carboxyl with formed ester bond or
Amide is bonded, and for the antitumor drug containing carboxyl, the linking arm selecting to have hydroxyl or primary amino radical is to form ester bond or acyl
Amine key；Described link arm is selected from aminoacid, succinic anhydrides, cis-aconitic, aconitic anhydride of taking advantage of a situation, hemisuccinic acid ester；
It is L-Glutamic Acid, L-Alanine, METHIONINE, L-phenylalanine, the bright ammonia of L- that described aminoacid selects aminoacid
Acid, L-Isoleucine, L-PROLINE, 1B or glycine.
Hyaluronic acid-antineoplastic agent the conjugate with tumor-targeting is it is characterised in that described antitumor drug is main
For paclitaxel, Docetaxel, amycin, daunorubicin, epirubicin, camptothecin, methotrexate, mitoxantrone or its spread out
Biology, preferably paclitaxel, Docetaxel, amycin, camptothecin.
There is the hyaluronic acid-antineoplastic agent conjugate of tumor-targeting it is characterised in that described hyaluronan molecule amount
For 4500～580000, hyaluronic acid or hyaluronic acid are treated that ammonium salt is selected from hyaluronic acid-TBAH salt
(HA-TBA), hyaluronic acid-cetyl trimethylammonium bromide salt (HA-CTA), hyaluronic acid-adipic dihydrazide (HA-
ADH), the preparation method of its ammonium salt is as follows：
A. weigh hyaluronate sodium and be dissolved in pure appropriate amount of water, plus the hydrochloric acid of 0.1mol/L is acidified, through cation exchange
Resin treatment, lyophilization, obtain final product hyaluronic acid (HA).
B.HA-TBA：Weigh hyaluronate sodium appropriate, be dissolved in the hydrochloric acid solution of 0.1mol/L, at acidifying under the conditions of 4 DEG C
Reason 15h.Plus TBAH (TBA) adjusts pH to 8.0, the dialysis treatment removal of impurity in 3 days, lyophilization, obtain final product HA-TBA.
C.HA-CTA：Weigh cetyl trimethylammonium bromide (CTAB), 40 DEG C are dissolved in pure water (a phase)；Separately weigh
Hyaluronate sodium is dissolved in pure water (b phase) in right amount；A is dropped in b, 40 DEG C of reaction 2h, separate out white precipitate, filter and must consolidate
Body, vacuum drying, obtain final product HA-CTA.
D.HA-ADH：Hyaluronate sodium is dissolved in pure water, plus adipic dihydrazide (ADH), use 0.05mol/L hydrochloric acid
It is 4.75 that solution is adjusted to pH, plus carbodiimide hydrochloride (EDC HCl), reacts 2～4h, plus 0.01mol/LNaOH adjusts pH
To neutral, deionized water dialysis (MWCO=6000), lyophilization, obtain final product HA-ADH.
The preparation method with tumor-targeting and multi-functional hyaluronic acid-antineoplastic agent conjugate that the present invention provides
As follows including step：
A. the preparation of the antitumor drug-HA conjugate containing hydroxyl
(1) medicine-linking arm intermediate product：Antitumor drug containing hydroxyl is dissolved in organic solvent, with Fmoc- ammonia
Base acid, dicarboxylic anhydride or half ester, for linking arm or with Boc- aminoacid, dicarboxylic anhydride as linking arm, add catalyst and dehydrant to carry out
Steglich react, temperature control at 0～30 DEG C, magnetic agitation react 0.5～50h, add inert organic solvents precipitate drug-
Linking arm product, isolates and purifies intermediate product using column chromatography, obtains final product pharmacological activation-linking arm intermediate；
Hyaluronic acid or hyaluronic acid ammonium salt activator：Hyaluronate sodium is dissolved in the hydrochloric acid solution of 0.1mol/L, acidifying
Obtain hyaluronic acid；By hyaluronic acid plus TBAH or cetyl trimethylammonium bromide, react 0.25～4h, vacuum
2～4h is dried, obtains final product hyaluronic acid ammonium salt activator；
(2) hyaluronic acid or hyaluronic acid ammonium salt activator are dissolved in organic solvent with medicine-linking arm intermediate product, plus
Catalyst and dehydrant, at 10～50 DEG C, magnetic agitation reacts 2～60h to temperature control, and reactant is put bag filter, deionization
The removal of impurity in 3 days of water dialysis treatment, lyophilization, obtain final product hyaluronic acid-antineoplastic agent conjugate；Or
B. the preparation of the antitumor drug-HA conjugate containing amino or hydroxyl
(1) antitumor drug containing amino or hydroxyl is dissolved in organic solvent, adds 9-fluorenylmethyloxycarbonyl chlorine, with ammonia
Base acid, dicarboxylic anhydride or half ester reaction, add catalyst and dehydrant, obtain final product N-Fmoc- antitumor drug intermediate；
Hyaluronic acid or hyaluronic acid ammonium salt activator：Hyaluronate sodium is dissolved in solvent and is acidified to obtain hyaluronic acid；Will
Hyaluronic acid adds TBAH or cetyl trimethylammonium bromide, reacts 0.25～6h, lyophilization, obtains final product transparent
Matter acid ammonium salt activator；
(2) derivatives of hyaluronic acids activator and medicine-linking arm intermediate product are dissolved in organic solvent, add catalyst
And dehydrant, plus piperidines, at 10～50 DEG C, magnetic agitation reacts 2～60h to temperature control.Using cross-linking dextran G-25 purification
Product, lyophilization, obtain final product hyaluronic acid-antineoplastic agent conjugate；Or
C. containing carboxyl antitumor drug-HA conjugate preparation
(1) antitumor drug containing carboxyl is dissolved in solvent, there is using Boc- aminoacid or end the carboxylic of sulfydryl
Acid is linking arm, carries out Steglich reaction, obtain final product in the middle of antitumor drug-linking arm in the presence of catalyst and dehydrant
Body；
Hyaluronate sodium is dissolved in pure water, plus 0.05mol/L hydrochloric acid, obtain HA；Add adipic dihydrazide, plus dehydration
0.25～6h is reacted in agent, adjusts pH to neutral, deionized water dialysis, lyophilization with 0.01mol/LNaOH, obtains final product hyalomitome
Acid-adipic dihydrazide activator；
(2) hyaluronic acid-adipic dihydrazide activator and medicine-linking arm intermediate product are dissolved in organic solvent,
The effect of dehydrant and catalyst is reacted, and temperature is 10～50 DEG C, and magnetic agitation reacts 2～60h, and reactant is put dialysis
Bag, the deionized water dialysis removal of impurity in 3 days, lyophilization, obtain final product hyaluronic acid-antitumor medicine conjugate.
