WO2016114355A1 - Composition for preventing adhesion - Google Patents
Composition for preventing adhesion Download PDFInfo
- Publication number
- WO2016114355A1 WO2016114355A1 PCT/JP2016/050998 JP2016050998W WO2016114355A1 WO 2016114355 A1 WO2016114355 A1 WO 2016114355A1 JP 2016050998 W JP2016050998 W JP 2016050998W WO 2016114355 A1 WO2016114355 A1 WO 2016114355A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- adhesion
- alginic acid
- composition
- metal salt
- monovalent metal
- Prior art date
Links
- 239000000203 mixture Substances 0.000 title claims abstract description 143
- 230000003405 preventing effect Effects 0.000 title claims abstract description 17
- 235000010443 alginic acid Nutrition 0.000 claims abstract description 136
- 229920000615 alginic acid Polymers 0.000 claims abstract description 136
- 150000003839 salts Chemical class 0.000 claims abstract description 129
- 150000004781 alginic acids Chemical class 0.000 claims abstract description 111
- 229960001126 alginic acid Drugs 0.000 claims abstract description 110
- 239000000783 alginic acid Substances 0.000 claims abstract description 110
- 229920002134 Carboxymethyl cellulose Polymers 0.000 claims abstract description 106
- 239000001768 carboxy methyl cellulose Substances 0.000 claims abstract description 105
- 235000010948 carboxy methyl cellulose Nutrition 0.000 claims abstract description 102
- 239000008112 carboxymethyl-cellulose Substances 0.000 claims abstract description 102
- 239000002158 endotoxin Substances 0.000 claims abstract description 80
- 229910052751 metal Inorganic materials 0.000 claims abstract description 75
- 239000002184 metal Substances 0.000 claims abstract description 75
- 238000000034 method Methods 0.000 claims description 52
- 210000004027 cell Anatomy 0.000 claims description 42
- 239000003795 chemical substances by application Substances 0.000 claims description 28
- 230000002265 prevention Effects 0.000 claims description 22
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 claims description 19
- 235000010413 sodium alginate Nutrition 0.000 claims description 19
- 239000000661 sodium alginate Substances 0.000 claims description 19
- 229940005550 sodium alginate Drugs 0.000 claims description 19
- 210000002901 mesenchymal stem cell Anatomy 0.000 claims description 18
- 239000000843 powder Substances 0.000 claims description 12
- 239000007788 liquid Substances 0.000 claims description 10
- 239000007787 solid Substances 0.000 claims description 10
- MZYRDLHIWXQJCQ-YZOKENDUSA-L potassium alginate Chemical compound [K+].[K+].O1[C@@H](C([O-])=O)[C@@H](OC)[C@H](O)[C@H](O)[C@@H]1O[C@@H]1[C@@H](C([O-])=O)O[C@@H](O)[C@@H](O)[C@H]1O MZYRDLHIWXQJCQ-YZOKENDUSA-L 0.000 claims description 6
- 239000012530 fluid Substances 0.000 claims description 5
- 235000010408 potassium alginate Nutrition 0.000 claims description 5
- 239000000737 potassium alginate Substances 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 4
- 239000002562 thickening agent Substances 0.000 claims description 3
- 230000000694 effects Effects 0.000 abstract description 11
- 239000000243 solution Substances 0.000 description 37
- 239000000499 gel Substances 0.000 description 28
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 description 26
- 229940072056 alginate Drugs 0.000 description 26
- 210000001519 tissue Anatomy 0.000 description 26
- 238000002156 mixing Methods 0.000 description 21
- 239000000463 material Substances 0.000 description 18
- 238000005259 measurement Methods 0.000 description 16
- 238000001356 surgical procedure Methods 0.000 description 15
- 241000700159 Rattus Species 0.000 description 13
- 206010061218 Inflammation Diseases 0.000 description 10
- 230000015572 biosynthetic process Effects 0.000 description 10
- 238000011156 evaluation Methods 0.000 description 10
- 230000004054 inflammatory process Effects 0.000 description 10
- 230000003068 static effect Effects 0.000 description 10
- 210000004185 liver Anatomy 0.000 description 9
- 239000000126 substance Substances 0.000 description 9
- 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 description 8
- 238000012752 Hepatectomy Methods 0.000 description 8
- 210000003815 abdominal wall Anatomy 0.000 description 8
- 230000037396 body weight Effects 0.000 description 8
- 229920002674 hyaluronan Polymers 0.000 description 8
- 229960003160 hyaluronic acid Drugs 0.000 description 8
- 238000000926 separation method Methods 0.000 description 8
- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 description 8
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 7
- 230000002440 hepatic effect Effects 0.000 description 7
- 208000014674 injury Diseases 0.000 description 7
- 238000000569 multi-angle light scattering Methods 0.000 description 7
- 238000012753 partial hepatectomy Methods 0.000 description 7
- 239000002953 phosphate buffered saline Substances 0.000 description 7
- 229920000642 polymer Polymers 0.000 description 7
- -1 polytetrafluoroethylene Polymers 0.000 description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 7
- 241000199919 Phaeophyceae Species 0.000 description 6
- 210000001015 abdomen Anatomy 0.000 description 6
- 238000001641 gel filtration chromatography Methods 0.000 description 6
- 239000002504 physiological saline solution Substances 0.000 description 6
- 230000008733 trauma Effects 0.000 description 6
- 208000034628 Celiac artery compression syndrome Diseases 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 5
- 230000000181 anti-adherent effect Effects 0.000 description 5
- 239000007864 aqueous solution Substances 0.000 description 5
- 210000001185 bone marrow Anatomy 0.000 description 5
- 239000003153 chemical reaction reagent Substances 0.000 description 5
- 238000009826 distribution Methods 0.000 description 5
- 238000002271 resection Methods 0.000 description 5
- 239000011734 sodium Substances 0.000 description 5
- 239000002904 solvent Substances 0.000 description 5
- IAJILQKETJEXLJ-SQOUGZDYSA-N L-guluronic acid Chemical compound O=C[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O IAJILQKETJEXLJ-SQOUGZDYSA-N 0.000 description 4
- 241001512709 Lessonia <stramenopiles> Species 0.000 description 4
- 241001465754 Metazoa Species 0.000 description 4
- 229920001218 Pullulan Polymers 0.000 description 4
- 239000004373 Pullulan Substances 0.000 description 4
- 230000004888 barrier function Effects 0.000 description 4
- AEMOLEFTQBMNLQ-UHFFFAOYSA-N beta-D-galactopyranuronic acid Natural products OC1OC(C(O)=O)C(O)C(O)C1O AEMOLEFTQBMNLQ-UHFFFAOYSA-N 0.000 description 4
- AEMOLEFTQBMNLQ-YBSDWZGDSA-N d-mannuronic acid Chemical compound O[C@@H]1O[C@@H](C(O)=O)[C@H](O)[C@@H](O)[C@H]1O AEMOLEFTQBMNLQ-YBSDWZGDSA-N 0.000 description 4
- 239000003480 eluent Substances 0.000 description 4
- 210000000416 exudates and transudate Anatomy 0.000 description 4
- 238000005227 gel permeation chromatography Methods 0.000 description 4
- 238000002347 injection Methods 0.000 description 4
- 239000007924 injection Substances 0.000 description 4
- XEEYBQQBJWHFJM-UHFFFAOYSA-N iron Substances [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 4
- 238000000691 measurement method Methods 0.000 description 4
- 230000036961 partial effect Effects 0.000 description 4
- WEXRUCMBJFQVBZ-UHFFFAOYSA-N pentobarbital Chemical compound CCCC(C)C1(CC)C(=O)NC(=O)NC1=O WEXRUCMBJFQVBZ-UHFFFAOYSA-N 0.000 description 4
- 229920001282 polysaccharide Polymers 0.000 description 4
- 239000005017 polysaccharide Substances 0.000 description 4
- 150000004804 polysaccharides Chemical class 0.000 description 4
- 235000019423 pullulan Nutrition 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 235000010344 sodium nitrate Nutrition 0.000 description 4
- 239000004317 sodium nitrate Substances 0.000 description 4
- 241001474374 Blennius Species 0.000 description 3
- 241001466453 Laminaria Species 0.000 description 3
- 241000239218 Limulus Species 0.000 description 3
- 241001491708 Macrocystis Species 0.000 description 3
- 229920002201 Oxidized cellulose Polymers 0.000 description 3
- 210000000683 abdominal cavity Anatomy 0.000 description 3
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 3
- 125000003277 amino group Chemical group 0.000 description 3
- 239000003431 cross linking reagent Substances 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 238000001035 drying Methods 0.000 description 3
- 150000002500 ions Chemical class 0.000 description 3
- 229910052742 iron Inorganic materials 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 229910021645 metal ion Inorganic materials 0.000 description 3
- 239000003960 organic solvent Substances 0.000 description 3
- 229940107304 oxidized cellulose Drugs 0.000 description 3
- 210000004303 peritoneum Anatomy 0.000 description 3
- 229920001343 polytetrafluoroethylene Polymers 0.000 description 3
- 239000004810 polytetrafluoroethylene Substances 0.000 description 3
- 230000002829 reductive effect Effects 0.000 description 3
- 238000013077 scoring method Methods 0.000 description 3
- 235000019812 sodium carboxymethyl cellulose Nutrition 0.000 description 3
- 229920001027 sodium carboxymethylcellulose Polymers 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 210000002536 stromal cell Anatomy 0.000 description 3
- RZVAJINKPMORJF-UHFFFAOYSA-N Acetaminophen Chemical compound CC(=O)NC1=CC=C(O)C=C1 RZVAJINKPMORJF-UHFFFAOYSA-N 0.000 description 2
- 206010002091 Anaesthesia Diseases 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 108010073385 Fibrin Proteins 0.000 description 2
- 102000009123 Fibrin Human genes 0.000 description 2
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 2
- 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 2
- 241000776474 Patescibacteria group Species 0.000 description 2
- QGMRQYFBGABWDR-UHFFFAOYSA-M Pentobarbital sodium Chemical compound [Na+].CCCC(C)C1(CC)C(=O)NC(=O)[N-]C1=O QGMRQYFBGABWDR-UHFFFAOYSA-M 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 239000000853 adhesive Substances 0.000 description 2
- 230000001070 adhesive effect Effects 0.000 description 2
- 210000000577 adipose tissue Anatomy 0.000 description 2
- 230000037005 anaesthesia Effects 0.000 description 2
- 229920001222 biopolymer Polymers 0.000 description 2
- 244000309464 bull Species 0.000 description 2
- 238000011088 calibration curve Methods 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 238000010828 elution Methods 0.000 description 2
- 210000004700 fetal blood Anatomy 0.000 description 2
- 229950003499 fibrin Drugs 0.000 description 2
- 239000000945 filler Substances 0.000 description 2
- 238000012812 general test Methods 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 230000002439 hemostatic effect Effects 0.000 description 2
- 229920001519 homopolymer Polymers 0.000 description 2
- 239000000017 hydrogel Substances 0.000 description 2
- 238000007912 intraperitoneal administration Methods 0.000 description 2
- 238000002350 laparotomy Methods 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 239000008176 lyophilized powder Substances 0.000 description 2
- 125000001288 lysyl group Chemical group 0.000 description 2
- 229930014626 natural product Natural products 0.000 description 2
- 231100000989 no adverse effect Toxicity 0.000 description 2
- 210000002747 omentum Anatomy 0.000 description 2
- 229960001412 pentobarbital Drugs 0.000 description 2
- 230000000704 physical effect Effects 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 239000011347 resin Substances 0.000 description 2
- 229920005989 resin Polymers 0.000 description 2
- 239000012266 salt solution Substances 0.000 description 2
- 238000001542 size-exclusion chromatography Methods 0.000 description 2
- 210000000813 small intestine Anatomy 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
- 159000000000 sodium salts Chemical class 0.000 description 2
- 239000011343 solid material Substances 0.000 description 2
- 238000001179 sorption measurement Methods 0.000 description 2
- 210000000130 stem cell Anatomy 0.000 description 2
- 239000011550 stock solution Substances 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 239000008215 water for injection Substances 0.000 description 2
- 230000004584 weight gain Effects 0.000 description 2
- 235000019786 weight gain Nutrition 0.000 description 2
- 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 1
- APKFDSVGJQXUKY-KKGHZKTASA-N Amphotericin-B Natural products O[C@H]1[C@@H](N)[C@H](O)[C@@H](C)O[C@H]1O[C@H]1C=CC=CC=CC=CC=CC=CC=C[C@H](C)[C@@H](O)[C@@H](C)[C@H](C)OC(=O)C[C@H](O)C[C@H](O)CC[C@@H](O)[C@H](O)C[C@H](O)C[C@](O)(C[C@H](O)[C@H]2C(O)=O)O[C@H]2C1 APKFDSVGJQXUKY-KKGHZKTASA-N 0.000 description 1
- 241000512260 Ascophyllum Species 0.000 description 1
- BSYNRYMUTXBXSQ-UHFFFAOYSA-N Aspirin Chemical compound CC(=O)OC1=CC=CC=C1C(O)=O BSYNRYMUTXBXSQ-UHFFFAOYSA-N 0.000 description 1
- 229920002498 Beta-glucan Polymers 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 241000282472 Canis lupus familiaris Species 0.000 description 1
- 241000700198 Cavia Species 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 241000699800 Cricetinae Species 0.000 description 1
- 241001512723 Ecklonia Species 0.000 description 1
- 241000230129 Eisenia <Phaeophyceae> Species 0.000 description 1
- 241000283086 Equidae Species 0.000 description 1
- PIICEJLVQHRZGT-UHFFFAOYSA-N Ethylenediamine Chemical compound NCCN PIICEJLVQHRZGT-UHFFFAOYSA-N 0.000 description 1
- 101150021185 FGF gene Proteins 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 229920002148 Gellan gum Polymers 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
- 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 description 1
- 102100021866 Hepatocyte growth factor Human genes 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101000898034 Homo sapiens Hepatocyte growth factor Proteins 0.000 description 1
- 101001076408 Homo sapiens Interleukin-6 Proteins 0.000 description 1
- 101000868152 Homo sapiens Son of sevenless homolog 1 Proteins 0.000 description 1
- 102000004218 Insulin-Like Growth Factor I Human genes 0.000 description 1
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 description 1
- 101000844802 Lacticaseibacillus rhamnosus Teichoic acid D-alanyltransferase Proteins 0.000 description 1
- 241001260563 Lessonia nigrescens Species 0.000 description 1
- 239000012901 Milli-Q water Substances 0.000 description 1
- 101100496858 Mus musculus Colec12 gene Proteins 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 229910000528 Na alloy Inorganic materials 0.000 description 1
- 229930193140 Neomycin Natural products 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 208000000450 Pelvic Pain Diseases 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 206010034650 Peritoneal adhesions Diseases 0.000 description 1
- 206010060932 Postoperative adhesion Diseases 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- 229920002385 Sodium hyaluronate Polymers 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 208000031737 Tissue Adhesions Diseases 0.000 description 1
- 102000004887 Transforming Growth Factor beta Human genes 0.000 description 1
- 108090001012 Transforming Growth Factor beta Proteins 0.000 description 1
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 description 1
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 description 1
- 241000947853 Vibrionales Species 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 229960001138 acetylsalicylic acid Drugs 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 229910052783 alkali metal Inorganic materials 0.000 description 1
- 229910052784 alkaline earth metal Inorganic materials 0.000 description 1
- 229960004821 amikacin Drugs 0.000 description 1
- LKCWBDHBTVXHDL-RMDFUYIESA-N amikacin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O1)O)NC(=O)[C@@H](O)CCN)[C@H]1O[C@H](CN)[C@@H](O)[C@H](O)[C@H]1O LKCWBDHBTVXHDL-RMDFUYIESA-N 0.000 description 1
- APKFDSVGJQXUKY-INPOYWNPSA-N amphotericin B Chemical compound O[C@H]1[C@@H](N)[C@H](O)[C@@H](C)O[C@H]1O[C@H]1/C=C/C=C/C=C/C=C/C=C/C=C/C=C/[C@H](C)[C@@H](O)[C@@H](C)[C@H](C)OC(=O)C[C@H](O)C[C@H](O)CC[C@@H](O)[C@H](O)C[C@H](O)C[C@](O)(C[C@H](O)[C@H]2C(O)=O)O[C@H]2C1 APKFDSVGJQXUKY-INPOYWNPSA-N 0.000 description 1
- 229960003942 amphotericin b Drugs 0.000 description 1
- 229940035676 analgesics Drugs 0.000 description 1
- 239000000730 antalgic agent Substances 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940124599 anti-inflammatory drug Drugs 0.000 description 1
- 230000001754 anti-pyretic effect Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 239000002221 antipyretic Substances 0.000 description 1
- 238000000149 argon plasma sintering Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 229920002988 biodegradable polymer Polymers 0.000 description 1
- 239000004621 biodegradable polymer Substances 0.000 description 1
- BBWBEZAMXFGUGK-UHFFFAOYSA-N bis(dodecylsulfanyl)-methylarsane Chemical compound CCCCCCCCCCCCS[As](C)SCCCCCCCCCCCC BBWBEZAMXFGUGK-UHFFFAOYSA-N 0.000 description 1
- 229920001400 block copolymer Polymers 0.000 description 1
- 210000004271 bone marrow stromal cell Anatomy 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 159000000007 calcium salts Chemical class 0.000 description 1
- 150000001735 carboxylic acids Chemical class 0.000 description 1
- 229940105329 carboxymethylcellulose Drugs 0.000 description 1
- 150000001768 cations Chemical class 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000003889 chemical engineering Methods 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 210000001072 colon Anatomy 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 210000001198 duodenum Anatomy 0.000 description 1
- 238000011013 endotoxin removal Methods 0.000 description 1
- 230000007717 exclusion Effects 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 238000002523 gelfiltration Methods 0.000 description 1
- 235000010492 gellan gum Nutrition 0.000 description 1
- 239000000216 gellan gum Substances 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 1
- 229920001477 hydrophilic polymer Polymers 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 210000003405 ileum Anatomy 0.000 description 1
- 208000000509 infertility Diseases 0.000 description 1
- 230000036512 infertility Effects 0.000 description 1
- 231100000535 infertility Toxicity 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- 210000001630 jejunum Anatomy 0.000 description 1
- 238000002356 laser light scattering Methods 0.000 description 1
- 229910003002 lithium salt Inorganic materials 0.000 description 1
- 159000000002 lithium salts Chemical class 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 238000002483 medication Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 229960004927 neomycin Drugs 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 125000004433 nitrogen atom Chemical group N* 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 210000000496 pancreas Anatomy 0.000 description 1
- 229960005489 paracetamol Drugs 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 210000003200 peritoneal cavity Anatomy 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 239000004848 polyfunctional curative Substances 0.000 description 1
- 239000002861 polymer material Substances 0.000 description 1
- 229920000193 polymethacrylate Polymers 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
- XAEFZNCEHLXOMS-UHFFFAOYSA-M potassium benzoate Chemical compound [K+].[O-]C(=O)C1=CC=CC=C1 XAEFZNCEHLXOMS-UHFFFAOYSA-M 0.000 description 1
- 238000007781 pre-processing Methods 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- ZNZJJSYHZBXQSM-UHFFFAOYSA-N propane-2,2-diamine Chemical compound CC(C)(N)N ZNZJJSYHZBXQSM-UHFFFAOYSA-N 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000008213 purified water Substances 0.000 description 1
- 238000010188 recombinant method Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 229940010747 sodium hyaluronate Drugs 0.000 description 1
- 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 description 1
- MSXHSNHNTORCAW-MPGIDXPLSA-M sodium;(3s,4s,5s,6r)-3,4,5,6-tetrahydroxyoxane-2-carboxylate Chemical compound [Na+].O[C@@H]1OC(C([O-])=O)[C@@H](O)[C@H](O)[C@@H]1O MSXHSNHNTORCAW-MPGIDXPLSA-M 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 230000003637 steroidlike Effects 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 229920001059 synthetic polymer Polymers 0.000 description 1
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 description 1
- 229960000707 tobramycin Drugs 0.000 description 1
- NLVFBUXFDBBNBW-PBSUHMDJSA-N tobramycin Chemical compound N[C@@H]1C[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N NLVFBUXFDBBNBW-PBSUHMDJSA-N 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/14—Blood; Artificial blood
- A61K35/15—Cells of the myeloid line, e.g. granulocytes, basophils, eosinophils, neutrophils, leucocytes, monocytes, macrophages or mast cells; Myeloid precursor cells; Antigen-presenting cells, e.g. dendritic cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
- A61K47/38—Cellulose; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/08—Solutions
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/70—Web, sheet or filament bases ; Films; Fibres of the matrix type containing drug
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
Definitions
- the present invention relates to an adhesion preventing composition.
- Adhesion refers to a state where the surfaces of tissues that should be separated from each other are connected or fused with a fibrous tissue. Adhesion occurs when exudate containing fibrin is generated on the surface of the tissue due to trauma or inflammation, and the exudate is organized and the tissue surfaces are connected or fused. Injuries that occur on the surface of tissues during surgery, inflammation caused by the trauma, and inflammation caused by drying of the tissue surface during surgery cause adhesions.
- Adhesion can sometimes cause infertility, bowel passage problems and chronic pelvic pain.
- another surgical operation may be required in order to peel off the adhesion that has occurred after the surgical operation. For these reasons, it is necessary to prevent adhesion, and various measures have been taken to prevent adhesion.
- Some such means for preventing adhesions are to place a physical barrier between the site of trauma or inflammation and its adjacent tissue to prevent tissue connection or fusion.
- a physical barrier As such a physical barrier, a sheet form and a gel form are known.
- the sheet-like material is a polytetrafluoroethylene (PTFE) film (Prelude (trade name) (WL Gore and Associates, Inc.)), a sheet (Seprafilm (product) containing hyaluronic acid (HA) and carboxymethyl cellulose (CMC). Name) (Genzyme GmbH)), regenerated oxidized cellulose sheet (Intraceded (trade name) (Johnson & Johnson)) and the like.
- PTFE film Prelude (trade name) (WL Gore and Associates, Inc.)
- a sheet Seprafilm (product) containing hyaluronic acid (HA) and carboxymethyl cellulose (CMC). Name) (Genzyme GmbH)
- regenerated oxidized cellulose sheet (Intraceded (trade name) (Johnson & Johnson)) and the like.
- the PTFE film is not biodegradable and has a problem of remaining in the body.
- the sheet containing HA and CMC and the regenerated oxidized cellulose sheet are biodegradable, serious adhesion such as adhesion occurring after hepatectomy cannot be completely prevented, and the effect of preventing adhesion can be improved. There was room.
- gels examples include iron ion cross-linked hyaluronic acid gel (Intergel (trade name) (Lifecore Biomedical, LLC)).
- Intergel trade name
- iron ion-crosslinked hyaluronic acid gels are not currently used in clinical practice because they cause severe inflammation mainly due to an increase in the concentration of iron ions in the peritoneal cavity and the long-term remaining of the hydrogel.
- a gel containing alginic acid, HA, CMC or a mixture thereof as a physical barrier for preventing adhesions (Patent Documents 1 and 2, and Non-Patent Documents 1 to 3)
- the optimum physical properties capable of sufficiently exerting the adhesion preventing effect were not clear.
- the surface of the tissue associated with the surgery may be treated with a biocompatible and pharmacologically acceptable water-soluble polypeptide, It has been proposed to coat with an aqueous solution of a hydrophilic polymer material selected from the group consisting of sugars, synthetic polymers, salts and complexes thereof, and mixtures thereof to prevent tissue adhesion due to drying. (Patent Document 3).
- the present inventors have found that (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) an adhesion using a combination of cells.
- the inventors have found that the prevention composition has a high adhesion prevention effect and have completed the present invention.
- An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell.
- a monovalent metal salt of low endotoxin alginic acid comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell.
- the cells are mesenchymal stem cells.
- the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
- the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
- An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose and a pharmaceutically acceptable salt thereof, and (c) a cell not used.
- composition according to [12] The composition according to [11] above, wherein the composition is in a liquid form having fluidity and has a viscosity of 5 mPa ⁇ s to 100,000 mPa ⁇ s.
- a subject in need of anti-adhesion using a composition comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells.
- a method for preventing adhesions comprising applying to.
- An anti-adhesion method comprising applying the composition to a subject in need of anti-adhesion.
- An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells.
- An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof and (c) a cell-free (a) monovalent metal salt of low endotoxin alginic acid. object.
- An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent.
- An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent, used in combination with cells.
- An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof.
- An anti-adhesion kit comprising (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells, and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof. .
- an anti-adhesion composition having a high anti-adhesion effect can be provided.
- Adhesion refers to a condition in which the surfaces of tissues that should be separated from each other are connected or fused with a fibrous tissue.
- causes of adhesion include trauma that can occur on the surface of a tissue during surgery, inflammation caused by the trauma, inflammation due to dry tissue surface during surgery, and the like. With these trauma and inflammation, exudate containing fibrin is generated on the surface of the tissue, and this exudate becomes organized and the tissue surface is connected or fused to form an adhesion.
- Anti-adhesion refers to reducing the formation of adhesions.
- adhesion prevention may be rephrased as adhesion reduction. For example, it is sufficient that at least one selected from the frequency, range, and degree of adhesion is reduced. “Adhesion prevention” means that, for example, when the adhesion grade evaluation described in the Examples is performed, the average adhesion grade is higher than the average adhesion grade when the composition of the present invention is not applied. It only has to be low.
- “adhesion prevention” means, for example, when the extent evaluation of adhesion described in the Examples is performed, compared to the extent of average adhesion when the composition of the present invention is not applied, the extent of average adhesion Should be lower.
- “Anti-adhesion” is preferably prevention of adhesions resulting from surgery, more preferably peritoneal adhesions resulting from surgery. That is, “adhesion prevention” is preferably prevention of postoperative adhesion.
- composition A of the present invention may be referred to as (a) a monovalent metal salt of low endotoxin alginic acid (hereinafter referred to as “component (a)”). ), (B) carboxymethylcellulose or a pharmaceutically acceptable salt thereof (hereinafter sometimes referred to as “(b) component”), and (c) a cell (herein referred to as “(c) component”)
- component (a) a monovalent metal salt of low endotoxin alginic acid
- component (b) component) carboxymethylcellulose or a pharmaceutically acceptable salt thereof
- cell herein referred to as “(c) component”.
- a composition according to another aspect of the present invention (sometimes referred to herein as “the composition B of the present invention”) is: (a) a monovalent metal salt of low endotoxin alginic acid, and (b) carboxymethylcellulose or its An anti-adhesion composition that uses a combination of pharmaceutically acceptable salts and does not use (c) cells.
- a composition according to another aspect of the present invention (sometimes referred to herein as “composition C of the present invention”) comprises (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell. And (a) a composition for preventing adhesion containing a monovalent metal salt of low endotoxin alginic acid.
- composition D of the present invention The composition according to another aspect of the present invention (hereinafter, sometimes referred to as “the composition D of the present invention”) is used in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, And (c) a composition for preventing adhesion containing (a) a monovalent metal salt of low endotoxin alginic acid without using cells.
- the composition A of the present invention, the composition B of the present invention, the composition C of the present invention and the composition D of the present invention may be collectively referred to as “the composition of the present invention”.
- the present invention also relates to an adhesion prevention method comprising applying the composition of the present invention to a subject in need of adhesion prevention.
- the method of certain aspects of the invention comprises (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell.
- An anti-adhesion method comprising applying a composition to be used to a subject in need of anti-adhesion.
- the method of another aspect of the present invention uses a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (C) An anti-adhesion method comprising applying a cell-free composition to a subject in need of anti-adhesion.
- Another aspect of the present invention provides (a) a monovalent monovalent form of low endotoxin alginic acid for use in adhesion prevention in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells. It is a metal salt.
- Another aspect of the present invention provides (a) a low endotoxin alginic acid for use in the prevention of adhesion in combination with (c) a cell, and (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof. It is a monovalent metal salt.
- “Use in combination” means to use in combination.
- the component (a), the component (b), and the component (c) are used in combination, the component (a), the component (b), and the component (c) are included in the composition when the composition is applied to a target. It means that it should be.
- the component (a) and the component (b) are used in combination, and the component (c) is not used, the component (a) and the component (b) are included in the composition when the composition is applied to the object.
- the component (c) should not be contained in the composition. Therefore, the term “used in combination” does not particularly refer to the form in the distribution stage including sales.
- (b) component may contain the hardening
- component, (b) component and (c) component when used in combination, (1) (a) component, (b) component and (c) component are combined in advance, (2) (a A form in which component (b) and component (b) are pre-combined and component (c) are combined into a kit, or prepared separately without being combined, and used in a mixed form when applied (3) (a) Either of the two types of compositions obtained by separately preparing the component and the component (b) and the component (c) are combined into a kit, or prepared separately without being combined and used in combination when applied. But you can.
- the component (b) may be provided in the form of a composition previously blended with the component (a), and two types of compositions obtained by separately preparing the component (a) and the component (b). You may provide as a kit which combined and packaged.
- the component (b) may further contain a curing agent, and in addition to the components (a), (b), and (c), a combination of curing agents is used. Also good. The same applies to the following examples.
- component and (b) component when (a) component and (b) component are used in combination and (c) component is not used, (1) a form in which (a) component and (b) component are combined in advance, (2) (a ) Component and (b) component may be prepared separately, and two types of compositions obtained may be combined into a kit, or may be prepared separately without combining and used in combination. That is, the component (b) may be provided in the form of a composition previously blended with the component (a), and two types of compositions obtained by separately preparing the component (a) and the component (b). You may provide as a kit which combined and packaged.
- each of the component (a) and the component (b) or a mixture thereof may be provided in a solution state using a solvent, or may be provided as a solid material such as a lyophilized product (particularly a lyophilized powder). May be.
- the composition of the present invention may be used in a state having fluidity such as a solution or a gel using a solvent at the time of administration.
- the component (a) and the component (b) are each a solid or a mixture selected from the group consisting of a sheet, sponge, powder, or powder Semi-solid; or gel.
- the solvent is not particularly limited as long as it is a solvent applicable to a living body.
- water for injection purified water, distilled water, ion-exchanged water (or deionized water), milli-Q water, physiological saline, phosphate buffered physiological Examples include saline (PBS).
- PBS saline
- Preferred are water for injection, distilled water, physiological saline and the like that can be used for treatment of humans and animals.
- the content of the monovalent metal salt of alginic acid in the composition is about 0.1 wt% to 10 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof The amount is about 0.1 wt% to 10 wt%. More preferably, the content of the monovalent metal salt of alginic acid in the composition is about 1 wt% to 5 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof is about 1 wt% to 5 wt%. .
- the content of the monovalent metal salt of alginic acid in the composition is about 0.1 wt% to 99.9 wt%, and carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
- the acceptable salt content is about 0.1 wt% to 99.9 wt%. More preferably, the content of the monovalent metal salt of alginic acid in the composition is about 1 wt% to 99 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof is about 1 wt% to 99 wt%. .
- Subjects are human or non-human warm-blooded animals, such as birds, and non-human mammals such as cows, monkeys, cats, mice, rats, guinea pigs, hamsters, pigs, dogs, rabbits, sheep, horses.
- the method of applying the composition of the present invention to a subject is not particularly limited, and for example, it may be directly injected onto the surface of a tissue related to surgery with a syringe, a gel pipette, a dedicated filler, or the like.
- tissue associated with surgery is tissue that has been traumatically injured on the surface during surgery, or tissue that has become or may be inflamed due to dryness of the surface during surgery.
- the tissues associated with the surgery are organs encased in the peritoneum, such as the stomach, jejunum, ileum, appendix, colon, liver, spleen, duodenum, and pancreas. is there.
- Particularly preferred compositions of the present invention can effectively prevent serious adhesions such as adhesions that occur after hepatectomy.
- the form of the composition of the present invention is, for example, a fluid liquid form (ie, a solution form); a solid form such as a sheet form, a sponge form, a powder (particularly freeze-dried powder) or a powder form; Solid; or gel.