The preparation process of hyaluronic acid-antineoplastic agent conjugate, described dehydrant is selected from DIC
(DIC), N, N- dicyclohexylcarbodiimide (DCC), 1- ethyl (3- dimethylamino-propyl) carbodiimide or carbodiimide class are spread out
Biological；Catalyst is selected from DMAP (DMAP), DMA, 1- hydroxyl phenylpropyl alcohol triazole, 4- pyrrolidine
Yl pyridines (PPY), piperidines, triethylamine；
Wherein, organic solvent is Methanamide, DMF, dimethyl sulfoxide, oxolane；Described inertia is organic
Solvent is methanol, ethanol, ether, acetone.
There is tumor-targeting and the hyaluronic acid-antineoplastic agent conjugate of multi-function action and composite nano-granule combines
Thing it is characterised in that described amphipathic-polyester be AB, ABA or ABC type di-block copolymer, triblock copolymer or grafting altogether
Polymers, wherein di-block copolymer are selected from mPEG-PLA, mPEG-PCL, mPEG-PLGA, mPEG-PAA, and triblock copolymer selects
From mPEG-PLLA-PMMD, PLA-PEG-PLA, PCL-PEG-PCL, PLGA-PEG-PLGA or PAA-PEG-PAA.
Hyaluronic acid-antineoplastic agent conjugate and the composite Nano with tumor-targeting and multi-function action of the present invention
Grain compositionss are it is characterised in that the preparation method of described composite nano-granule compositionss is as follows：
(1) solvent diffuse-film dispersion method-dialysis are adopted：By hyaluronic acid-antineoplastic agent conjugate dissolving or dispersion
In pure water, amphipathic polyester is dissolved in organic solvent, by above-mentioned two-phase mixtures magnetic agitation 0.5～4h, 30～50 DEG C of rotations
Turn evaporation and remove organic solvent, Probe Ultrasonic Searching 5～60min, deionized water dialysis (MWCO is 6000), obtain final product composite nano-granule and be suspended
Liquid；
(2) Freeze Drying Technique：Composite nano-granule suspension is added freeze drying protectant, lyophilization, obtains final product compound receiving
Grain of rice lyophilized powder；
Wherein said organic solvent is one or more of dichloromethane, chloroform, methanol, ethanol, acetonitrile, acetone etc.
Mixed solvent；Wherein solvent described in dialysis is deionized water, purified water, and dialysis time is 12～120h, preferably 24～72h.
Described composite nano-granule freeze drying protectant be selected from one of Mannitol, glucose, Lactose, sucrose or trehalose or
Several.
Hyaluronic acid provided by the present invention-antineoplastic agent conjugate and composite nano-granule compositionss are it is characterised in that institute
State hyaluronic acid-antineoplastic agent conjugate or assemble single nanoparticle with amphipathic polyester or be assembled into composite nano-granule, have
Improve anticarcinogen dissolubility, drug loading is more than 12%, maximum drug loading is 35%, and mean diameter is less than 200nm, good stability etc.
Multiple advantages.In vivo can biodegradation, this composite nano-granule, it can be avoided that the phagocytosis of reticuloendothelial system (RES) and kidney
Excretion, has long circulating action, internal drug effect is apparently higher than antineoplastic agent normal injection agent.Its clinically acceptable administration way
There are drug administration by injection, oral administration or mucosal drug delivery in footpath；Dosage form is injection, infusion solution, frozen powder for injection injection, tablet, capsule
Agent or powder spray.
Hyaluronic acid provided by the present invention-antineoplastic agent conjugate and composite nano-granule compositionss are it is characterised in that be somebody's turn to do
Macromole conjugate nanoparticle or composite nano-granule can be used for treating nonsmall-cell lung cancer, breast carcinoma, ovarian cancer, cervical cancer,
The single or drug combination of the malignant tumor such as gastric cancer, hepatocarcinoma, leukemia.Novel Drug Delivery Systems provide to the clinical practice of anticarcinogen
Experiment basis and theoretical foundation, have major clinical significance.
Brief description
Fig. 1 is hyaluronic acid-antineoplastic agent Conjugate Molecules structural representation
Fig. 2 is paclitaxel Multifunctional composite nanometer kernel structure schematic diagram
Fig. 3 is paclitaxel injection (PTX- injection), hyaluronic acid-paclitaxel conjugate (HA-PTX) and composite Nano
Grain (mPP-HA-PTX-NPs) cytotoxicity experiment figure (n=3)
Fig. 4 is that hyaluronic acid-paclitaxel composite nano-granule digests lab diagram (n=3) in vitro
Specific embodiment
Embodiment 1
Hyaluronic acid-paclitaxel conjugate (HA-PTX) preparation (linking arm is L-Valine)
(1) 100mg fluorenylmethyloxycarbonyl-L-Valine (Fmoc-Val) and 170mg paclitaxel (PTX) are dissolved in dichloromethane
In, plus 24mg dimethylamino naphthyridine (DMAP) and 46mg carbodiimide hydrochloride (EDC HCl), ice bath reaction 1h, stir 8h.
With hydrochloric acid solution and the deionized water wash reactant liquor of 0.05mol/L, organic layer is dried with anhydrous magnesium sulfate, filters, concentrate, plus pyrrole
Pyrrolidone dichloromethane solution, stirs 3h.It is vacuum dried to obtain crude product.Using column chromatography purification product, obtain 2 '-valyl
Paclitaxel (2 '-Val-PTX).
(2) hyaluronate sodium is dissolved in purified water, cation exchange resin is processed, lyophilization obtains final product hyaluronic acid
(HA).600mg hyaluronic acid (HA) is dissolved in Methanamide, plus 38.4mg carbodiimide hydrochloride (EDC HCl) and 23mgN-
N-Hydroxysuccinimide (NHS), room temperature activates 30min, and 2 '-Val-PTX are dissolved in appropriate DMF, by it under nitrogen stream protection
Add in HA solution, stir 24h, plus acetone removes intermediate, 4 DEG C of standing 1h, sucking filtration, deionized water is redissolved and taken precipitation, dialysis
Process 72h (MWCO=6000), lyophilization, obtain final product hyaluronic acid-paclitaxel conjugate (HA-PTX).