- “having fluidity” means having a property of changing its form into an indefinite form. For example, it is desirable to have fluidity so that the composition can be encapsulated in a syringe or the like and injected into the surface of tissue associated with the surgical procedure.
- composition of the present invention that is in the form of a gel or solution can be easily applied to the surface of a tissue with a syringe, a pipette for gel, a dedicated syringe or the like.
- the powder and the powdery composition can also be applied to the tissue surface with a spray device or the like.
- Sheet-like, sponge-like, powder, powder-like, gel-like, and fluid liquid compositions are suitable for any surface shape and are applied to the entire surface of the tissue to which the composition of the present invention is applied. You can also touch.
- the sheet form is a flat plate having a suitable thickness and flexibility, and the sponge form is processed into a porous state.
- composition can also be made into a semi-solid state with a freezing or hardening
- the gel can be prepared by covalently bonding the composition with a polyvalent cation or a cross-linking reagent as a curing agent (cross-linking agent).
- Monovalent metal salt of alginic acid is a water-soluble substance formed by ion exchange of the hydrogen atom of the 6-position carboxylic acid of alginic acid with monovalent metal ions such as Na + and K + . It is salt.
- Specific examples of the monovalent metal salt of alginic acid include sodium alginate and potassium alginate. In particular, sodium alginate that is commercially available is preferable.
- a solution of a monovalent metal salt of alginic acid forms a gel when mixed with a crosslinking agent.
- Alginic acid used in the present invention is a biodegradable polymer polysaccharide, a polymer in which two types of uronic acids, D-mannuronic acid (M) and L-guluronic acid (G), are polymerized in a straight chain. It is. More specifically, D-mannuronic acid homopolymer fraction (MM fraction), L-guluronic acid homopolymer fraction (GG fraction), and D-mannuronic acid and L-guluronic acid are randomly arranged. This is a block copolymer in which the fractions (MG fraction) are bound arbitrarily.
- the composition ratio (M / G ratio) of D-mannuronic acid and L-guluronic acid of alginic acid varies depending on the type of organism mainly derived from seaweed, etc. It ranges from a high G type with an M / G ratio of about 0.4 to a high M type with an M / G ratio of about 5.
- the monovalent metal salt of alginic acid is a high molecular weight polysaccharide, and it is difficult to accurately determine the molecular weight, but generally the weight average molecular weight is 10 to 10 million, preferably 10,000 to 10 million, more preferably 2 It ranges from 10,000 to 3,000,000.
- a preferable range of the weight average molecular weight measured by gel permeation chromatography (GPC) method is 300,000 to 2,000,000.
- GPC-MALS method is 1,000 to 300,000, and a more preferable range is 50,000 to 300,000.
- a measurement error of 10 to 20% or more may occur.
- the value may vary in the range of about 32 to 480,000 for 400,000, 400,000 to 600,000 for 500,000, and about 800 to 1,200,000 for 1,000,000. Therefore, a suitable weight average molecular weight range for the monovalent metal salt of alginic acid is at least 20,000, more preferably 50,000, and even more preferably 100,000. If the molecular weight is too high, it is difficult to produce and causes problems such as excessively high viscosity when used as an aqueous solution, poor solubility, and difficulty in maintaining physical properties during long-term storage. Is preferably 5 million or less, more preferably 3 million or less.
- a polymer substance derived from a natural product does not have a single molecular weight but is an aggregate of molecules having various molecular weights, and thus is measured as a molecular weight distribution having a certain width.
- a typical measurement technique is gel filtration chromatography. Representative information on the molecular weight distribution obtained by gel filtration chromatography includes weight average molecular weight (Mw), number average molecular weight (Mn), and dispersion ratio (Mw / Mn).
- the weight average molecular weight places importance on the contribution to the average molecular weight of a polymer having a large molecular weight and is represented by the following formula.
- the number average molecular weight is calculated by dividing the total weight of the polymer by the total number of polymers.
- W is the total weight of the polymer
- Wi is the weight of the i-th polymer
- Mi is the molecular weight at the i-th elution time
- Ni is the number of molecular weights Mi
- Hi is the height at the i-th elution time.
- the molecular weight of a high molecular weight substance derived from a natural product may vary depending on the measurement method (examples of hyaluronic acid: Chikako YOMOTA et.al., Bull. Natl. Health Sci., Vol. 117, pp 135-139 (1999), Chikako YOMOTA et.al., Bull. Natl. Inst. Health Sci., Vol. 121, pp 30-33 (2003)).
- the molecular weight measurement of alginate there is a method of calculating from intrinsic viscosity (Intrinsic Viscosity), SEC-MALLS (Size Exclusion Chromatography with Multiple Laser Light Scattering, which is calculated by the Ft.
- the molecular weight of the alginate is specified, it is a weight average molecular weight calculated by gel filtration chromatography unless otherwise specified.
- Typical conditions for gel filtration chromatography include using a calibration curve with pullulan as a standard substance.
- the molecular weight of pullulan used as the standard substance is preferably at least 1.6 million, 788,000, 404,000, 212,000 and 112,000 as the standard substance.
- eluent 200 mM sodium nitrate solution
- column conditions it is preferable to use a polymethacrylate resin filler and use at least one to three columns having an exclusion limit molecular weight of 10 million or more.
- Typical columns are TSKgel GMPW x1 (diameter 7.8 mm ⁇ 300 mm) and G2500PW XL (diameter 7.8 mm ⁇ 300 mm) (manufactured by Tosoh Corporation).
- the monovalent metal salt of alginic acid initially has a large molecular weight and a high viscosity when extracted from brown algae, but the molecular weight becomes small and the viscosity becomes low in the process of drying by heat, lyophilization and purification. Therefore, monovalent metal salts of alginic acid having different molecular weights can be produced by performing appropriate temperature control in each production process.
- a monovalent metal salt of alginic acid having a large molecular weight can be obtained by controlling the temperature in each step of the production to be lower, and a monovalent metal salt of alginic acid having a smaller molecular weight can be obtained as the temperature is increased.
- the monovalent metal salt of alginic acid from which molecular weight differs can also be manufactured by the method of selecting suitably the brown algae used as a raw material, or performing the fractionation by molecular weight in a manufacturing process. Furthermore, the molecular weight or viscosity of the monovalent metal salt of alginic acid produced by each method is measured, and then mixed with another lot of monovalent metal salt of alginic acid having a different molecular weight or viscosity to have the target molecular weight. It is also possible to use a monovalent metal salt of alginic acid.
- the alginic acid used in the present invention may be naturally derived or synthetic, but is preferably naturally derived.
- naturally occurring alginic acid include those extracted from brown algae.
- brown alga containing alginic acid is nowadays in coastal areas around the world, seaweed that can actually be used as a raw material for alginic acid is limited, such as Lessonia in South America, Macrocystis in North America, Laminaria and Ascofilum in Europe, Typical examples are Daviglia.
- brown algae used as a raw material for alginic acid include, for example, the genus Lesonia, the genus Macrocystis, the genus Laminaria (comb), the genus Ascophyllum, the genus Durvillea, Examples include the genus Eisenia and the genus Ecklonia.
- Carboxymethylcellulose or a pharmaceutically acceptable salt thereof is carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
- the carboxymethyl cellulose or a pharmaceutically acceptable salt thereof used in the present invention is not particularly limited, but it is preferable to use a carboxymethyl cellulose having a weight average molecular weight of 10,000 to 1,500,000.
- a carboxymethyl cellulose having a weight average molecular weight of 10,000 to 1,500,000 about the measuring method of the weight average molecular weight of such carboxymethylcellulose or its pharmaceutically acceptable salt, the measuring method similar to the weight average molecular weight of the said monovalent metal salt of alginic acid can be utilized.
- Such carboxymethylcellulose or a pharmaceutically acceptable salt thereof can be produced by using various known methods, or a commercially available product can be purchased (for example, Sigma-Aldrich, Daiichi Kogyo Seiyaku Co., Ltd.).
- Examples of pharmaceutically acceptable salts of carboxymethyl cellulose include alkali metal salts such as sodium salt, potassium salt and lithium salt, and alkaline earth metal salts such as calcium salt.
- a particularly preferable salt is a sodium salt.
- composition A of the present invention uses component (c), ie, cells.
- composition B of the present invention does not use component (c), that is, cells.
- Examples of cells include stem cells and stromal cells, and the origin is not particularly limited, but examples thereof include bone marrow, adipose tissue, and cord blood.
- examples of cells include ES cells and iPS cells.
- Preferred examples of the cells include mesenchymal stem cells and mesenchymal stromal cells, and more preferred examples include bone marrow mesenchymal stem cells and bone marrow mesenchymal stromal cells.
- Using cells means collecting the target cells from bone marrow, adipose tissue, umbilical cord blood, etc., concentrating them, or culturing them to increase the amount, and adding the prepared cells to the composition of the present invention Say to do. Specifically, for example, 1 ⁇ 10 4 cells / ml or more, or 1 ⁇ 10 5 cells / ml or more, preferably 1 ⁇ 10 4 cells / ml to 1 ⁇ 10 7 cells / ml are used in the present invention. It is said to be contained in the composition.
- the composition of the present invention may contain a factor that promotes cell growth.
- factors include BMP, FGF, VEGF, HGF, TGF- ⁇ , IGF-1, PDGF, CDMP, CSF, EPO, IL, and IF. These factors may be produced by recombinant methods or purified from the protein composition. It should be noted that the compositions of some embodiments of the present invention do not contain these growth factors.
- composition of some embodiments of the present invention does not contain a curing agent of a monovalent metal salt of alginic acid.
- the form of the composition that does not contain a curing agent is, for example, a liquid having fluidity, a solid such as a lyophilized product (particularly lyophilized powder).
- the composition of some other embodiments of the present invention contains a curing agent of a monovalent metal salt of alginic acid.
- the form of the composition containing the curing agent is, for example, a freeze-dried product (particularly freeze-dried powder), a solid material such as a sheet or sponge; or a gel.
- the curing agent is immobilized by crosslinking a solution of a monovalent metal salt of alginic acid.
- the curing agent examples include divalent or higher valent metal ion compounds such as Ca 2+ , Mg 2+ , Ba 2+ , and Sr 2+, and crosslinkable reagents having 2 to 4 amino groups in the molecule. More specifically, CaCl 2 , MgCl 2 , CaSO 4 , BaCl 2 , SrCl 2, etc. (preferably CaCl 2 , CaSO 4 , BaCl 2, etc.) are used as the divalent or higher valent metal ion compound in the molecule.
- Diaminoalkanes that may have a lysyl group (—COCH (NH 2 ) — (CH 2 ) 4 —NH 2 ) on the nitrogen atom as crosslinkable reagents having ⁇ 4 amino groups, ie diaminoalkanes And derivatives in which the amino group is substituted with a lysyl group to form a lysylamino group, and specific examples include diaminoethane, diaminopropane, N- (lysyl) -diaminoethane, and the like.
- the content of the curing agent in the composition of the present invention is, for example, 0.1 wt% to 10 wt%, preferably 1 wt% to 3 wt%.
- the monovalent metal salt of alginic acid used in the present invention is a monovalent metal salt of low endotoxin alginic acid.
- Carboxymethylcellulose or a pharmaceutically acceptable salt thereof used in the present invention is also preferably a low endotoxin carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
- Low endotoxins are those in which endotoxin levels are reduced to such an extent that they do not substantially cause inflammation or fever. That is, it has been subjected to low endotoxin treatment.
- the low endotoxin treatment can be performed by a known method or a method analogous thereto.
- the method of Takada et al. See, for example, JP-A-9-32001 for purifying sodium hyaluronate
- the method of Yoshida et al. Eg, JP-A-8-269102 for purifying ⁇ 1,3-glucan. Etc.
- a method of William et al. for example, see JP-T-2002-530440, etc.
- biopolymer salts such as alginate, gellan gum, etc.
- James et al. For example, international publication for purifying polysaccharides, etc.
- the low endotoxin treatment of the present invention is not limited thereto, but is washed, filtered with a filter (such as an endotoxin removal filter or a charged filter), ultrafiltration, a column (an endotoxin adsorption affinity column, a gel filtration column, a column with an ion exchange resin, etc.) ), Adsorption to hydrophobic substances, resin or activated carbon, organic solvent treatment (extraction with organic solvent, precipitation / precipitation by addition of organic solvent, etc.), surfactant treatment (for example, JP-A-2005-036036) It can be carried out by a known method such as a gazette) or a combination thereof. These processing steps may be appropriately combined with known methods such as centrifugation. It is desirable to select appropriately according to the type of alginic acid, the type of carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
- the endotoxin level can be confirmed by a known method, and can be measured, for example, by a method using Limulus reagent (LAL), a method using Enspercy (registered trademark) ES-24S set (Seikagaku Corporation), or the like. .
- LAL Limulus reagent
- Enspercy registered trademark
- ES-24S set Seikagaku Corporation
- the method for treating the endotoxin of the monovalent metal salt of alginic acid contained in the composition of the present invention is not particularly limited.
- the endotoxin content of the monovalent metal salt of alginic acid is measured by the endorsin assay using the Limulus reagent (LAL) Is preferably 500 endotoxin units (EU) / g or less, more preferably 100 EU / g or less, particularly preferably 50 EU / g or less, and particularly preferably 30 EU / g or less.
- LAL Limulus reagent
- Low endotoxin-treated sodium alginate is commercially available, for example, Sea Matrix (sterilized) (Kimika Co., Ltd. Mochida International), PRONOVA TM UP LVG (FMC), and the like.
- the endotoxin content of carboxymethylcellulose or a pharmaceutically acceptable salt thereof contained in the composition of the present invention is not limited.
- the endotoxin content of carboxymethylcellulose or a pharmaceutically acceptable salt thereof is not limited. Is preferably 500 endotoxin units (EU) / g or less, more preferably 100 EU / g or less, particularly preferably 50 EU / g or less, particularly when endotoxin measurement using Limulus reagent (LAL) is performed. 30 EU / g or less.
- EU endotoxin units
- Such carboxymethylcellulose or a pharmaceutically acceptable salt thereof can be obtained by subjecting it to the above endotoxin treatment.
- composition of some embodiments of the invention is a fluid liquid, ie, a solution.
- the viscosity of the composition of the present invention of this embodiment is not particularly limited as long as the effects of the present invention can be obtained.
- the viscosity is 5 mPa ⁇ s to 100,000 mPa ⁇ s, preferably 10 mPa ⁇ s to 20000 mPa ⁇ s.
- the composition of some embodiments of the present invention is a viscosity that can also be applied to a subject, such as with a syringe.
- the viscosity of the composition of the present invention is preferably about 10 mPa ⁇ s or more from the viewpoint of adhesion, and preferably about 20000 mPa ⁇ s or less from the viewpoint of easy handling of the composition, and more preferably 2000 mPa ⁇ s to 10,000 mPa. S, 1000 mPa ⁇ s to 5000 mPa ⁇ s, or 10 mPa ⁇ s to 10000 mPa ⁇ s.
- the viscosity of the composition of the present invention includes, for example, alginic acid concentration, alginic acid molecular weight, alginic acid M / G ratio, carboxymethylcellulose or pharmaceutically acceptable salt concentration, carboxymethylcellulose or pharmaceutically acceptable salt thereof.
- alginic acid concentration alginic acid molecular weight
- alginic acid M / G ratio carboxymethylcellulose or pharmaceutically acceptable salt concentration
- carboxymethylcellulose or pharmaceutically acceptable salt thereof By adjusting the molecular weight, the mixing ratio of the monovalent metal salt of alginic acid and carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and the like.
- the viscosity of the monovalent metal salt solution of alginic acid is high when the concentration of alginic acid in the solution is high, and is low when the concentration of alginic acid in the solution is low. Also, it is high when the molecular weight of alginic acid is large, and low when it is small. For example, in order to obtain a viscosity of 10 mPa ⁇ s to 20000 mPa ⁇ s using alginic acid having a molecular weight of about 200 to 500,000 Da, an aqueous alginate solution of about 1% w / v to 3% w / v may be used.
- the viscosity of the aqueous alginate solution can be measured by a known method using, for example, a rotational viscometer (cone plate type) (TVE-20LT, TOKI SANGYO CO., LTD. JAPAN).
- a known method is, for example, the 16th revision Japanese Pharmacopoeia, General Test Method Viscosity Measurement Method (cone-plate rotational viscometer).
- the M / G ratio of alginic acid used in the present invention is about 0.4 to 4.0, preferably about 0.8 to 3.0, more preferably about 1.0 to 1.6.
- the type of brown algae used as a raw material affects the viscosity of a monovalent metal salt solution of alginic acid.
- the alginic acid used in the present invention is preferably derived from a brown algae of the genus Lessonia, Macrocystis, Laminaria, Ascofilum, Davilia, more preferably a brown alga of the genus Lessonia, particularly preferably Lessonia It is derived from Niscens (Lessonia nigrescens).
- the viscosity of the solution of carboxymethylcellulose or a pharmaceutically acceptable salt thereof is high when the carboxymethylcellulose concentration in the solution is high, and is low when the carboxymethylcellulose concentration in the solution is low. Moreover, it is high when the molecular weight of carboxymethyl cellulose is large, and low when it is small.
- an aqueous carboxymethyl cellulose solution of about 1% w / v to 2% w / v may be used. .
- the viscosity of the aqueous carboxymethyl cellulose solution can be measured by a known method using, for example, a rotational viscosity measuring device (corn plate type) (TVE-20LT, TOKI SANGYO CO., LTD. JAPAN).
- a known method is, for example, the 16th revision Japanese Pharmacopoeia, General Test Method Viscosity Measurement Method (cone-plate rotational viscometer).
- the viscosity of the composition obtained by mixing the monovalent metal salt of alginic acid with carboxymethylcellulose or a pharmaceutically acceptable salt thereof is almost the same as before mixing if the viscosity of each solution before mixing is the same. Become. When the viscosity of one solution before mixing is high, the viscosity of the composition obtained can be lowered
- the mixing method of the component (a) and the component (b) is not particularly limited as long as the component (a) and the component (b) can be mixed uniformly.
- Examples of the mixing method include a method using a static mixer and a method using a double syringe.
- a static mixer refers to a static mixing device without a drive unit. More specifically, a static mixer usually consists of a tube and a mixing element without a drive fixed in the tube, thereby splitting the flow and diverting or reversing the flow direction, so that the flow is longitudinal. It is a mixing device that mixes fluids by repeating division, conversion and inversion in the horizontal direction. Some types of static mixers are equipped with a jacket for heat exchange on the outer periphery of the tube, and others are equipped with a tube for heat exchange through which the heat medium passes through the mixing element itself. . Details on the static mixer can be found in, for example, Thakur et al. (2003) Trans IC ChemE, 81, Part A: 787-826, which can be referred to.
- a known mixer can be used as the static mixer.
- the known static mixer include, for example, Kenix type (for example, manufactured by Nordson and Noritake); Kenics (Chemineer Inc.); low pressure drop ( Ross Engineering Inc.); SMV (Koch-Glitsch Inc.); SMXL (Koch-Glitsch Inc.); Interfacial Surface Generator. InC (R) Eng. ); Inliner series 50 (Lightnin Inc.); Inliner series 45 . Lightnin Inc); Custody transfer mixer (Komax systems Inc.);. And SMR (Koch-Glitsch, Inc) and the like.
- the double syringe is a device that has containers for separately filling the components to be mixed, and is uniformly mixed and released when the components are released from the double syringe.
- a well-known thing can be used as a double syringe. Examples of known double syringes include those manufactured by Baxter. Alternatively, the static mixer and double syringe can be custom made to mix the compositions of the present invention.
- antibiotics such as streptomycin, penicillin, tobramycin, amikacin, gentamicin, neomycin, and amphotericin B, aspirin
- non- Concomitant drugs such as steroidal antipyretic analgesics (NSAIDs) and anti-inflammatory drugs such as acetaminophen may be administered.
- NSAIDs steroidal antipyretic analgesics
- anti-inflammatory drugs such as acetaminophen
- Rat partial hepatectomy model A rat partial hepatectomy model was used to evaluate the formation of adhesions.
- the rat partial hepatectomy model is a model capable of reproducibly observing the formation of adhesions that cause severe inflammation and have high strength (Shimizu A et al., (2014) Surg Today. (44): 314-314. 323). Specifically, the formation of adhesions was evaluated as follows.
- MSC mesenchymal stem cells
- MDSC mesenchymal stem cells
- AL20 Lit NO. 3H19
- Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich.
- Seprafilm (Sodium Carboxymethyl Cellulose (Sigma) 419338-100G, weight average molecular weight: 700,000, substitution degree 0.9, counter ion Na type)
- Seprafilm (trade name) is a sheet-like material obtained by mixing carboxymethylcellulose (CMC) and hyaluronic acid, and was obtained from Genzyme GmbH.
- Interlaced (trade name) is a regenerated oxidized cellulose sheet and was obtained from Johnson & Johnson.
- the double syringe used was DUPLOJECT System manufactured by Baxter Corporation.
- Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
- PBS phosphate buffered saline
- physiological saline 0.9 w / v% NaCl
- Alginate / CMC group (n 4): Prevention adhesion and low endotoxin alginic acid sodium 3 wt% solution 0.5mL, the Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL Applied as a material. The content of sodium alginate in the Alginate / CMC gel was 3.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
- Alginate / CMC + MSC group (n 4): a low endotoxin alginic acid 3 wt% solution 0.5mL, mesenchymal in Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL Stem cells 5.0 ⁇ 10 6 cells were encapsulated and applied as an adhesion preventive material.
- MSC i. p. Group (n 4): Immediately after hepatectomy, mesenchymal stem cells 5.0 ⁇ 10 6 cells suspended in 1 mL of physiological saline were intraperitoneally administered and the abdomen was closed. MSC i. v.
- Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points.
- the abdomen was immediately closed and the treatment was terminated.
- the anti-adhesive material was applied after removing the gauze.
- MSC i. p. Group and MSC i. v. In the group, the mesenchymal stem cells were then immediately administered by injection. Next, the abdominal wall and skin were sutured twice and closed. A biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin.
- rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance. Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
- Adhesion was evaluated visually. An adhesion score was assigned to each of the following sites in the abdominal cavity based on the following scoring method. Site: Hepatic section, omentum, peritoneum, small intestine, directly under the median wound Adhesion score: Grade 0: No adhesion is seen at all. Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion) Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion) Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
- Rat partial hepatectomy model The rat partial hepatectomy model described in Example 1 was used to evaluate the formation of adhesions. Specifically, the formation of adhesions was evaluated as follows.
- Low endotoxin sodium alginate was obtained from Mochida Pharmaceutical.
- AL20 Low endotoxin sodium alginate (sterile product) AL20 (Lot NO. 3H19), endotoxin amount 7 EU / g) (Low endotoxin sodium alginate (sterile product) AL100 (Lot NO. 5H08), endotoxin amount 7 EU / g) (low endotoxin alginic acid Sodium (sterilized product) AL500 (Lot NO. 3H17), endotoxin amount 15 EU / g) Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich.
- Low endotoxin CMC was obtained from Mochida Pharmaceutical.
- Low endotoxin CMC non-sterile
- As Seprafilm trade name
- Interceed trade name
- Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
- PBS phosphate buffered saline
- physiological saline 0.9 w / v% NaCl
- the composition of the PBS described in Example 1 was used.
- the weight average molecular weight measured by each following method was described. [Pre-processing method] An eluent was added to the sample and dissolved, and then filtered through a 0.45 ⁇ m membrane filter to obtain a measurement solution.
- Control group (n 8): The margin of the left lateral lobe was measured 3 cm, separated and coagulated and hemostatic (untreated control group).
- Alginate / CMC group (n 5 ⁇ 9): About AL20 and AL100 means a sodium alginate 3 wt% solution 0.5 mL, a solution containing the 100mMCaCl 2 / 1wt% CMC solution (100mMCaCl 2 and 1 wt% CMC, The same applies hereinafter) 1 mL of Alginate / CMC gel prepared by mixing 0.5 mL with a double syringe was applied as an adhesion preventing material.
- the content of sodium alginate in the Alginate / CMC gel was 3.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
- the AL500 a sodium alginate 1 wt% solution 0.5mL, was applied Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL as antiadhesive material.
- the content of sodium alginate in the Alginate / CMC gel was 1.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
- Alginate / CMC group a combination of alginic acid treated with low endotoxin (AL20) and CMC not treated with low endotoxin.
- AL20 ( ⁇ ) / CMC ( ⁇ ) Combination of low endotoxin-treated alginic acid (AL20) and low endotoxin-treated CMC.
- Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points.
- the abdomen was immediately closed and the treatment was terminated.
- the anti-adhesive material was applied after removing the gauze.
- the abdominal wall and skin were sutured twice and closed.
- a biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin.
- rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance. Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
- Adhesion was evaluated as follows. (1) Separation Section The following (a) and (b) were evaluated for the liver separation section described in [Procedure] above. (A) Adhesion grade Adhesion was evaluated visually. An adhesion score was assigned to the hepatic section based on the following scoring method. Adhesion score: Grade 0: No adhesion is seen at all.
- Grade 1 Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
- Grade 2 Adhesion that can be peeled off with tweezers (blunt adhesion)
- Grade 3 Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
- B Adhesive Extent Of the 3 cm hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm (thus, the maximum extent of the section is 30 mm).
- Non-separation section Other than the hepatic section, the following evaluations (a) and (b) were performed on the liver surface, omentum, peritoneum, small intestine, directly under the midline, and the like.
- Adhesion grade Adhesion was evaluated visually. Adhesion scores were assigned based on the following scoring method for sites other than the hepatic section. The site was not specified, and the maximum value of the observed adhesion score was recorded as the adhesion score of the test animal. Adhesion score: Grade 0: No adhesion is seen at all.
- Grade 1 Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
- Grade 2 Adhesion that can be peeled off with tweezers (blunt adhesion)
- Grade 3 Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
- B Adhesive Extent For sites other than the hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm. Similar to (2) and (a) above, the site was not specified, and the maximum value of the width at which the observed adhesion was formed was recorded as the adhesion extent of the test animal.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Hematology (AREA)
- Cell Biology (AREA)
- Developmental Biology & Embryology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Inorganic Chemistry (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Zoology (AREA)
- Vascular Medicine (AREA)
- Surgery (AREA)
- Heart & Thoracic Surgery (AREA)
- Materials For Medical Uses (AREA)
- Medicinal Preparation (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The present invention provides a composition for preventing adhesion using a combination of (a) a monovalent metal salt of low-endotoxin alginic acid, (b) carboxymethylcellulose or a pharmacologically acceptable salt thereof, and (c) cells. A composition for preventing adhesion having a high adhesion-preventing effect is thereby provided.
Description
本発明は、癒着防止用組成物に関する。
The present invention relates to an adhesion preventing composition.
癒着とは、互いに分離しているべき組織の表面が線維性の組織で連結または融合された状態のことをいう。癒着は、外傷や炎症に伴い組織の表面にフィブリンを含む滲出液が生じ、この滲出液が器質化して組織表面が連結または融合されることにより生じる。外科手術において組織の表面にできる外傷や、外傷により引き起こされる炎症、および外科手術において組織表面が乾燥することによる炎症は、癒着が生じる原因となっている。
Adhesion refers to a state where the surfaces of tissues that should be separated from each other are connected or fused with a fibrous tissue. Adhesion occurs when exudate containing fibrin is generated on the surface of the tissue due to trauma or inflammation, and the exudate is organized and the tissue surfaces are connected or fused. Injuries that occur on the surface of tissues during surgery, inflammation caused by the trauma, and inflammation caused by drying of the tissue surface during surgery cause adhesions.
癒着は、時に、不妊、腸の通過障害、慢性骨盤痛の原因となり得る。また、外科手術後に生じた癒着を剥離するために、再度の外科手術が必要となることもある。これらのことから、癒着を防止する必要があり、癒着防止のためにこれまでに様々な手段が講じられている。
Adhesion can sometimes cause infertility, bowel passage problems and chronic pelvic pain. In addition, another surgical operation may be required in order to peel off the adhesion that has occurred after the surgical operation. For these reasons, it is necessary to prevent adhesion, and various measures have been taken to prevent adhesion.
癒着防止のためのそのような手段のいくつかは、外傷または炎症部位とその隣接組織の間に配置して組織の連結または融合を防止する物理的バリアを設けることである。そのような物理的バリアとしては、シート状のものおよびゲル状のものが知られている。
Some such means for preventing adhesions are to place a physical barrier between the site of trauma or inflammation and its adjacent tissue to prevent tissue connection or fusion. As such a physical barrier, a sheet form and a gel form are known.
シート状のものは、ポリテトラフルオロエチレン（ＰＴＦＥ）フィルム（Ｐｒｅｃｌｕｄｅ（商品名）（ＷＬ Ｇｏｒｅ ａｎｄ Ａｓｓｏｃｉａｔｅｓ，Ｉｎｃ．））、ヒアルロン酸（ＨＡ）とカルボキシメチルセルロース（ＣＭＣ）を含有するシート（Ｓｅｐｒａｆｉｌｍ（商品名）（Ｇｅｎｚｙｍｅ ＧｍｂＨ））、再生酸化セルロースシート（Ｉｎｔｒａｃｅｅｄ（商品名）（Ｊｏｈｎｓｏｎ＆Ｊｏｈｎｓｏｎ））などがある。このうちＰＴＦＥフィルムは、生分解性でないため、体内に残存するという問題がある。ＨＡととＣＭＣを含有するシートおよび再生酸化セルロースシートは、生分解性であるものの、肝切除後に生じる癒着のような重篤な癒着を完全に防止できず、癒着防止の効果の点で改善の余地があった。
The sheet-like material is a polytetrafluoroethylene (PTFE) film (Prelude (trade name) (WL Gore and Associates, Inc.)), a sheet (Seprafilm (product) containing hyaluronic acid (HA) and carboxymethyl cellulose (CMC). Name) (Genzyme GmbH)), regenerated oxidized cellulose sheet (Intraceded (trade name) (Johnson & Johnson)) and the like. Among these, the PTFE film is not biodegradable and has a problem of remaining in the body. Although the sheet containing HA and CMC and the regenerated oxidized cellulose sheet are biodegradable, serious adhesion such as adhesion occurring after hepatectomy cannot be completely prevented, and the effect of preventing adhesion can be improved. There was room.
ゲル状のものは、鉄イオン架橋ヒアルロン酸ゲル（Ｉｎｔｅｒｇｅｌ（商品名）（Ｌｉｆｅｃｏｒｅ Ｂｉｏｍｅｄｉｃａｌ，ＬＬＣ））などがある。しかしながら、鉄イオン架橋ヒアルロン酸ゲルは、主に腹腔内の鉄イオンの濃度上昇とハイドロゲルの長期残存により重篤な炎症を引き起こすことから、現在では、臨床において使用されていない。この他、アルギン酸、ＨＡ、ＣＭＣまたはこれらの混合物を含有するゲルを、癒着防止の物理的バリアとして用いることが提案されているものの（特許文献１および２、ならびに非特許文献１～３）、これらのゲルについては、癒着防止効果を十分に発揮できる最適な物性は、明らかではなかった。
Examples of gels include iron ion cross-linked hyaluronic acid gel (Intergel (trade name) (Lifecore Biomedical, LLC)). However, iron ion-crosslinked hyaluronic acid gels are not currently used in clinical practice because they cause severe inflammation mainly due to an increase in the concentration of iron ions in the peritoneal cavity and the long-term remaining of the hydrogel. In addition, although it has been proposed to use a gel containing alginic acid, HA, CMC or a mixture thereof as a physical barrier for preventing adhesions ( Patent Documents 1 and 2, and Non-Patent Documents 1 to 3), For these gels, the optimum physical properties capable of sufficiently exerting the adhesion preventing effect were not clear.