Through proton nmr spectra1H NMR and infrared FT-IR identifies to its structure：
1H NMR (400Hz, D20)：8.12 (dd, J=7.08,1.44,2H), 7.75 (dd, J=7.08,1.44,2H), 7.61
(s, 1H), 7.58-7.49 (m, 4H), 7.47-7.32 (m, 6H), 7.13 (d, 9.21H2, 1H), 6.30 (s, 1H), 6.24 (t, J=
9.0,1H), 6.03 (dd, J=92H2, 3.5H2, 1H), 5.67 (d, J=7.11,1H), 5.54 (d, J=3.6H2,1H, C2), 4.98
(dd, J=9.57,1.44,1H), 4.45 (m, 1H), 4.30 (d, J=8.46,1H), 4.21 (d, J=8.46,1H), 3.81 (d, J=
6.9H2,1H), 3.73 (m, 1H, α-Met), 2.59 (m, 2H, γ-Met), 2.38 (s, 3H), 2.23 (s, 3H), 2.12 (m, 2H,
β-Met), 2.01 (s, 3H, δ-Met), 1.95 (s, 3H), 1.68 (s, 3H), 1.24 (s, 3H), 1.15 (s, 3H).
FT-IR：3423cm-1、2961cm-1、2927cm-1、1655cm-1、1490cm-1、1155cm-1、1082cm-1、
1048cm-1、949cm-1、711cm-1.
Embodiment 2
Hyaluronic acid-paclitaxel conjugate (HA-PTX) preparation (linking arm is methionine)
(1) 110mg fluorenylmethyloxycarbonyl-methionine (Fmoc-Met) and 170mg paclitaxel (PTX) are dissolved in dichloromethane
In, add 24mg dimethylamino naphthyridine (DMAP) and 46mg carbodiimide hydrochloride (EDC HCl), ice bath reacts 1h, room temperature is stirred
Mix 8h.With hydrochloric acid solution and the deionized water wash reactant liquor of 0.05mol/L, anhydrous magnesium sulfate is dried organic layer and uses, and filters, dense
Contracting, plus ethanolamine dichloromethane solution, stir 4h.Vacuum drying obtains crude product, using column chromatography purification product, obtains
2 '-methionyl paclitaxel (2 '-Met-PTX).
(2) weigh cetyl trimethylammonium bromide (CTAB), under the conditions of 40 DEG C, be dissolved in purified water (A phase)；Separately weigh
Hyaluronate sodium is dissolved in purified water (B phase)；40 DEG C add to A in B, react 2h, filter, obtain white solid, use hot water injection
Precipitation 2～3 times, vacuum drying, obtain final product HA-CTA.1.2g HA-CTA is dissolved in DMF, plus 38.4mg carbodiimide hydrochloride
(EDC HCl) and 23mg N-hydroxy-succinamide (NHS), activates 30min, 2 '-Met-PTX is dissolved in DMF, nitrogen stream
It is added under protection in HA-CTA, stirs 24h, add acetone to remove intermediate, 4 DEG C of placement 1h, sucking filtration, deionized water is multiple
Molten take precipitation, dialysis treatment 72h (MWCO=6000), lyophilization, obtain final product hyaluronic acid-paclitaxel conjugate (HA-PTX).Warp
Proton nmr spectra1H NMR and infrared FT-IR identifies to its structure.
Embodiment 3
Hyaluronic acid-Docetaxel conjugate (HA-DTX) preparation (linking arm is leucine)
(1) 105mg fluorenylmethyloxycarbonyl-leucine (Fmoc-Leu) and 160mg Docetaxel (DTX) are dissolved in dichloromethane
In alkane, plus 24mg dimethylamino naphthyridine (DMAP) and 46mg carbodiimide hydrochloride (EDC HCl), condition of ice bath reaction 1h, room
Temperature stirring 8h.Use hydrochloric acid solution and the deionized water wash of 0.05mol/L respectively, anhydrous magnesium sulfate is dried, filter, concentrate, add
Strong aqua ammonia dioxane/4M NaOH (30: 9: 1), stirs 2h.Vacuum drying, obtains crude product, is produced using column chromatography purification
Thing, obtains 2 '-leucyl paclitaxel (2 '-Leu-DTX).Through proton nmr spectra1H NMR and infrared FTIR is carried out to its structure
Identification.
(2) hyaluronate sodium is dissolved in 0.1mol/L hydrochloric acid solution, 4 DEG C of acidifying 15h.Plus TBA (tetrabutylammonium hydroxide
Ammonium) adjust pH to 8.0, dialysis treatment 3 days, the removal of impurity, lyophilization, obtain product HA-TBA.1.0g HA-TBA is dissolved in DMF
In, plus 38.4mg carbodiimide hydrochloride (EDC HCl) and 23mgN- N-Hydroxysuccinimide (NHS), ice bath activation 30min.
The DMF solution of 2 '-Leu-PTX is added in HA-TBA solution, 35 DEG C of stirrings 24h, deionized water dialysis treatment 72h (MWCO=
6000), lyophilization, obtains final product hyaluronic acid-paclitaxel conjugate (HA-DTX).Through proton nmr spectra1H NMR and infrared
FT-IR identifies to its structure.
Embodiment 4
Hyaluronic acid-amycin conjugate (HA-DOX) preparation (linking arm is cis-aconitic acid anhydride)
(1) by 50mg doxorubicin hydrochloride (DOX) and the molten (NaHCO of 8mg sodium bicarbonate3) in purified water, stirring；By 29mg9-
Fluorenylmethyloxycarbonyl chlorine (9-Fmoc) is dissolved in ethyl acetate, adds to (pH is 7～8) in above-mentioned solution, plus saturation NaHCO3Solution,
Reaction 6h, extracts ethyl acetate layer, recrystallization in 0.1% trifluoroacetic acid, and ice ester rinses crystallization, vacuum drying, obtains N-
Fmoc-DOX.It is added into, containing in 36mg cis-aconitic acid anhydride (CAA), 12mg dimethylamino naphthyridine (DMAP) DMF, stirring 12h,
Vacuum drying, carries out purification, ethyl acetate using silicagel column：Normal hexane (90: 10) eluting, obtains final product N- fluorenylmethyloxycarbonyl -13- suitable
Formula Aconitum carmichjaelii Debx. acyl amycin (N-Fmoc-CA-DOX).Through proton nmr spectra1HNMR and infrared FT-IR identifies to its structure.
(2) hyaluronic acid is prepared with embodiment 1.600mg hyaluronic acid (HA) is dissolved in distilled water, adds 16mg diisopropyl
Base carbodiimide (DIC) and 23mg N-hydroxy-succinamide (NHS), ice bath activates 30min, adds N- under the conditions of 50 DEG C
Fmoc-DOX-CA product, 50 DEG C of stirring 12h, plus appropriate piperidines, stir 2h.Using cross-linking dextran G-25 purified product, obtain
Hyaluronic acid-Ah can's mycin conjugate (HA-DOX).Through proton nmr spectra1H NMR and infrared FT-IR reflects to its structure
Fixed.