このような物理的バリアの他に、外科手術中に組織を処置する前に、外科手術に関連する組織の表面を、生体適合性があり且つ薬理学上許容される水溶性のポリペプチド、多糖類、合成ポリマー、これらの塩および錯体、これらの混合物からなる群の中から選択される親水性ポリマー材料の水溶液で被覆して、乾燥に起因する組織の癒着を防止することが提案されている（特許文献３）。
In addition to such a physical barrier, prior to treating the tissue during surgery, the surface of the tissue associated with the surgery may be treated with a biocompatible and pharmacologically acceptable water-soluble polypeptide, It has been proposed to coat with an aqueous solution of a hydrophilic polymer material selected from the group consisting of sugars, synthetic polymers, salts and complexes thereof, and mixtures thereof to prevent tissue adhesion due to drying. (Patent Document 3).
このような状況の下、癒着防止効果の高い癒着防止用組成物が求められていた。
Under such circumstances, an anti-adhesion composition having a high anti-adhesion effect has been demanded.
本発明者らは、鋭意検討を重ねた結果、（ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる癒着防止用組成物が癒着防止効果の高いものであることなどを見出し、本発明を完成するに至った。
As a result of extensive studies, the present inventors have found that (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) an adhesion using a combination of cells. The inventors have found that the prevention composition has a high adhesion prevention effect and have completed the present invention.
本発明は以下のとおりである。
［１］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる癒着防止用組成物。
［２］ 前記細胞が、間葉系幹細胞である、上記［１］に記載の組成物。
［３］ 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、上記［１］または［２］に記載の組成物。
［４］ 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、上記［１］～［３］のいずれか１項に記載の組成物。
［５］ 前記組成物が、流動性を有する液体状である、上記［１］～［４］のいずれか１項に記載の組成物。
［６］ 前記組成物の粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、上記［５］に記載の組成物。
［７］ さらに、アルギン酸の１価金属塩の硬化剤を含有する、上記［１］～［４］のいずれか１項に記載の組成物。
［８］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースおよびその薬学的に許容される塩を組合せて用い、かつ（ｃ）細胞を用いない、癒着防止用組成物。
［９］ 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、上記［８］に記載の組成物。
［１０］ 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、上記［８］または［９］に記載の組成物。
［１１］ 前記組成物が、流動性を有する液体状；シート状、スポンジ状、粉体およびパウダー状からなる群から選択される固形物状；半固形状；またはゲル状である、上記［８］～［１０］のいずれか１項に記載の組成物。
［１２］ 前記組成物が、流動性を有する液体状であり、その粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、上記［１１］に記載の組成物。
［１３］ さらに、アルギン酸の１価金属塩の硬化剤を含有する、上記［８］～［１１］のいずれか１項に記載の組成物。
［１４］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。
［１５］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩から選択される増粘剤を組合せて用い、かつ（ｃ）細胞を用いない組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。
［１６］ （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。
［１７］ （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて用いられ、かつ（ｃ）細胞を用いない、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。
［１８］ 上記［１］～［１３］、[１６]および[１７]のいずれか１項に記載の組成物を製造するために用いられる、低エンドトキシンアルギン酸の１価金属塩を含有する製剤。
［１９］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。
［２０］ （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。
［２１］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。
［２２］ （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。
The present invention is as follows.
[1] An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell.
[2] The composition according to [1] above, wherein the cells are mesenchymal stem cells.
[3] The composition according to [1] or [2] above, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
[4] The composition according to any one of [1] to [3], wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
[5] The composition according to any one of [1] to [4] above, wherein the composition is in a liquid form having fluidity.
[6] The composition described in [5] above, wherein the viscosity of the composition is 5 mPa · s to 100,000 mPa · s.
[7] The composition according to any one of [1] to [4], further comprising a curing agent of a monovalent metal salt of alginic acid.
[8] An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose and a pharmaceutically acceptable salt thereof, and (c) a cell not used.
[9] The composition according to [8] above, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
[10] The composition according to [8] or [9] above, wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
[11] The above [8], wherein the composition is in a liquid form having fluidity; a solid form selected from the group consisting of a sheet form, a sponge form, a powder form and a powder form; a semisolid form; or a gel form. ] To [10] The composition according to any one of [10].
[12] The composition according to [11] above, wherein the composition is in a liquid form having fluidity and has a viscosity of 5 mPa · s to 100,000 mPa · s.
[13] The composition according to any one of [8] to [11], further comprising a curing agent of a monovalent metal salt of alginic acid.
[14] A subject in need of anti-adhesion using a composition comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells. A method for preventing adhesions, comprising applying to.
[15] A combination of (a) a monovalent metal salt of low endotoxin alginic acid, and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) no cells. An anti-adhesion method comprising applying the composition to a subject in need of anti-adhesion.
[16] An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells.
[17] An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof and (c) a cell-free (a) monovalent metal salt of low endotoxin alginic acid. object.
[18] A preparation containing a monovalent metal salt of low endotoxin alginic acid, which is used for producing the composition according to any one of [1] to [13], [16] and [17].
[19] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent.
[20] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent, used in combination with cells.
[21] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof.
[22] An anti-adhesion kit comprising (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells, and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof. .
［１］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる癒着防止用組成物。
［２］ 前記細胞が、間葉系幹細胞である、上記［１］に記載の組成物。
［３］ 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、上記［１］または［２］に記載の組成物。
［４］ 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、上記［１］～［３］のいずれか１項に記載の組成物。
［５］ 前記組成物が、流動性を有する液体状である、上記［１］～［４］のいずれか１項に記載の組成物。
［６］ 前記組成物の粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、上記［５］に記載の組成物。
［７］ さらに、アルギン酸の１価金属塩の硬化剤を含有する、上記［１］～［４］のいずれか１項に記載の組成物。
［８］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースおよびその薬学的に許容される塩を組合せて用い、かつ（ｃ）細胞を用いない、癒着防止用組成物。
［９］ 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、上記［８］に記載の組成物。
［１０］ 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、上記［８］または［９］に記載の組成物。
［１１］ 前記組成物が、流動性を有する液体状；シート状、スポンジ状、粉体およびパウダー状からなる群から選択される固形物状；半固形状；またはゲル状である、上記［８］～［１０］のいずれか１項に記載の組成物。
［１２］ 前記組成物が、流動性を有する液体状であり、その粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、上記［１１］に記載の組成物。
［１３］ さらに、アルギン酸の１価金属塩の硬化剤を含有する、上記［８］～［１１］のいずれか１項に記載の組成物。
［１４］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。
［１５］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩から選択される増粘剤を組合せて用い、かつ（ｃ）細胞を用いない組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。
［１６］ （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。
［１７］ （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて用いられ、かつ（ｃ）細胞を用いない、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。
［１８］ 上記［１］～［１３］、[１６]および[１７]のいずれか１項に記載の組成物を製造するために用いられる、低エンドトキシンアルギン酸の１価金属塩を含有する製剤。
［１９］ （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。
［２０］ （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。
［２１］ （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。
［２２］ （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。
The present invention is as follows.
[1] An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell.
[2] The composition according to [1] above, wherein the cells are mesenchymal stem cells.
[3] The composition according to [1] or [2] above, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
[4] The composition according to any one of [1] to [3], wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
[5] The composition according to any one of [1] to [4] above, wherein the composition is in a liquid form having fluidity.
[6] The composition described in [5] above, wherein the viscosity of the composition is 5 mPa · s to 100,000 mPa · s.
[7] The composition according to any one of [1] to [4], further comprising a curing agent of a monovalent metal salt of alginic acid.
[8] An anti-adhesion composition comprising a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose and a pharmaceutically acceptable salt thereof, and (c) a cell not used.
[9] The composition according to [8] above, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
[10] The composition according to [8] or [9] above, wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
[11] The above [8], wherein the composition is in a liquid form having fluidity; a solid form selected from the group consisting of a sheet form, a sponge form, a powder form and a powder form; a semisolid form; or a gel form. ] To [10] The composition according to any one of [10].
[12] The composition according to [11] above, wherein the composition is in a liquid form having fluidity and has a viscosity of 5 mPa · s to 100,000 mPa · s.
[13] The composition according to any one of [8] to [11], further comprising a curing agent of a monovalent metal salt of alginic acid.
[14] A subject in need of anti-adhesion using a composition comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells. A method for preventing adhesions, comprising applying to.
[15] A combination of (a) a monovalent metal salt of low endotoxin alginic acid, and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) no cells. An anti-adhesion method comprising applying the composition to a subject in need of anti-adhesion.
[16] An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells.
[17] An anti-adhesion composition comprising (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof and (c) a cell-free (a) monovalent metal salt of low endotoxin alginic acid. object.
[18] A preparation containing a monovalent metal salt of low endotoxin alginic acid, which is used for producing the composition according to any one of [1] to [13], [16] and [17].
[19] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent.
[20] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent, used in combination with cells.
[21] An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof.
[22] An anti-adhesion kit comprising (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells, and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof. .
本発明によれば、癒着防止効果の高い癒着防止用組成物を提供することができる。
According to the present invention, an anti-adhesion composition having a high anti-adhesion effect can be provided.
以下、本発明を詳細に説明するが、以下の実施の形態は本発明を説明するための例示であり、本発明はその要旨を逸脱しない限りさまざまな形態で実施することができる。
Hereinafter, the present invention will be described in detail. The following embodiments are examples for explaining the present invention, and the present invention can be implemented in various forms without departing from the gist thereof.
１．癒着防止
「癒着」とは、互いに分離しているべき組織の表面が線維性の組織で連結または融合された状態のことをいう。癒着の原因は、外科手術において組織の表面にできる外傷や、外傷により引き起こされる炎症、外科手術において組織表面が乾燥することによる炎症などである。これらの外傷や炎症に伴い組織の表面にフィブリンを含む滲出液が生じ、この滲出液が器質化して組織表面が連結または融合されることにより癒着が形成される。
「癒着防止」とは、癒着の形成を減少させることをいう。癒着防止は、必ずしも癒着の形成を完全に防止することまでも必要とせず、本発明の組成物を適用しなかった場合の状態と比較して、癒着の形成が防止されていればよい。すなわち「癒着防止」は癒着の軽減と言い換えてもよく、例えば、癒着の頻度、範囲および程度から選ばれる少なくとも１つが軽減されていればよい。「癒着防止」は、例えば、実施例に記載した癒着のグレード評価を行った場合に、本発明の組成物を適用しなかった場合の平均癒着のグレードと比較して、平均癒着のグレードがより低くなっていればよい。あるいは、「癒着防止」は、例えば、実施例に記載した癒着のＥｘｔｅｎｔ評価を行った場合に、本発明の組成物を適用しなかった場合の平均癒着のＥｘｔｅｎｔと比較して、平均癒着のＥｘｔｅｎｔがより低くなっていればよい。「癒着防止」は、好ましくは、外科手術に起因して生じる癒着、より好ましくは、外科手術に起因して生じる腹膜癒着の防止である。すなわち、「癒着防止」は、好ましくは、術後の癒着防止である。 1. Anti-adhesion “Adhesion” refers to a condition in which the surfaces of tissues that should be separated from each other are connected or fused with a fibrous tissue. Causes of adhesion include trauma that can occur on the surface of a tissue during surgery, inflammation caused by the trauma, inflammation due to dry tissue surface during surgery, and the like. With these trauma and inflammation, exudate containing fibrin is generated on the surface of the tissue, and this exudate becomes organized and the tissue surface is connected or fused to form an adhesion.
“Anti-adhesion” refers to reducing the formation of adhesions. The prevention of adhesion does not necessarily need to completely prevent the formation of adhesions, and it is only necessary that the formation of adhesions be prevented as compared with the state where the composition of the present invention is not applied. That is, “adhesion prevention” may be rephrased as adhesion reduction. For example, it is sufficient that at least one selected from the frequency, range, and degree of adhesion is reduced. “Adhesion prevention” means that, for example, when the adhesion grade evaluation described in the Examples is performed, the average adhesion grade is higher than the average adhesion grade when the composition of the present invention is not applied. It only has to be low. Alternatively, “adhesion prevention” means, for example, when the extent evaluation of adhesion described in the Examples is performed, compared to the extent of average adhesion when the composition of the present invention is not applied, the extent of average adhesion Should be lower. “Anti-adhesion” is preferably prevention of adhesions resulting from surgery, more preferably peritoneal adhesions resulting from surgery. That is, “adhesion prevention” is preferably prevention of postoperative adhesion.
「癒着」とは、互いに分離しているべき組織の表面が線維性の組織で連結または融合された状態のことをいう。癒着の原因は、外科手術において組織の表面にできる外傷や、外傷により引き起こされる炎症、外科手術において組織表面が乾燥することによる炎症などである。これらの外傷や炎症に伴い組織の表面にフィブリンを含む滲出液が生じ、この滲出液が器質化して組織表面が連結または融合されることにより癒着が形成される。
「癒着防止」とは、癒着の形成を減少させることをいう。癒着防止は、必ずしも癒着の形成を完全に防止することまでも必要とせず、本発明の組成物を適用しなかった場合の状態と比較して、癒着の形成が防止されていればよい。すなわち「癒着防止」は癒着の軽減と言い換えてもよく、例えば、癒着の頻度、範囲および程度から選ばれる少なくとも１つが軽減されていればよい。「癒着防止」は、例えば、実施例に記載した癒着のグレード評価を行った場合に、本発明の組成物を適用しなかった場合の平均癒着のグレードと比較して、平均癒着のグレードがより低くなっていればよい。あるいは、「癒着防止」は、例えば、実施例に記載した癒着のＥｘｔｅｎｔ評価を行った場合に、本発明の組成物を適用しなかった場合の平均癒着のＥｘｔｅｎｔと比較して、平均癒着のＥｘｔｅｎｔがより低くなっていればよい。「癒着防止」は、好ましくは、外科手術に起因して生じる癒着、より好ましくは、外科手術に起因して生じる腹膜癒着の防止である。すなわち、「癒着防止」は、好ましくは、術後の癒着防止である。 1. Anti-adhesion “Adhesion” refers to a condition in which the surfaces of tissues that should be separated from each other are connected or fused with a fibrous tissue. Causes of adhesion include trauma that can occur on the surface of a tissue during surgery, inflammation caused by the trauma, inflammation due to dry tissue surface during surgery, and the like. With these trauma and inflammation, exudate containing fibrin is generated on the surface of the tissue, and this exudate becomes organized and the tissue surface is connected or fused to form an adhesion.
“Anti-adhesion” refers to reducing the formation of adhesions. The prevention of adhesion does not necessarily need to completely prevent the formation of adhesions, and it is only necessary that the formation of adhesions be prevented as compared with the state where the composition of the present invention is not applied. That is, “adhesion prevention” may be rephrased as adhesion reduction. For example, it is sufficient that at least one selected from the frequency, range, and degree of adhesion is reduced. “Adhesion prevention” means that, for example, when the adhesion grade evaluation described in the Examples is performed, the average adhesion grade is higher than the average adhesion grade when the composition of the present invention is not applied. It only has to be low. Alternatively, “adhesion prevention” means, for example, when the extent evaluation of adhesion described in the Examples is performed, compared to the extent of average adhesion when the composition of the present invention is not applied, the extent of average adhesion Should be lower. “Anti-adhesion” is preferably prevention of adhesions resulting from surgery, more preferably peritoneal adhesions resulting from surgery. That is, “adhesion prevention” is preferably prevention of postoperative adhesion.
２．本発明の組成物
本発明は、癒着防止に好ましく用いられる組成物に関する。
本発明のある態様の組成物（以下、「本発明の組成物Ａ」という場合がある）は、（ａ）低エンドトキシンアルギン酸の１価金属塩（以下、「（ａ）成分」という場合がある）、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩（以下、「（ｂ）成分」という場合がある）、および（ｃ）細胞（本明細書中、「（ｃ）成分」という場合がある）を組合せて用いる癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｂ」という場合がある）は、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩を組合せて用い、かつ（ｃ）細胞を用いない、癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｃ」という場合がある）は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｄ」という場合がある）は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて用いられ、かつ（ｃ）細胞を用いない、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物である。
尚、本発明の組成物Ａ、本発明の組成物Ｂ、本発明の組成物Ｃおよび本発明の組成物Ｄを合わせて、「本発明の組成物」という場合がある。 2. The composition of this invention TECHNICAL FIELD This invention relates to the composition used preferably for adhesion prevention.