Embodiment 5
The synthesis (linking arm is succinic anhydrides) of hyaluronic acid-amycin conjugate (HA-DOX)
(1) by 50mg doxorubicin hydrochloride (DOX) and 8mg sodium bicarbonate (NaHCO3) be dissolved in purified water, stirring；Will
29mg9- fluorenylmethyloxycarbonyl chlorine (9-Fmoc) is dissolved in ethyl acetate, drops to above-mentioned aqueous solution (pH is 7～8), plus saturation
NaHCO3Solution, reacts 6h, extracts ethyl acetate layer, recrystallization in 0.1% trifluoroacetic acid, it is residual that ice ester rinses clean surface
The 9-Fmoc staying, vacuum drying, obtain N-Fmoc-DOX.It is added into succinic anhydrides containing 8mg (SA), 12mg dimethylamino pyrrole
In pyridine (DMAP) DMF, 12h is stirred at room temperature, vacuum drying.Purification, ethyl acetate are carried out using silicagel column：Normal hexane (95: 5) is washed
De-, obtain final product N- fluorenylmethyloxycarbonyl -13- succinyl amycin (N-Fmoc-SA-DOX).
(2) 600mg hyaluronic acid (HA) is dissolved in purified water, plus 38.4mg carbodiimide hydrochloride (EDC HCl) and
23mg N-hydroxy-succinamide (NHS), ice bath activates 30min, adds N-Fmoc-SA-DOX, 50 DEG C of stirring 24h, plus piperidines
3mL, is stirred at room temperature 2h.Using cross-linking dextran G-25 purified product, obtain final product hyaluronic acid-amycin conjugate (HA-DOX).
Through proton nmr spectra1H NMR and infrared FT-IR identifies to its structure.
Embodiment 6
Hyaluronic acid-camptothecin conjugates (HA-CPT) preparation (linking arm is hemisuccinic acid ester)
(1) 37mg hemisuccinic acid list tertiary butyl ester and 50mg camptothecine (CPT) are dissolved in dichloromethane, add 24mg bis-
Methylamino pyridine (DMAP) and DIC, ice bath reacts 1h, room temperature magnetic agitation 8h.Salt with 0.05mol/L
Acid solution and deionized water wash reactant liquor, anhydrous magnesium sulfate dry filter, rotary evaporation removes dichloromethane, and residue adds acetone
Recrystallization, filters, vacuum drying, obtains solidss, plus containing in 40% dichloromethane solution of trifluoroacetic acid, stands 1h, evaporates
It is dried, plus recrystallizing methanol, filter, vacuum drying, obtain 20- hemisuccinic acid ester-CPT.Through proton nmr spectra1H NMR and
Infrared FT-IR identifies to its structure.
(2) HA-CTA is prepared with embodiment 2.Will be molten for 1.2g hyaluronic acid-cetyl trimethylammonium bromide (HA-CTA)
In DMF, add 26.6mg N, N- dicyclohexylcarbodiimide (DCC) and 23mg N-hydroxy-succinamide (NHS), ice bath
Activation 30min.Plus the DMF solution of 20- hemisuccinic acid ester-CPT is to above-mentioned HA-CTA solution, 35 DEG C of stirring 24h, plus saturation chlorine
Change sodium solution, stir 30min, plus 50mL dehydrated alcohol, filter, after DMF and washing with alcohol, residue is dissolved in deionized water
In, dialysis purification (MWCO=6000), lyophilization, obtain hyaluronic acid-camptothecin conjugates (HA-CPT).Through hydrogen nuclear magnetic resonance
Spectrum1H NMR and infrared FT-IR identifies to its structure.
Embodiment 7
Hyaluronic acid-camptothecin conjugates (HA-CPT) preparation (linking arm is aconitic anhydride of taking advantage of a situation)
(1) 50mg camptothecine (CPT) and 55mg aconitic anhydride (CAA) of taking advantage of a situation is dissolved in dichloromethane, adds 12mg bis-
Methylamino pyridine (DMAP) and 23mg carbodiimide hydrochloride (EDC HCl), ice bath reacts 1h, is transferred to room temperature, and nitrogen is protected
Lower stirring 24h, with 0.05mol/L hydrochloric acid extractive reaction liquid, rotation is evaporated off dichloromethane, obtains solid product, silica gel column chromatography,
Dichloromethane：Methanol (100: 1) purification, obtains final product cis-aconitic anhydride-camptothecine (CA-CPT).Through proton nmr spectra1H NMR
With infrared FT-IR, its structure is identified.
(2) hyaluronic acid is prepared with embodiment 1.Take 600mg hyaluronic acid (HA) to be dissolved in distilled water, add 260mg own
Dihydrazi (ADH), adjusting pH with 0.05mol/L hydrochloric acid is 4.75, plus 50mg carbodiimide hydrochloride (EDC HCl), room temperature
React 2.5h, plus 0.01mol/LNaOH adjusts pH to neutral, deionized water dialysis, lyophilization, obtains final product HA-ADH.By HA-
ADH is dissolved in DMF, adds CA-CPT, 38.4mg carbodiimide hydrochloride (EDC HCl) and 23mg N-hydroxy-succinamide
(NHS), the lower 35 DEG C of activation 30min of N2, magnetic agitation 24h, deionized water dialysis treatment 72h (MWCO=6000), lyophilization,
Obtain final product hyaluronic acid-paclitaxel conjugate (HA-CPT).Through proton nmr spectra1HNMR and infrared FT-IR is carried out to its structure
Identification.
Embodiment 8
Hyaluronic acid-paclitaxel and the preparation of amphipathic polyester composite nano-granule
40mg HA-Met-PTX conjugate is scattered in purified water, adds in the acetonitrile solution of 40mg mPEG-PLA,
24h, Probe Ultrasonic Searching 20min are stirred at room temperature, 37 DEG C of rotary evaporations remove acetonitrile, Probe Ultrasonic Searching 10min, obtain final product mPEG-PLA-HA-
PTX composite nano-granule (mPP-HA-PTX-NPs).Particle diameter distribution and Zeta electricity are measured using ZetaSizer laser particle size analyzer
Position.Its particle diameter is 146.2 ± 7.8nm, and PI is 0.12, and Zeta potential is -26.7mV.