A composition of an embodiment of the present invention (hereinafter sometimes referred to as “the composition A of the present invention”) may be referred to as (a) a monovalent metal salt of low endotoxin alginic acid (hereinafter referred to as “component (a)”). ), (B) carboxymethylcellulose or a pharmaceutically acceptable salt thereof (hereinafter sometimes referred to as “(b) component”), and (c) a cell (herein referred to as “(c) component”) The anti-adhesion composition used in combination.
A composition according to another aspect of the present invention (sometimes referred to herein as “the composition B of the present invention”) is: (a) a monovalent metal salt of low endotoxin alginic acid, and (b) carboxymethylcellulose or its An anti-adhesion composition that uses a combination of pharmaceutically acceptable salts and does not use (c) cells.
A composition according to another aspect of the present invention (sometimes referred to herein as “composition C of the present invention”) comprises (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell. And (a) a composition for preventing adhesion containing a monovalent metal salt of low endotoxin alginic acid.
The composition according to another aspect of the present invention (hereinafter, sometimes referred to as “the composition D of the present invention”) is used in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, And (c) a composition for preventing adhesion containing (a) a monovalent metal salt of low endotoxin alginic acid without using cells.
The composition A of the present invention, the composition B of the present invention, the composition C of the present invention and the composition D of the present invention may be collectively referred to as “the composition of the present invention”.
本発明は、癒着防止に好ましく用いられる組成物に関する。
本発明のある態様の組成物（以下、「本発明の組成物Ａ」という場合がある）は、（ａ）低エンドトキシンアルギン酸の１価金属塩（以下、「（ａ）成分」という場合がある）、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩（以下、「（ｂ）成分」という場合がある）、および（ｃ）細胞（本明細書中、「（ｃ）成分」という場合がある）を組合せて用いる癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｂ」という場合がある）は、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩を組合せて用い、かつ（ｃ）細胞を用いない、癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｃ」という場合がある）は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物である。
本発明の別の態様の組成物（本明細書中、「本発明の組成物Ｄ」という場合がある）は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて用いられ、かつ（ｃ）細胞を用いない、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物である。
尚、本発明の組成物Ａ、本発明の組成物Ｂ、本発明の組成物Ｃおよび本発明の組成物Ｄを合わせて、「本発明の組成物」という場合がある。 2. The composition of this invention TECHNICAL FIELD This invention relates to the composition used preferably for adhesion prevention.
A composition of an embodiment of the present invention (hereinafter sometimes referred to as “the composition A of the present invention”) may be referred to as (a) a monovalent metal salt of low endotoxin alginic acid (hereinafter referred to as “component (a)”). ), (B) carboxymethylcellulose or a pharmaceutically acceptable salt thereof (hereinafter sometimes referred to as “(b) component”), and (c) a cell (herein referred to as “(c) component”) The anti-adhesion composition used in combination.
A composition according to another aspect of the present invention (sometimes referred to herein as “the composition B of the present invention”) is: (a) a monovalent metal salt of low endotoxin alginic acid, and (b) carboxymethylcellulose or its An anti-adhesion composition that uses a combination of pharmaceutically acceptable salts and does not use (c) cells.
A composition according to another aspect of the present invention (sometimes referred to herein as “composition C of the present invention”) comprises (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell. And (a) a composition for preventing adhesion containing a monovalent metal salt of low endotoxin alginic acid.
The composition according to another aspect of the present invention (hereinafter, sometimes referred to as “the composition D of the present invention”) is used in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, And (c) a composition for preventing adhesion containing (a) a monovalent metal salt of low endotoxin alginic acid without using cells.
The composition A of the present invention, the composition B of the present invention, the composition C of the present invention and the composition D of the present invention may be collectively referred to as “the composition of the present invention”.
本発明は、本発明の組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法にも関する。
The present invention also relates to an adhesion prevention method comprising applying the composition of the present invention to a subject in need of adhesion prevention.
より具体的には、本発明のある態様の方法は、（ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法である。
More specifically, the method of certain aspects of the invention comprises (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell. An anti-adhesion method comprising applying a composition to be used to a subject in need of anti-adhesion.
本発明の別の態様の方法は、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩から選択される増粘剤を組合せて用い、かつ（ｃ）細胞を用いない組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法である。
また、本発明の別の態様は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて、癒着防止に用いるための、（ａ）低エンドトキシンアルギン酸の１価金属塩である。
また、本発明の別の態様は、（ｃ）細胞を用いず、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて、癒着防止に用いるための、（ａ）低エンドトキシンアルギン酸の１価金属塩である。 The method of another aspect of the present invention uses a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (C) An anti-adhesion method comprising applying a cell-free composition to a subject in need of anti-adhesion.
Another aspect of the present invention provides (a) a monovalent monovalent form of low endotoxin alginic acid for use in adhesion prevention in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells. It is a metal salt.
Another aspect of the present invention provides (a) a low endotoxin alginic acid for use in the prevention of adhesion in combination with (c) a cell, and (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof. It is a monovalent metal salt.
また、本発明の別の態様は、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて、癒着防止に用いるための、（ａ）低エンドトキシンアルギン酸の１価金属塩である。
また、本発明の別の態様は、（ｃ）細胞を用いず、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて、癒着防止に用いるための、（ａ）低エンドトキシンアルギン酸の１価金属塩である。 The method of another aspect of the present invention uses a combination of (a) a monovalent metal salt of low endotoxin alginic acid and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (C) An anti-adhesion method comprising applying a cell-free composition to a subject in need of anti-adhesion.
Another aspect of the present invention provides (a) a monovalent monovalent form of low endotoxin alginic acid for use in adhesion prevention in combination with (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) cells. It is a metal salt.
Another aspect of the present invention provides (a) a low endotoxin alginic acid for use in the prevention of adhesion in combination with (c) a cell, and (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof. It is a monovalent metal salt.
「組合せて用いる」とは、併用することである。(ａ)成分と（ｂ）成分と（ｃ）成分を組合せて用いる場合、組成物を対象に適用する際に、(ａ)成分と（ｂ）成分と（ｃ）成分が組成物に含まれていればよいことを意味する。(ａ)成分と（ｂ）成分とを組合せて用い、かつ（ｃ）成分を用いない場合、組成物を対象に適用する際に、(ａ)成分と（ｂ）成分が組成物に含まれ、かつ（ｃ）成分が組成物に含まれていなければよいことを意味する。したがって、「組合せて用いる」という用語は、販売を含む流通段階における形態を特に問うものではない。なお、（ｂ）成分は、さらに硬化剤を含んでいてもよく、また、（ａ）成分、（ｂ）成分、（ｃ）成分とは別に、硬化剤を組み合わせて用いてもよい。
“Use in combination” means to use in combination. When the component (a), the component (b), and the component (c) are used in combination, the component (a), the component (b), and the component (c) are included in the composition when the composition is applied to a target. It means that it should be. When the component (a) and the component (b) are used in combination, and the component (c) is not used, the component (a) and the component (b) are included in the composition when the composition is applied to the object. In addition, it means that the component (c) should not be contained in the composition. Therefore, the term “used in combination” does not particularly refer to the form in the distribution stage including sales. In addition, (b) component may contain the hardening | curing agent further, and you may use it combining a hardening | curing agent separately from (a) component, (b) component, and (c) component.
例えば(ａ)成分と（ｂ）成分と（ｃ）成分を組合せて用いる場合、（１）(ａ)成分と（ｂ）成分と（ｃ）成分があらかじめ組み合わされた形態、（２）(ａ)成分と（ｂ）成分があらかじめ組み合わされた形態と（ｃ）成分を組合せてキットとし、または組合せないで別々に用意し、適用する際に混合して使用する形態、（３）(ａ)成分と（ｂ）成分を別々に用意して得られる２種類の組成物と（ｃ）成分を組合せてキットとし、または組合せないで別々に用意し、適用する際に組み合わせて使用する形態のいずれでもよい。すなわち、（ｂ）成分は、(ａ)成分と予め配合した組成物の形態で提供してもよく、(ａ)成分と（ｂ）成分とを別々に用意して得られる２種類の組成物を組合せてパッケージ化したキットとして提供してもよい。これらの各形態についても、（ｂ）成分は、さらに硬化剤を含んでいてもよく、また、（ａ）成分、（ｂ）成分、（ｃ）成分とは別に、硬化剤を組み合わせて用いてもよい。以下の例示についても同様である。
For example, when (a) component, (b) component and (c) component are used in combination, (1) (a) component, (b) component and (c) component are combined in advance, (2) (a A form in which component (b) and component (b) are pre-combined and component (c) are combined into a kit, or prepared separately without being combined, and used in a mixed form when applied (3) (a) Either of the two types of compositions obtained by separately preparing the component and the component (b) and the component (c) are combined into a kit, or prepared separately without being combined and used in combination when applied. But you can. That is, the component (b) may be provided in the form of a composition previously blended with the component (a), and two types of compositions obtained by separately preparing the component (a) and the component (b). You may provide as a kit which combined and packaged. In each of these forms, the component (b) may further contain a curing agent, and in addition to the components (a), (b), and (c), a combination of curing agents is used. Also good. The same applies to the following examples.
例えば(ａ)成分と（ｂ）成分とを組合せて用い、かつ（ｃ）成分を用いない場合、（１）(ａ)成分と（ｂ）成分があらかじめ組み合わされた形態、（２）(ａ)成分と（ｂ）成分を別々に用意して得られる２種類の組成物を組合せてキットとし、または組合せないで別々に用意し、適用する際に組み合わせて使用する形態のいずれでもよい。すなわち、（ｂ）成分は、(ａ)成分と予め配合した組成物の形態で提供してもよく、(ａ)成分と（ｂ）成分とを別々に用意して得られる２種類の組成物を組合せてパッケージ化したキットとして提供してもよい。
For example, when (a) component and (b) component are used in combination and (c) component is not used, (1) a form in which (a) component and (b) component are combined in advance, (2) (a ) Component and (b) component may be prepared separately, and two types of compositions obtained may be combined into a kit, or may be prepared separately without combining and used in combination. That is, the component (b) may be provided in the form of a composition previously blended with the component (a), and two types of compositions obtained by separately preparing the component (a) and the component (b). You may provide as a kit which combined and packaged.
(ａ)成分と（ｂ）成分は、それぞれが、または混合物が、溶媒を用いて溶液状態で提供されてもよいし、凍結乾燥体（特には、凍結乾燥粉体）などの固形物として提供されてもよい。固形物として提供される場合、本発明の組成物は、投与時には溶媒を用いて、溶液状、ゲル状などの流動性を有する状態で使用されてもよい。
本発明のいくつかの態様の組成物では、(ａ)成分と（ｂ）成分は、それぞれが、または混合物が、シート状、スポンジ状、粉体またはパウダー状からなる群から選択される固形物状；半固形状；またはゲル状で使用されてもよい。 Each of the component (a) and the component (b) or a mixture thereof may be provided in a solution state using a solvent, or may be provided as a solid material such as a lyophilized product (particularly a lyophilized powder). May be. When provided as a solid, the composition of the present invention may be used in a state having fluidity such as a solution or a gel using a solvent at the time of administration.
In the composition of some embodiments of the present invention, the component (a) and the component (b) are each a solid or a mixture selected from the group consisting of a sheet, sponge, powder, or powder Semi-solid; or gel.
本発明のいくつかの態様の組成物では、(ａ)成分と（ｂ）成分は、それぞれが、または混合物が、シート状、スポンジ状、粉体またはパウダー状からなる群から選択される固形物状；半固形状；またはゲル状で使用されてもよい。 Each of the component (a) and the component (b) or a mixture thereof may be provided in a solution state using a solvent, or may be provided as a solid material such as a lyophilized product (particularly a lyophilized powder). May be. When provided as a solid, the composition of the present invention may be used in a state having fluidity such as a solution or a gel using a solvent at the time of administration.
In the composition of some embodiments of the present invention, the component (a) and the component (b) are each a solid or a mixture selected from the group consisting of a sheet, sponge, powder, or powder Semi-solid; or gel.
溶媒は、生体へ適用可能な溶媒であれば特に限定されないが、例えば、注射用水、精製水、蒸留水、イオン交換水（または脱イオン化水）、ミリＱ水、生理食塩水、リン酸緩衝生理食塩水（ＰＢＳ）などが挙げられる。好ましくは、ヒトおよび動物の治療に用いることが可能な注射用水、蒸留水、生理食塩水などである。
The solvent is not particularly limited as long as it is a solvent applicable to a living body. For example, water for injection, purified water, distilled water, ion-exchanged water (or deionized water), milli-Q water, physiological saline, phosphate buffered physiological Examples include saline (PBS). Preferred are water for injection, distilled water, physiological saline and the like that can be used for treatment of humans and animals.
本発明の好ましい態様の組成物では、当該組成物中でのアルギン酸の１価金属塩の含有量は０．１ｗｔ％～１０ｗｔ％程度であり、カルボキシメチルセルロースまたはその薬学的に許容される塩の含有量は０．１ｗｔ％～１０ｗｔ％程度である。より好ましくは、当該組成物中でのアルギン酸の１価金属塩の含有量は１ｗｔ％～５ｗｔ％程度、カルボキシメチルセルロースまたはその薬学的に許容される塩の含有量は１ｗｔ％～５ｗｔ％程度である。本発明の別のいくつかの態様の組成物では、当該組成物中でのアルギン酸の１価金属塩の含有量は０．１ｗｔ％～９９．９ｗｔ％程度であり、カルボキシメチルセルロースまたはその薬学的に許容される塩の含有量は０．１ｗｔ％～９９．９ｗｔ％程度である。より好ましくは、当該組成物中でのアルギン酸の１価金属塩の含有量は１ｗｔ％～９９ｗｔ％程度、カルボキシメチルセルロースまたはその薬学的に許容される塩の含有量は１ｗｔ％～９９ｗｔ％程度である。
In the composition of a preferred embodiment of the present invention, the content of the monovalent metal salt of alginic acid in the composition is about 0.1 wt% to 10 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof The amount is about 0.1 wt% to 10 wt%. More preferably, the content of the monovalent metal salt of alginic acid in the composition is about 1 wt% to 5 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof is about 1 wt% to 5 wt%. . In the composition of another aspect of the present invention, the content of the monovalent metal salt of alginic acid in the composition is about 0.1 wt% to 99.9 wt%, and carboxymethylcellulose or a pharmaceutically acceptable salt thereof. The acceptable salt content is about 0.1 wt% to 99.9 wt%. More preferably, the content of the monovalent metal salt of alginic acid in the composition is about 1 wt% to 99 wt%, and the content of carboxymethyl cellulose or a pharmaceutically acceptable salt thereof is about 1 wt% to 99 wt%. .
「対象」は、ヒトまたは非ヒト温血動物、例えばトリ、およびウシ、サル、ネコ、マウス、ラット、モルモット、ハムスター、ブタ、イヌ、ウサギ、ヒツジ、ウマなどの非ヒト哺乳動物である。
“Subjects” are human or non-human warm-blooded animals, such as birds, and non-human mammals such as cows, monkeys, cats, mice, rats, guinea pigs, hamsters, pigs, dogs, rabbits, sheep, horses.
本発明の組成物を対象に適用する方法は、特に限定されないが、例えば、シリンジ、ゲル用ピペット、専用充填器などで外科手術に関連した組織の表面に直接注入するようにしてよい。「外科手術に関連した組織」とは、外科手術において表面に外傷を負った組織や、外科手術において表面が乾燥することにより炎症が生じたまたは炎症が生じる恐れのある組織である。本発明のいくつかの態様の組成物の場合、外科手術に関連した組織は、腹膜に包まれている臓器、例えば、胃、空腸、回腸、虫垂、結腸、肝臓、脾臓、十二指腸、および膵臓である。本発明の特に好ましい組成物は、肝切除後に生じる癒着のような重篤な癒着を効果的に防止することができる。
The method of applying the composition of the present invention to a subject is not particularly limited, and for example, it may be directly injected onto the surface of a tissue related to surgery with a syringe, a gel pipette, a dedicated filler, or the like. “Tissue associated with surgery” is tissue that has been traumatically injured on the surface during surgery, or tissue that has become or may be inflamed due to dryness of the surface during surgery. In the case of the composition of some embodiments of the present invention, the tissues associated with the surgery are organs encased in the peritoneum, such as the stomach, jejunum, ileum, appendix, colon, liver, spleen, duodenum, and pancreas. is there. Particularly preferred compositions of the present invention can effectively prevent serious adhesions such as adhesions that occur after hepatectomy.
本発明の組成物の形態は、例えば、流動性のある液体状（すなわち、溶液状）；シート状、スポンジ状、粉体（特に、凍結乾燥粉体）またはパウダー状などの固形物状；半固形状；またはゲル状である。本発明において「流動性を有する」とは、その形態を不定形に変化させる性質を持つことを意味する。例えば、組成物をシリンジなどに封入し、外科手術に関連した組織の表面へ注入することができるような流動性を有することが望ましい。ゲル状または溶液状である本発明の組成物は、シリンジ、ゲル用ピペット、専用注射器などで組織の表面に容易に適用することができる。粉体およびパウダー状の組成物は、噴霧装置などで組織の表面に適用することもできる。また、シート状、スポンジ状、紛体、パウダー状、ゲル状、および流動性のある液体状の組成物は、いずれの表面形状にも適合し、本発明の組成物を適用する組織の表面全体に接触することもできる。シート状とは適した厚さ、柔軟性を有する平板の状態であり、スポンジ状は多孔性を有した状態に加工したものを示す。また、組成物を溶液にしたものを凍結または硬化剤により半固形状態にすることもできる。さらにゲル状は組成物を硬化剤（架橋剤）となる多価陽イオンや架橋性試薬で共有結合させることにより作製することができる。
The form of the composition of the present invention is, for example, a fluid liquid form (ie, a solution form); a solid form such as a sheet form, a sponge form, a powder (particularly freeze-dried powder) or a powder form; Solid; or gel. In the present invention, “having fluidity” means having a property of changing its form into an indefinite form. For example, it is desirable to have fluidity so that the composition can be encapsulated in a syringe or the like and injected into the surface of tissue associated with the surgical procedure. The composition of the present invention that is in the form of a gel or solution can be easily applied to the surface of a tissue with a syringe, a pipette for gel, a dedicated syringe or the like. The powder and the powdery composition can also be applied to the tissue surface with a spray device or the like. Sheet-like, sponge-like, powder, powder-like, gel-like, and fluid liquid compositions are suitable for any surface shape and are applied to the entire surface of the tissue to which the composition of the present invention is applied. You can also touch. The sheet form is a flat plate having a suitable thickness and flexibility, and the sponge form is processed into a porous state. Moreover, what made the composition a solution can also be made into a semi-solid state with a freezing or hardening | curing agent. Further, the gel can be prepared by covalently bonding the composition with a polyvalent cation or a cross-linking reagent as a curing agent (cross-linking agent).
３．アルギン酸の１価金属塩
「アルギン酸の１価金属塩」は、アルギン酸の６位のカルボン酸の水素原子を、Ｎａ＋やＫ＋などの１価金属イオンとイオン交換することでつくられる水溶性の塩である。アルギン酸の１価金属塩としては、具体的には、アルギン酸ナトリウム、アルギン酸カリウムなどを挙げることができるが、特には、市販品により入手可能なアルギン酸ナトリウムが好ましい。アルギン酸の１価金属塩の溶液は、架橋剤と混合したときにゲルを形成する。 3. Monovalent metal salt of alginic acid “Monovalent metal salt of alginic acid” is a water-soluble substance formed by ion exchange of the hydrogen atom of the 6-position carboxylic acid of alginic acid with monovalent metal ions such as Na + and K + . It is salt. Specific examples of the monovalent metal salt of alginic acid include sodium alginate and potassium alginate. In particular, sodium alginate that is commercially available is preferable. A solution of a monovalent metal salt of alginic acid forms a gel when mixed with a crosslinking agent.
「アルギン酸の１価金属塩」は、アルギン酸の６位のカルボン酸の水素原子を、Ｎａ＋やＫ＋などの１価金属イオンとイオン交換することでつくられる水溶性の塩である。アルギン酸の１価金属塩としては、具体的には、アルギン酸ナトリウム、アルギン酸カリウムなどを挙げることができるが、特には、市販品により入手可能なアルギン酸ナトリウムが好ましい。アルギン酸の１価金属塩の溶液は、架橋剤と混合したときにゲルを形成する。 3. Monovalent metal salt of alginic acid “Monovalent metal salt of alginic acid” is a water-soluble substance formed by ion exchange of the hydrogen atom of the 6-position carboxylic acid of alginic acid with monovalent metal ions such as Na + and K + . It is salt. Specific examples of the monovalent metal salt of alginic acid include sodium alginate and potassium alginate. In particular, sodium alginate that is commercially available is preferable. A solution of a monovalent metal salt of alginic acid forms a gel when mixed with a crosslinking agent.
本発明に用いる「アルギン酸」は、生分解性の高分子多糖類であって、Ｄ－マンヌロン酸（Ｍ）とＬ－グルロン酸（Ｇ）という２種類のウロン酸が直鎖状に重合したポリマーである。より具体的には、Ｄ－マンヌロン酸のホモポリマー画分（ＭＭ画分）、Ｌ－グルロン酸のホモポリマー画分（ＧＧ画分）、およびＤ－マンヌロン酸とＬ－グルロン酸がランダムに配列した画分（ＭＧ画分）が任意に結合したブロック共重合体である。アルギン酸のＤ－マンヌロン酸とＬ－グルロン酸の構成比（Ｍ／Ｇ比）は、主に海藻等の由来となる生物の種類によって異なり、また、その生物の生育場所や季節による影響を受け、Ｍ／Ｇ比が約０．４の高Ｇ型からＭ／Ｇ比が約５の高Ｍ型まで高範囲にわたる。
“Alginic acid” used in the present invention is a biodegradable polymer polysaccharide, a polymer in which two types of uronic acids, D-mannuronic acid (M) and L-guluronic acid (G), are polymerized in a straight chain. It is. More specifically, D-mannuronic acid homopolymer fraction (MM fraction), L-guluronic acid homopolymer fraction (GG fraction), and D-mannuronic acid and L-guluronic acid are randomly arranged. This is a block copolymer in which the fractions (MG fraction) are bound arbitrarily. The composition ratio (M / G ratio) of D-mannuronic acid and L-guluronic acid of alginic acid varies depending on the type of organism mainly derived from seaweed, etc. It ranges from a high G type with an M / G ratio of about 0.4 to a high M type with an M / G ratio of about 5.
アルギン酸の１価金属塩は高分子多糖類であり、分子量を正確に定めることは困難であるが、一般的に重量平均分子量で１０００～１０００万、好ましくは１万～１０００万、より好ましくは２万～３００万の範囲である。例えば、ゲル浸透クロマトグラフィー（ＧＰＣ）法により測定した重量平均分子量の好ましい範囲として、３０万～２００万が挙げられる。また、例えば、ＧＰＣ－ＭＡＬＳ法により測定した重量平均分子量の好ましい範囲として、１０００～３０万、より好ましい範囲として５万～３０万が挙げられる。
The monovalent metal salt of alginic acid is a high molecular weight polysaccharide, and it is difficult to accurately determine the molecular weight, but generally the weight average molecular weight is 10 to 10 million, preferably 10,000 to 10 million, more preferably 2 It ranges from 10,000 to 3,000,000. For example, a preferable range of the weight average molecular weight measured by gel permeation chromatography (GPC) method is 300,000 to 2,000,000. Further, for example, a preferable range of the weight average molecular weight measured by GPC-MALS method is 1,000 to 300,000, and a more preferable range is 50,000 to 300,000.
通常、高分子多糖類の分子量をゲルろ過クロマトグラフィーにより算出する場合、１０～２０％以上の測定誤差を生じうる。例えば、４０万であれば３２～４８万、５０万であれば４０～６０万、１００万であれば８０～１２０万程度の範囲で値の変動が生じうる。したがって、アルギン酸の１価金属塩について、好適な重量平均分子量範囲は、少なくとも２万以上、より好ましくは５万以上、さらに好ましくは１０万以上である。分子量が高すぎるものは製造が困難であるとともに、水溶液とする際に粘度が高くなりすぎる、溶解性が低下する、長期間の保存で物性を維持しにくいなどの問題を生じるため、重量平均分子量が５００万以下であることが好ましく、より好ましくは３００万以下である。
Usually, when the molecular weight of the high molecular polysaccharide is calculated by gel filtration chromatography, a measurement error of 10 to 20% or more may occur. For example, the value may vary in the range of about 32 to 480,000 for 400,000, 400,000 to 600,000 for 500,000, and about 800 to 1,200,000 for 1,000,000. Therefore, a suitable weight average molecular weight range for the monovalent metal salt of alginic acid is at least 20,000, more preferably 50,000, and even more preferably 100,000. If the molecular weight is too high, it is difficult to produce and causes problems such as excessively high viscosity when used as an aqueous solution, poor solubility, and difficulty in maintaining physical properties during long-term storage. Is preferably 5 million or less, more preferably 3 million or less.
一般に天然物由来の高分子物質は、単一の分子量を持つのではなく、種々の分子量を持つ分子の集合体であるため、ある一定の幅を持った分子量分布として測定される。代表的な測定手法はゲルろ過クロマトグラフィーである。ゲルろ過クロマトグラフィーにより得られる分子量分布の代表的な情報としては、重量平均分子量（Ｍｗ）、数平均分子量（Ｍｎ）、分散比（Ｍｗ／Ｍｎ）が挙げられる。
Generally, a polymer substance derived from a natural product does not have a single molecular weight but is an aggregate of molecules having various molecular weights, and thus is measured as a molecular weight distribution having a certain width. A typical measurement technique is gel filtration chromatography. Representative information on the molecular weight distribution obtained by gel filtration chromatography includes weight average molecular weight (Mw), number average molecular weight (Mn), and dispersion ratio (Mw / Mn).
分子量の大きい高分子の平均分子量への寄与を重視したのが重量平均分子量であり、下記式で表される。
Ｍｗ＝Σ（ＷｉＭｉ）／Ｗ＝Σ（ＨｉＭｉ）／Σ（Ｈｉ）
数平均分子量は、高分子の総重量を高分子の総数で除して算出される。
Ｍｎ＝Ｗ／ΣＮｉ＝Σ（ＭｉＮｉ）／ΣＮｉ＝Σ（Ｈｉ）／Σ（Ｈｉ／Ｍｉ）
ここで、Ｗは高分子の総重量、Ｗｉはｉ番目の高分子の重量、Ｍｉはｉ番目の溶出時間における分子量、Ｎｉは分子量Ｍｉの個数、Ｈｉはｉ番目の溶出時間における高さである。 The weight average molecular weight places importance on the contribution to the average molecular weight of a polymer having a large molecular weight and is represented by the following formula.
Mw = Σ (WiMi) / W = Σ (HiMi) / Σ (Hi)
The number average molecular weight is calculated by dividing the total weight of the polymer by the total number of polymers.
Mn = W / ΣNi = Σ (MiNi) / ΣNi = Σ (Hi) / Σ (Hi / Mi)
Here, W is the total weight of the polymer, Wi is the weight of the i-th polymer, Mi is the molecular weight at the i-th elution time, Ni is the number of molecular weights Mi, and Hi is the height at the i-th elution time. .
Ｍｗ＝Σ（ＷｉＭｉ）／Ｗ＝Σ（ＨｉＭｉ）／Σ（Ｈｉ）
数平均分子量は、高分子の総重量を高分子の総数で除して算出される。
Ｍｎ＝Ｗ／ΣＮｉ＝Σ（ＭｉＮｉ）／ΣＮｉ＝Σ（Ｈｉ）／Σ（Ｈｉ／Ｍｉ）
ここで、Ｗは高分子の総重量、Ｗｉはｉ番目の高分子の重量、Ｍｉはｉ番目の溶出時間における分子量、Ｎｉは分子量Ｍｉの個数、Ｈｉはｉ番目の溶出時間における高さである。 The weight average molecular weight places importance on the contribution to the average molecular weight of a polymer having a large molecular weight and is represented by the following formula.
Mw = Σ (WiMi) / W = Σ (HiMi) / Σ (Hi)
The number average molecular weight is calculated by dividing the total weight of the polymer by the total number of polymers.
Mn = W / ΣNi = Σ (MiNi) / ΣNi = Σ (Hi) / Σ (Hi / Mi)
Here, W is the total weight of the polymer, Wi is the weight of the i-th polymer, Mi is the molecular weight at the i-th elution time, Ni is the number of molecular weights Mi, and Hi is the height at the i-th elution time. .
天然物由来の高分子物質の分子量測定では、測定方法により値に違いが生じうることが知られている（ヒアルロン酸の例：Ｃｈｉｋａｋｏ ＹＯＭＯＴＡ ｅｔ．ａｌ．，Ｂｕｌｌ．Ｎａｔｌ．Ｈｅａｌｔｈ Ｓｃｉ．，Ｖｏｌ．１１７，ｐｐ１３５－１３９（１９９９）、Ｃｈｉｋａｋｏ ＹＯＭＯＴＡ ｅｔ．ａｌ．，Ｂｕｌｌ．Ｎａｔｌ．Ｉｎｓｔ．Ｈｅａｌｔｈ Ｓｃｉ．，Ｖｏｌ．１２１，ｐｐ３０－３３（２００３））。アルギン酸塩の分子量測定については、固有粘度（Ｉｎｔｒｉｎｓｉｃ ｖｉｓｃｏｓｉｔｙ）から算出する方法、ＳＥＣ－ＭＡＬＬＳ（Ｓｉｚｅ Ｅｘｃｌｕｓｉｏｎ Ｃｈｒｏｍａｔｏｇｒａｐｈｙ ｗｉｔｈ Ｍｕｌｔｉｐｌｅ Ａｎｇｌｅ Ｌａｓｅｒ Ｌｉｇｈｔ Ｓｃａｔｔｅｒｉｎｇ Ｄｅｔｅｃｔｉｏｎ）により算出する方法が記載された文献がある（ＡＳＴＭ Ｆ２０６４－００ （２００６），ＡＳＴＭ Ｉｎｔｅｒｎａｔｉｏｎａｌ発行）。なお、当該文献では、サイズ排除クロマトグラフィー（＝ゲルろ過クロマトグラフィー）により分子量を測定するにあたっては、プルランを標準物質として用いた較正曲線に加え多角度光散乱検出器（Ｍｕｌｔｉ Ａｎｇｌｅ Ｌｉｇｈｔ Ｓｃａｔｔｅｒｉｎｇ：ＭＡＬＳ）を併用すること（＝ＳＥＣ－ＭＡＬＳによる測定）を推奨している。また、ＳＥＣ－ＭＡＬＳによる分子量を、アルギン酸塩のカタログ上の規格値として用いている例もある（ＦＭＣ Ｂｉｏｐｏｌｙｍｅｒ社、ＰＲＯＮＯＶＡＴＭ ｓｏｄｉｕｍ ａｌｇｉｎａｔｅｓ ｃａｔａｌｏｇｕｅ）。
It is known that the molecular weight of a high molecular weight substance derived from a natural product may vary depending on the measurement method (examples of hyaluronic acid: Chikako YOMOTA et.al., Bull. Natl. Health Sci., Vol. 117, pp 135-139 (1999), Chikako YOMOTA et.al., Bull. Natl. Inst. Health Sci., Vol. 121, pp 30-33 (2003)). Regarding the molecular weight measurement of alginate, there is a method of calculating from intrinsic viscosity (Intrinsic Viscosity), SEC-MALLS (Size Exclusion Chromatography with Multiple Laser Light Scattering, which is calculated by the Ft. (2006), issued by ASTM International). In this document, when measuring the molecular weight by size exclusion chromatography (= gel filtration chromatography), in addition to a calibration curve using pullulan as a standard substance, a multi-angle light scattering detector (MULTIS) (= Measurement by SEC-MALS) is recommended. In addition, there is an example in which the molecular weight by SEC-MALS is used as a standard value on a catalog of alginate (FMC Biopolymer, PRONOVA ™ sodium alloys catalog).