Embodiment 9
Hyaluronic acid-Docetaxel and the preparation of amphipathic polyester composite nano-granule
40mg HA-DTX is dissolved in purified water water, takes 20mg mPEG-PLGA to be dissolved in dichloromethane, both mixing,
Stirring 3h, Probe Ultrasonic Searching 30min, 37 DEG C of hydration 2h, rotary evaporation removes dichloromethane, Probe Ultrasonic Searching 10min, obtains final product mPEG-
PLGA-HA-DTX composite nano-granule (mPP-HA-DTX-NPs).Particle diameter distribution is measured using ZetaSizer laser particle size analyzer
And Zeta potential.Result shows, its mean diameter is 238.9 ± 12.3nm, and PI is 0.25, and Zeta potential is -21.7mV.
Embodiment 10
Hyaluronic acid-amycin and the preparation of amphipathic polyester composite nano-granule
40mg HA-DOX is dissolved in distilled water, separately takes 40mg mPEG-PCL to be dissolved in chloroform, both mixing, stirring
3h, Probe Ultrasonic Searching 15min, 37 DEG C of hydration 1h, rotary evaporation removes chloroform, Probe Ultrasonic Searching 20min, obtains final product mPEG-PCL-HA-
DOX composite nano-granule (mPP-HA-DOX-NPs).Particle diameter and Zeta potential are measured using ZetaSizer laser particle size analyzer,
Its mean diameter is 184.2 ± 9.3nm, and PI is 0.18, and Zeta potential is -22.7mV.
Embodiment 11
Hyaluronic acid-paclitaxel composite nano-granule digests experiment in vitro
HA-PTX conjugate is dissolved in normal saline in right amount, crosses 0.8 μm of filter membrane, plus hyaluronidase (50IU/mL) and ester
Enzyme (20IU/mL), is placed in bag filter and is dialysed.With 0.2% Tween 80 phosphate buffer (PBS, pH5.0, pH6.8 and
PH7.4) it is release medium.Bag filter is placed in release medium, 37 DEG C of water-baths vibration dialysis treatment (MWCO=3000), respectively
In 0.5,1,2,4,8,12,24,48,72h sampling 5mL, add equivalent medium, at ultraviolet spectrophotometer 230nm simultaneously
Determination sample absorbance, investigates enzymolysis behavior under different pH condition for the HA-PTX conjugate.
Result is as shown in Figure 4.In the presence of hyaluronidase and esterase, HA-PTX can reach quick release and go out Ramulus et folium taxi cuspidatae
Alcohol, the burst size that dissociates respectively in 3 days is 82.1% (pH7.4), 90.3% (pH6.8) and 96.7% (pH5.0).In acid condition
(pH5.0) hydrolysis is the fastest, and 24h is 90% enzymolysis；Raise with pH value, digest reduced rate, illustrate hyaluronidase in pH5.0 bar
Under part, activity is best.Result shows that HA-PTX is easy to digest at the tumor tissues position of slant acidity, is conducive to antitumor action
Play.
Embodiment 12
Vitro cytotoxicity is studied
Take the logarithm the non-small cell lung cancer cell strain A549 cell of trophophase, 1 × 104Individual/hole is inoculated in 96 orifice plates, culture
After 24h, every hole adds the medicine (PTX injection, HA-PTX and mPP-HA-PTX-NPs) of respective concentration, each group sample paclitaxel
Concentration is respectively 0.1,1,5,10,20 μ g/mL.Continue incubation 72h, add MTT solution (1mg/mL) 0.1mL, after incubation 4h, inhale
Except in the hole supernatant, each hole adds DMSO100 μ L, in 570nm microplate reader determination sample absorbance (ODtest).And with phase
Measure the absorbance of blank group (acellular strain) and matched group (n.s) with method, be designated as OD respectivelyblankAnd ODcontrol.
Calculate cell strain survival rate according to the following equation, evaluate the cytotoxicity of sample.
Result as shown in figure 3, increase with concentration, deposit by PTX injection, the cell of HA-PTX and mPP-HA-PTX-NPs
Motility rate all declines, and shows the cytotoxicity with concentration dependent, that is, be in concentration-effect relation, its IC50Value is respectively 13.71
± 1.09 μ g/mL, 3.02 ± 0.25 μ g/mL and 6.52 ± 0.51 μ g/mL.In 0.1～20 μ g/mL concentration range, three kinds of systems
Agent suppresses the ability of A549 cell growth to have significant difference (P<0.01), HA-PTX and mPP-HA-PTX-NPs has to A549
There is stronger cytotoxicity, the modification of HA can make nanoparticle pass through the receptor-mediated endocytosis of CD44 to enter cell in a large number, make
HA-PTX-NPs can in the cell in the presence of enzyme degraded dissociate active pharmaceutical ingredient, play antitumor curative effect.mPP-
The anti tumor activity in vitro of HA-PTX-NPs is less than HA-PTX-NPs, the anti-tumor in vivo activity no shadow to mPP-HA-TPX-NPs
Ring.
Embodiment 13
Internal pharmacodynamic study
Experimental program：S180 mice with tumor is randomly divided into 4 groups, every group 5, tail vein injection saline respectively
(0.15mL), PTX injection (7.5mg/kg), HA-PTX (dosage 7.5mg/kg) and mPP-HA-PTX-NPs (dosage 7.5mg/
Kg), it is administered respectively at tail vein injection on the 0,2,4,6,8th.In the 9th day by sacrifice, strip and separate tumor, weigh, calculate
Tumour inhibiting rate, carries out pharmacodynamic evaluation.
The pharmacodynamic evaluation (n=5) of the different formulation for paclitaxel of table 1.
Illustrated by table 1, HA-PTX and mPP-HA-PTX-NPs has obvious inhibition, Mice Body to S180 mice with tumor
Weight average has notable growth, shows that toxicity is remarkably decreased.MPP-HA-PTX-NPs has more preferable anti-tumor in vivo compared with HA-PTX and lives
Property and lower toxicity (mPP-HA-PTX-NPs Mouse Weight increases than HA-PTX group high 11.28%).Pharmacodynamics in vitro is tested
Deducibility PEG- polyester can increase the stability of composite nano-granule.MPP-HA-PTX-NPs can strengthen tumor-targeting, fall
Low toxicity side effect, improves antitumor drug effect.
Claims (5)
1. have tumor-targeting effect hyaluronic acid-antineoplastic agent conjugate composite nano-granule compositionss it is characterised in that：
This conjugate is using the active group hydroxyl of antitumor drug, amino and hydroxyl, amino or carboxyl, is formed anti-by linking arm
Tumor medicine-linking arm intermediate product, the hydroxyl of hyaluronic acid or its ammonium salt or carboxyl are reacted with intermediate product, by covalently bonded
Conjunction forms hyaluronic acid-antineoplastic agent conjugate；Described hyaluronic acid-antineoplastic agent conjugate and amphipathic ester copolymer group
Fill as Multifunctional composite nanometer grain compositionss；
Described antitumor drug is paclitaxel, Docetaxel, amycin and camptothecine；Described hyaluronan molecule amount is 4500
～580000, the weight that hyaluronic acid is coupled with antineoplastic agent is than for 1: 0.05～1；Hyaluronic acid is fed intake with antitumor drug
Than for 1: 20～1: 1, hyaluronic acid substitution value is 3%～48%；Hyaluronic acid-antineoplastic agent conjugate is with amphipathic polyester altogether
The composition weight of polymers is than for 1: 0.2～5；
Described linking arm selects aminoacid, succinic anhydrides, cis-aconitic, aconitic anhydride of taking advantage of a situation, hemisuccinic acid ester；Described amino
Acid selects L-Glutamic Acid, L-Alanine, METHIONINE, L-phenylalanine, L-Leu, L-Isoleucine, L-PROLINE, L-
Lysine or L- glycine；
Wherein, amphipathic ester copolymer is selected from di-block copolymer or triblock copolymer；Described di-block copolymer is selected from
MPEG-PLA, mPEG-PCL, mPEG-PLGA, mPEG-PAA, triblock copolymer is selected from mPEG-PLLA-PMMD, PLA-PEG-
PLA, PCL-PEG-PCL, PLGA-PEG-PLGA or PAA-PEG-PAA；PMMD be polyalanine, polylysine, polyglycine,
Polyglutamic acid, m is 500～20000.
2. hyaluronic acid described in claim 1-antineoplastic agent conjugate composite nano-granule compositionss are it is characterised in that hyalomitome
The preparation process of acid-antineoplastic agent conjugate comprises the steps：
A. the preparation of the antitumor drug-HA conjugate containing hydroxyl
(1) medicine-linking arm intermediate product：Antitumor drug containing hydroxyl is dissolved in organic solvent, with Fmoc- amino
Acid, dicarboxylic anhydride or half ester, for linking arm or with Boc- aminoacid, dicarboxylic anhydride as linking arm, add catalyst and dehydrant to carry out
Steglich react, temperature control at 0～30 DEG C, magnetic agitation react 0.5～50h, add inert organic solvents precipitate drug-
Linking arm product, isolates and purifies intermediate product using column chromatography, obtains final product pharmacological activation-linking arm intermediate；
Hyaluronic acid or hyaluronic acid ammonium salt activator：Hyaluronate sodium is dissolved in the hydrochloric acid solution of 0.1mol/L, is acidified thoroughly
Bright matter acid；By hyaluronic acid plus TBAH or cetyl trimethylammonium bromide, react 0.25～4h, vacuum drying
2～4h, obtains final product hyaluronic acid ammonium salt activator；
Hyaluronic acid or the treated ammonium salt of hyaluronic acid are selected from hyaluronic acid-TBAH salt, hyaluronic acid-ten six
Alkyl trimethyl ammonium bromide salt, hyaluronic acid-adipic dihydrazide；
(2) hyaluronic acid or hyaluronic acid ammonium salt activator are dissolved in organic solvent with medicine-linking arm intermediate product, plus catalysis
Agent and dehydrant, at 10～50 DEG C, magnetic agitation reacts 2～60h to temperature control, reactant is put bag filter, deionized water is saturating
Analysis processes the removal of impurity in 3 days, lyophilization, obtains final product hyaluronic acid-antineoplastic agent conjugate；Or
B. the preparation of the antitumor drug-HA conjugate containing amino or hydroxyl
(1) antitumor drug containing amino or hydroxyl is dissolved in organic solvent, adds 9-fluorenylmethyloxycarbonyl chlorine, with amino
Acid, dicarboxylic anhydride or half ester reaction, add catalyst and dehydrant, obtain final product N-Fmoc- antitumor drug intermediate；
Hyaluronic acid or hyaluronic acid ammonium salt activator：Hyaluronate sodium is dissolved in solvent and is acidified to obtain hyaluronic acid；Will be transparent
Matter acid plus TBAH or cetyl trimethylammonium bromide, react 0.25～6h, lyophilization, obtain final product hyaluronic acid
Ammonium salt activator；
(2) derivatives of hyaluronic acids activator and medicine-linking arm intermediate product are dissolved in organic solvent, add catalyst and take off
Water preparation, plus piperidines, at 10～50 DEG C, magnetic agitation reacts 2～60h to temperature control, using cross-linking dextran G-25 purified product,
Lyophilization, obtains final product hyaluronic acid-antineoplastic agent conjugate；Or
C. the preparation of the antitumor drug-HA conjugate containing carboxyl
(1) antitumor drug containing carboxyl is dissolved in solvent, using the carboxylic acid that Boc- aminoacid or end have sulfydryl is
Linking arm, carries out Steglich reaction in the presence of catalyst and dehydrant, obtains final product antitumor drug-linking arm intermediate；
Hyaluronate sodium is dissolved in pure water, plus 0.05mol/L hydrochloric acid, obtain HA；Add adipic dihydrazide, plus dehydrant is anti-
Answer 0.25～6h, adjust pH to neutral with 0.01mol/LNaOH, deionized water is dialysed, lyophilization, obtain final product hyaluronic acid-own
Dihydrazi activator；
(2) hyaluronic acid-adipic dihydrazide activator and medicine-linking arm intermediate product are dissolved in organic solvent, in dehydration
The effect of agent and catalyst is reacted, and temperature is 10～50 DEG C, and magnetic agitation reacts 2～60h, and reactant is put bag filter,
The deionized water dialysis removal of impurity in 3 days, lyophilization, obtain final product hyaluronic acid-antitumor medicine conjugate；
Wherein said dehydrant is selected from DIC, N, N- dicyclohexylcarbodiimide, 1- ethyl (3- dimethylamino
Propyl group) carbodiimide or carbodiimide analog derivative；Catalyst is selected from DMAP, DMA, 1- hydroxyl
Base phenylpropyl alcohol triazole, 4- pyrollidinopyridine, piperidines, triethylamine；
Described organic solvent is Methanamide, DMF, dimethyl sulfoxide, oxolane；Described inertia organic solvent
For methanol, ethanol, ether, acetone, acetonitrile one or two.