本明細書中においてアルギン酸塩の分子量を特定する場合は、特段のことわりがない限り、ゲルろ過クロマトグラフィーにより算出される重量平均分子量である。
In the present specification, when the molecular weight of the alginate is specified, it is a weight average molecular weight calculated by gel filtration chromatography unless otherwise specified.
ゲルろ過クロマトグラフィーの代表的な条件としては、プルランを標準物質とした較正曲線を用いることが挙げられる。標準物質として用いるプルランの分子量としては、少なくとも１６０万、７８．８万、４０．４万、２１．２万および１１．２万のものを標準物質として用いることが好ましい。その他、溶離液（２００ｍＭ硝酸ナトリウム溶液）、カラム条件などを特定できる。カラム条件としては、ポリメタクリレート樹脂系充填剤を用い、排除限界分子量１０００万以上のカラムを少なくとも１本ないし３本用いることが好ましい。代表的なカラムは、ＴＳＫｇｅｌ ＧＭＰＷｘ１（直径７．８ｍｍ×３００ｍｍ）およびＧ２５００ＰＷXL(直径７．８ｍｍ×３００ｍｍ）（東ソー株式会社製）である。
Typical conditions for gel filtration chromatography include using a calibration curve with pullulan as a standard substance. The molecular weight of pullulan used as the standard substance is preferably at least 1.6 million, 788,000, 404,000, 212,000 and 112,000 as the standard substance. In addition, eluent (200 mM sodium nitrate solution), column conditions, etc. can be specified. As column conditions, it is preferable to use a polymethacrylate resin filler and use at least one to three columns having an exclusion limit molecular weight of 10 million or more. Typical columns are TSKgel GMPW x1 (diameter 7.8 mm × 300 mm) and G2500PW XL (diameter 7.8 mm × 300 mm) (manufactured by Tosoh Corporation).
アルギン酸の１価金属塩は、褐藻類から抽出された当初は、分子量が大きく、粘度が高めだが、熱による乾燥、凍結乾燥、精製などの過程で、分子量が小さくなり、粘度は低めとなる。したがって、製造の各工程において適切な温度管理をすることにより、分子量の異なるアルギン酸の１価金属塩を製造することができる。製造の各工程における温度が低めとなるよう管理することで分子量の大きいアルギン酸の１価金属塩が得られ、温度が高くなるほど分子量の小さいアルギン酸の１価金属塩が得られる。また、原料とする褐藻類を適宜選択する、あるいは、製造工程において、分子量による分画を行う、などの手法によっても、分子量の異なるアルギン酸の１価金属塩を製造することができる。さらに、各手法で製造したアルギン酸の１価金属塩について、分子量あるいは粘度を測定した後、異なる分子量あるいは粘度を持つ別ロットのアルギン酸の１価金属塩と混合することにより、目的とする分子量を有するアルギン酸の１価金属塩とすることも可能である。
The monovalent metal salt of alginic acid initially has a large molecular weight and a high viscosity when extracted from brown algae, but the molecular weight becomes small and the viscosity becomes low in the process of drying by heat, lyophilization and purification. Therefore, monovalent metal salts of alginic acid having different molecular weights can be produced by performing appropriate temperature control in each production process. A monovalent metal salt of alginic acid having a large molecular weight can be obtained by controlling the temperature in each step of the production to be lower, and a monovalent metal salt of alginic acid having a smaller molecular weight can be obtained as the temperature is increased. Moreover, the monovalent metal salt of alginic acid from which molecular weight differs can also be manufactured by the method of selecting suitably the brown algae used as a raw material, or performing the fractionation by molecular weight in a manufacturing process. Furthermore, the molecular weight or viscosity of the monovalent metal salt of alginic acid produced by each method is measured, and then mixed with another lot of monovalent metal salt of alginic acid having a different molecular weight or viscosity to have the target molecular weight. It is also possible to use a monovalent metal salt of alginic acid.
本発明に用いるアルギン酸は、天然由来でも合成物であってもよいが、天然由来であるのが好ましい。天然由来のアルギン酸としては、例えば、褐藻類から抽出されるものを挙げることができる。アルギン酸を含有する褐藻類は世界中の沿岸域に繁茂しているが、実際にアルギン酸原料として使用できる海藻は限られており、南米のレッソニア、北米のマクロシスティス、欧州のラミナリアやアスコフィラム、豪のダービリアなどが代表的なものである。アルギン酸の原料となる褐藻類としては、例えば、レッソニア（Ｌｅｓｓｏｎｉａ）属、マクロシスティス（Ｍａｃｒｏｃｙｓｔｉｓ）属、ラミナリア（Ｌａｍｉｎａｒｉａ）属（コンブ属）、アスコフィラム（Ａｓｃｏｐｈｙｌｌｕｍ）属、ダービリア（Ｄｕｒｖｉｌｌｅａ）属、アラメ（Ｅｉｓｅｎｉａ）属、カジメ（Ｅｃｋｌｏｎｉａ）属などがあげられる。
The alginic acid used in the present invention may be naturally derived or synthetic, but is preferably naturally derived. Examples of naturally occurring alginic acid include those extracted from brown algae. Although brown alga containing alginic acid is prosperous in coastal areas around the world, seaweed that can actually be used as a raw material for alginic acid is limited, such as Lessonia in South America, Macrocystis in North America, Laminaria and Ascofilum in Europe, Typical examples are Daviglia. Examples of the brown algae used as a raw material for alginic acid include, for example, the genus Lesonia, the genus Macrocystis, the genus Laminaria (comb), the genus Ascophyllum, the genus Durvillea, Examples include the genus Eisenia and the genus Ecklonia.
４．カルボキシメチルセルロースまたはその薬学的に許容される塩
本発明で用いるｂ）成分は、カルボキシメチルセルロースまたはその薬学的に許容される塩である。 4). Carboxymethylcellulose or a pharmaceutically acceptable salt thereof The component b) used in the present invention is carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
本発明で用いるｂ）成分は、カルボキシメチルセルロースまたはその薬学的に許容される塩である。 4). Carboxymethylcellulose or a pharmaceutically acceptable salt thereof The component b) used in the present invention is carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
本発明に用いられるカルボキシメチルセルロースまたはその薬学的に許容される塩は、特に限定されるものではないが、重量平均分子量で、１万～１５０万であるものを用いるのが好ましい。このようなカルボキシメチルセルロースまたはその薬学的に許容される塩の重量平均分子量の測定方法については、前記アルギン酸の１価金属塩の重量平均分子量と同様の測定方法を利用することができる。このようなカルボキシメチルセルロースまたはその薬学的に許容される塩は、各種公知の方法を用いて製造することができ、あるいは市販のものを購入して（例えば、Ｓｉｇｍａ－Ａｌｄｒｉｃｈ、第一工業製薬株式会社、ダイセルファインケム株式会社、ニチリン化学工業株式会社、および日本製紙株式会社から）入手することも可能である。
カルボキシメチルセルロースの薬学的に許容される塩としては、ナトリウム塩、カリウム塩、リチウム塩等のアルカリ金属塩、カルシウム塩等のアルカリ土類金属塩を挙げることができる。この中で、特に好ましい塩は、ナトリウム塩である。 The carboxymethyl cellulose or a pharmaceutically acceptable salt thereof used in the present invention is not particularly limited, but it is preferable to use a carboxymethyl cellulose having a weight average molecular weight of 10,000 to 1,500,000. About the measuring method of the weight average molecular weight of such carboxymethylcellulose or its pharmaceutically acceptable salt, the measuring method similar to the weight average molecular weight of the said monovalent metal salt of alginic acid can be utilized. Such carboxymethylcellulose or a pharmaceutically acceptable salt thereof can be produced by using various known methods, or a commercially available product can be purchased (for example, Sigma-Aldrich, Daiichi Kogyo Seiyaku Co., Ltd.). , Daicel Finechem Co., Ltd., Nichirin Chemical Industry Co., Ltd., and Nippon Paper Industries Co., Ltd.).
Examples of pharmaceutically acceptable salts of carboxymethyl cellulose include alkali metal salts such as sodium salt, potassium salt and lithium salt, and alkaline earth metal salts such as calcium salt. Among these, a particularly preferable salt is a sodium salt.
カルボキシメチルセルロースの薬学的に許容される塩としては、ナトリウム塩、カリウム塩、リチウム塩等のアルカリ金属塩、カルシウム塩等のアルカリ土類金属塩を挙げることができる。この中で、特に好ましい塩は、ナトリウム塩である。 The carboxymethyl cellulose or a pharmaceutically acceptable salt thereof used in the present invention is not particularly limited, but it is preferable to use a carboxymethyl cellulose having a weight average molecular weight of 10,000 to 1,500,000. About the measuring method of the weight average molecular weight of such carboxymethylcellulose or its pharmaceutically acceptable salt, the measuring method similar to the weight average molecular weight of the said monovalent metal salt of alginic acid can be utilized. Such carboxymethylcellulose or a pharmaceutically acceptable salt thereof can be produced by using various known methods, or a commercially available product can be purchased (for example, Sigma-Aldrich, Daiichi Kogyo Seiyaku Co., Ltd.). , Daicel Finechem Co., Ltd., Nichirin Chemical Industry Co., Ltd., and Nippon Paper Industries Co., Ltd.).
Examples of pharmaceutically acceptable salts of carboxymethyl cellulose include alkali metal salts such as sodium salt, potassium salt and lithium salt, and alkaline earth metal salts such as calcium salt. Among these, a particularly preferable salt is a sodium salt.
５．細胞
本発明の組成物Ａは、成分（ｃ）、すなわち細胞を用いる。一方、本発明の組成物Ｂは、成分（ｃ）、すなわち細胞を用いない。 5. Cells Composition A of the present invention uses component (c), ie, cells. On the other hand, the composition B of the present invention does not use component (c), that is, cells.
本発明の組成物Ａは、成分（ｃ）、すなわち細胞を用いる。一方、本発明の組成物Ｂは、成分（ｃ）、すなわち細胞を用いない。 5. Cells Composition A of the present invention uses component (c), ie, cells. On the other hand, the composition B of the present invention does not use component (c), that is, cells.
細胞としては、例えば、幹細胞、間質細胞などが挙げられ、由来は特に限定されないが、骨髄、脂肪組織、臍帯血などを挙げることができる。また、細胞として、ＥＳ細胞およびｉＰＳ細胞を挙げることもできる。細胞として、好ましくは、間葉系幹細胞および間葉系間質細胞を挙げることができ、より好ましくは、骨髄間葉系幹細胞および骨髄間葉系間質細胞を挙げることができる。
Examples of cells include stem cells and stromal cells, and the origin is not particularly limited, but examples thereof include bone marrow, adipose tissue, and cord blood. In addition, examples of cells include ES cells and iPS cells. Preferred examples of the cells include mesenchymal stem cells and mesenchymal stromal cells, and more preferred examples include bone marrow mesenchymal stem cells and bone marrow mesenchymal stromal cells.
「細胞を用いる」とは、骨髄、脂肪組織、臍帯血などから目的とする細胞を回収し濃縮する処理や、培養して量を増やす処理を行い、調製した細胞を本発明の組成物に添加することを言う。具体的には、例えば、１×１０４個／ｍｌ以上、または１×１０５個／ｍｌ以上、好ましくは、１×１０４個／ｍｌ～１×１０７個／ｍｌの細胞を本発明の組成物に含有させることを言う。
“Using cells” means collecting the target cells from bone marrow, adipose tissue, umbilical cord blood, etc., concentrating them, or culturing them to increase the amount, and adding the prepared cells to the composition of the present invention Say to do. Specifically, for example, 1 × 10 4 cells / ml or more, or 1 × 10 5 cells / ml or more, preferably 1 × 10 4 cells / ml to 1 × 10 7 cells / ml are used in the present invention. It is said to be contained in the composition.
本発明の組成物は、細胞の成長を促進する因子を含ませることもできる。そのような因子としては、例えば、ＢＭＰ、ＦＧＦ、ＶＥＧＦ、ＨＧＦ、ＴＧＦ－β、ＩＧＦ－１，ＰＤＧＦ，ＣＤＭＰ，ＣＳＦ，ＥＰＯ、ＩＬおよびＩＦ等が挙げられる。これらの因子は、組み換え法により製造してもよく、あるいは蛋白組成物から精製してもよい。尚、本発明のいくつかの態様の組成物は、これらの成長因子を含まない。
The composition of the present invention may contain a factor that promotes cell growth. Examples of such factors include BMP, FGF, VEGF, HGF, TGF-β, IGF-1, PDGF, CDMP, CSF, EPO, IL, and IF. These factors may be produced by recombinant methods or purified from the protein composition. It should be noted that the compositions of some embodiments of the present invention do not contain these growth factors.
６．硬化剤
本発明のいくつかの態様の組成物は、アルギン酸の１価金属塩の硬化剤を含有しない。硬化剤を含有しない組成物の形態は、例えば、流動性を有する液体状、凍結乾燥体（特に、凍結乾燥粉体）などの固形物である。本発明の別のいくつかの態様の組成物は、アルギン酸の１価金属塩の硬化剤を含有する。硬化剤を含有する組成物の形態は、例えば、凍結乾燥体（特に、凍結乾燥粉体）、シート状、スポンジ状などの固形物；またはゲル状である。
硬化剤は、アルギン酸の１価金属塩の溶液を架橋することにより、固定化するものである。硬化剤は、例えば、Ｃａ２＋、Ｍｇ２＋、Ｂａ２＋、Ｓｒ２＋などの２価以上の金属イオン化合物、分子内に２～４個のアミノ基を有する架橋性試薬などが挙げられる。より具体的には、２価以上の金属イオン化合物として、ＣａＣｌ２、ＭｇＣｌ２、ＣａＳＯ４、ＢａＣｌ２、ＳｒＣｌ２等（好ましくは、ＣａＣｌ２、ＣａＳＯ４、ＢａＣｌ２等）を、分子内に２～４個のアミノ基を有する架橋性試薬として、窒素原子上にリジル（ｌｙｓｙｌ）基（－ＣＯＣＨ（ＮＨ２）－（ＣＨ２）４－ＮＨ２）を有することもあるジアミノアルカン、すなわちジアミノアルカンおよびそのアミノ基がリジル基で置換されてリジルアミノ基を形成している誘導体が包含され、具体的にはジアミノエタン、ジアミノプロパン、Ｎ－（リジル）－ジアミノエタン等を挙げることができる。
硬化剤を含有する場合、硬化剤の含有量は、本発明の組成物の粘度に応じて適宜調節するのが望ましい。本発明の組成物における、硬化剤の含有量は、例えば、０．１ｗｔ％～１０ｗｔ％、好ましくは、１ｗｔ％～３ｗｔ％である。
6). Hardener
The composition of some embodiments of the present invention does not contain a curing agent of a monovalent metal salt of alginic acid. The form of the composition that does not contain a curing agent is, for example, a liquid having fluidity, a solid such as a lyophilized product (particularly lyophilized powder). The composition of some other embodiments of the present invention contains a curing agent of a monovalent metal salt of alginic acid. The form of the composition containing the curing agent is, for example, a freeze-dried product (particularly freeze-dried powder), a solid material such as a sheet or sponge; or a gel.
The curing agent is immobilized by crosslinking a solution of a monovalent metal salt of alginic acid. Examples of the curing agent include divalent or higher valent metal ion compounds such as Ca 2+ , Mg 2+ , Ba 2+ , and Sr 2+, and crosslinkable reagents having 2 to 4 amino groups in the molecule. More specifically, CaCl 2 , MgCl 2 , CaSO 4 , BaCl 2 , SrCl 2, etc. (preferably CaCl 2 , CaSO 4 , BaCl 2, etc.) are used as the divalent or higher valent metal ion compound in the molecule. Diaminoalkanes that may have a lysyl group (—COCH (NH 2