3. hyaluronic acid according to claim 1-antineoplastic agent conjugate composite nano-granule compositionss are it is characterised in that described
The preparation method of composite nano-granule compositionss is as follows：
(1) solvent diffuse-film dispersion method-dialysis are adopted：Hyaluronic acid-antineoplastic agent conjugate is dissolved or dispersed in pure
In water purification, amphipathic ester copolymer is dissolved in organic solvent, by above-mentioned two-phase mixtures magnetic agitation 0.5～4h, 30～50 DEG C
Rotary evaporation removes organic solvent, Probe Ultrasonic Searching 5～50min, and deionized water is dialysed, and obtains final product composite nano-granule suspension；
Wherein said organic solvent be methanol, ethanol, acetonitrile, acetone, ether, dichloromethane, chloroform one or two
Mixed solvent；
(2) Freeze Drying Technique：Composite nano-granule suspension is added freeze drying protectant, lyophilization, obtains final product composite nano-granule
Lyophilized powder；
Described composite nano-granule freeze drying protectant is selected from one of Mannitol, glucose, Lactose, sucrose or trehalose or several
Kind.
4. according to claim 3 hyaluronic acid-antineoplastic agent conjugate composite nano-granule compositionss it is characterised in that described
Bright matter acid-antineoplastic agent conjugate assembles single nanoparticle with amphipathic ester copolymer or is assembled into composite nano-granule, and it faces
On bed, acceptable route of administration has drug administration by injection, oral administration or mucosa delivery；Dosage form be injection, tablet, capsule or
Powder spray.
5. according to claim 4 hyaluronic acid-antineoplastic agent conjugate composite nano-granule compositionss it is characterised in that should
Nanoparticle or composite nano-granule compositionss can be used for treating nonsmall-cell lung cancer, breast carcinoma, ovarian cancer, cervical cancer, gastric cancer, liver
Cancer, leukemic single or drug combination.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310186800.2A CN103751795B (en) | 2013-05-20 | 2013-05-20 | Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310186800.2A CN103751795B (en) | 2013-05-20 | 2013-05-20 | Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103751795A CN103751795A (en) | 2014-04-30 |
| CN103751795B true CN103751795B (en) | 2017-02-08 |
Family
ID=50519216
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310186800.2A Expired - Fee Related CN103751795B (en) | 2013-05-20 | 2013-05-20 | Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103751795B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107375199A (en) * | 2017-09-07 | 2017-11-24 | 中国药科大学 | A kind of nanogel delivery system for polymerizeing chloroquine and preparation method thereof |
Families Citing this family (37)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104013960B (en) * | 2014-05-17 | 2016-08-17 | 广西师范大学 | A kind of targeting photo-thermal therapy water-soluble compound and preparation method and application |
| CN104056275B (en) * | 2014-05-30 | 2017-09-19 | 中国药科大学 | Multi-functional active targeting hyaluronic acid PLA carrier synthesis and its antineoplastic micella preparation method |
| CN104306983A (en) * | 2014-10-31 | 2015-01-28 | 山东大学 | Redox-sensitive hyaluronic acid-docetaxel conjugate and preparation method thereof |
| CN104497167B (en) * | 2014-11-24 | 2017-05-17 | 山东省药学科学院 | Hyaluronic acid derivative and therapeutic thereof |
| CN104644560A (en) * | 2015-03-05 | 2015-05-27 | 东华大学 | Hyaluronic acid modified hectorite amide nanoparticle and preparation and application of hyaluronic acid modified hectorite amide nanoparticle |
| CN105153428B (en) * | 2015-07-02 | 2018-02-16 | 天津大学 | A kind of pH response macromolecule micelles for mucus infiltration and preparation method thereof |
| CN106924731A (en) * | 2015-12-30 | 2017-07-07 | 复旦大学 | One kind is based on PDT and chemotherapeutic combination tumor targeted therapy system |
| CN105664174B (en) * | 2016-02-16 | 2019-06-18 | 中山大学孙逸仙纪念医院 | Lf-HA-DOX macromolecular prodrug compound and its construction method and the application in treatment glioma |
| CN106727314B (en) * | 2017-01-13 | 2021-01-01 | 中国药科大学 | Preparation and characterization of photosensitizer/anti-tumor drug self-assembly nano drug delivery system based on hyaluronic acid |
| CN107115296B (en) * | 2017-07-05 | 2019-08-02 | 扬州大学 | A kind of preparation method of intermediate hydrophobic drug substance stable nano suspension |
| CN108186571B (en) * | 2018-02-09 | 2020-07-14 | 苏州大学 | Application of reversible cross-linked asymmetric vesicle in preparation of acute leukemia treatment drug |
| CN108542895A (en) * | 2018-05-09 | 2018-09-18 | 中国药科大学 | Hyaluronic acid-curcumin-polyethylene glycol carrier preparation method and applications |
| CN110960506A (en) * | 2018-09-28 | 2020-04-07 | 中山大学 | Nanosuppreparation for the treatment of diseases caused by mitochondrial dysfunction |
| CN109678987B (en) * | 2018-12-03 | 2020-12-25 | 河南大学 | Hydrogen sulfide releasing agent HA-ADT, preparation method and application thereof |
| CN109893662B (en) * | 2019-03-13 | 2022-03-15 | 苏州大学 | Preparation method and application of prodrug-carrying brain metastasis targeted drug delivery system for inhibiting Mfsd2a |
| CN110124054B (en) * | 2019-06-21 | 2022-02-18 | 天津科技大学 | Preparation method and application of targeted nano particles self-assembled layer by layer |
| CN110354095A (en) * | 2019-07-01 | 2019-10-22 | 东南大学 | Target the pH sensitive transparent matter acid nanoparticle and preparation method thereof of atherosclerosis |
| CN110917360B (en) * | 2019-11-27 | 2021-04-27 | 武汉理工大学 | Preparation method and application of adriamycin-methotrexate combined drug delivery nano delivery system |
| CN111467500B (en) * | 2020-03-23 | 2022-08-09 | 北京工业大学 | Low-oxygen dual-targeting AGT inhibitor conjugate and preparation method and application thereof |
| CN111870616A (en) * | 2020-05-15 | 2020-11-03 | 吴宏伟 | Composite nano particle for targeted inhibition of bone tumor and preparation method thereof |
| CN111529486A (en) * | 2020-05-20 | 2020-08-14 | 郑州大学 | Preparation method and application of dissociable nano micelle based on pH/MMP response |
| CN111892668B (en) * | 2020-07-03 | 2022-07-12 | 广东工业大学 | Compound, preparation method thereof, fluorescent probe and antitumor drug |
| CN115867319B (en) * | 2020-07-15 | 2024-01-02 | 上海椿安生物医药科技有限公司 | Drug delivery system for local delivery of therapeutic agents and uses thereof |
| CN112263547B (en) * | 2020-11-26 | 2022-12-20 | 深圳大学 | Multifunctional nano-drug carrier, preparation method thereof and drug-loaded composition |
| CN112516310B (en) * | 2020-12-11 | 2022-11-29 | 武汉理工大学 | Preparation method and application of nano prodrug with response to tumor acid environment |
| CN112641760B (en) * | 2021-01-08 | 2022-04-26 | 东南大学 | Ferrocene-berberine/indometacin @ glucose oxidase @ hyaluronic acid nano-drug, preparation method and application |
| CN113087820B (en) * | 2021-04-12 | 2022-03-08 | 四川大学 | Dendrimer-modified hyaluronic acid-docetaxel conjugate and preparation method thereof |
| CN113045760B (en) * | 2021-04-12 | 2022-03-25 | 华南理工大学 | Polymer brush based on hyaluronic acid and alpha-D-galactose and preparation method and application thereof |
| CN113171342A (en) * | 2021-05-18 | 2021-07-27 | 天津科技大学 | Tumor-targeted nano micelle based on hyaluronic acid and preparation and application thereof |
| CN114904012B (en) * | 2021-07-22 | 2023-12-15 | 四川大学华西医院 | Active oxygen self-supplementing amphiphilic block copolymer-drug conjugate, preparation method and application thereof |
| CN114177302B (en) * | 2021-11-12 | 2023-07-18 | 中山大学 | Glycosaminoglycan modified substance for antitumor drug delivery, and preparation method and application thereof |
| CN114569736A (en) * | 2021-12-17 | 2022-06-03 | 兰州大学 | Cis-aconitic anhydride bond adriamycin/human serum albumin (CAD/HSA) composition and preparation method and application thereof |
| CN114522219B (en) * | 2022-02-23 | 2024-03-26 | 宜春学院 | Co-delivery polymer prodrug, and preparation method and application thereof |
| CN114870031B (en) * | 2022-05-20 | 2023-07-07 | 四川大学 | CD44 targeted taxane nanocrystal and preparation method and application thereof |
| CN115581660B (en) * | 2022-10-08 | 2024-02-27 | 中国药科大学 | Microneedle anti-tumor composite drug delivery system and preparation method and application thereof |
| CN115487171A (en) * | 2022-11-10 | 2022-12-20 | 郑州大学 | Hyaluronic acid-isoliquiritigenin conjugate, dissolving type microneedle patch, preparation method and application |
| CN116763725B (en) * | 2023-08-16 | 2023-11-24 | 四川大学华西医院 | Intelligent response type injectable hydrogel and preparation method and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1421201A (en) * | 2002-12-16 | 2003-06-04 | 天津大学 | Taxane injecta preparation |
-
2013
- 2013-05-20 CN CN201310186800.2A patent/CN103751795B/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1421201A (en) * | 2002-12-16 | 2003-06-04 | 天津大学 | Taxane injecta preparation |
Non-Patent Citations (1)
| Title |
|---|
| 《Hyaluronic Acid-Paclitaxel Conjugate Micelles：Synthesis，Characterization，and Antitumor Activity》;Hyukjin et al.;《Bioconjugate chem.》;20080516;第19卷(第6期);全文 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107375199A (en) * | 2017-09-07 | 2017-11-24 | 中国药科大学 | A kind of nanogel delivery system for polymerizeing chloroquine and preparation method thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103751795A (en) | 2014-04-30 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103751795B (en) | Preparation and application of hyaluronic acid-antitumor drug conjugate and composite nanoparticle composition | |
| CN101791411B (en) | Preparation and application of amphiphilic polysaccharide conjugate and medicinal compositions thereof | |
| CN105727309B (en) | The preparation and application of sensitive amphiphilic polysaccharide-adriamycin conjugate and its pharmaceutical compositions | |
| Zhang et al. | Tumor microenvironment-labile polymer–doxorubicin conjugate thermogel combined with docetaxel for in situ synergistic chemotherapy of hepatoma | |
| Zhang et al. | Dendrimer–doxorubicin conjugate as enzyme-sensitive and polymeric nanoscale drug delivery vehicle for ovarian cancer therapy | |
| CN102988999B (en) | Curcumin-polysaccharide conjugate as well as preparation method and application thereof | |
| CN102060991B (en) | Amphiphilic prodrug of 7- ethyl-10-hydroxycamptothecin and preparation method thereof | |
| Wang et al. | Norcantharidin-associated galactosylated chitosan nanoparticles for hepatocyte-targeted delivery | |
| US8466127B2 (en) | Pegylated and fatty acid grafted chitosan oligosaccharide, synthesis method and application for drug delivery system | |
| CN102120036B (en) | Nano micelle of biodegradable macromolecular-bonding Pt(IV) anti-cancer medicament and preparation method thereof | |
| Li et al. | Effective deactivation of A549 tumor cells in vitro and in vivo by RGD-decorated chitosan-functionalized single-walled carbon nanotube loading docetaxel | |
| CN104971353B (en) | The preparation and application of the amphiphilic polysaccharide derivative carrier and its pharmaceutical compositions of target tumor new vessels | |
| CN103301472A (en) | Amphiphilic polysaccharide-anti-tumor medicament conjugate capable of releasing medicines specifically at lesion site of living body, as well as preparation method and application of medicinal composition of amphiphilic polysaccharide-anti-tumor medicament conjugate | |
| CN103804472B (en) | A kind of taxone precursor | |
| US20100093935A1 (en) | Method of preparing polyglutamate conjugates | |
| CN103705943A (en) | Preparation method and application of reduction-response-type pegylation (PEG) nanomedicine composition | |
| CN105860057B (en) | Biodegradable polymer based on the hydrophilic polyaminoacid of hydrophobic function small molecule and its preparation method and application | |
| CN107952082A (en) | A kind of multi-functional synergistic pharmaceutical combination and its construction method based on adriamycin | |
| EP3735989A1 (en) | Novel cationic polyphosphazene compound, polyphosphazenes-drug conjugate compound and method for preparing same | |
| CN1879889A (en) | Kidney-targeted medicine vector and the formed prodrug, preparation method and uses | |
| Zhong et al. | Redox-responsive self-assembled polymeric nanoprodrug for delivery of gemcitabine in B-cell lymphoma therapy | |
| CN106176602A (en) | A kind of targeting is in the Docetaxel chitosan nano-micelle of stomach organization and preparation method and application | |
| CN103083682B (en) | Folic acid modified chitosan quaternary ammonium salt-taxol polymer medicine, as well as preparation method and application thereof | |
| CN112999159A (en) | HA-mediated targeted double-drug-loading cationic liposome coating and preparation method thereof | |
| Li et al. | Polysaccharides: A carrier for gene therapy |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20170208 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |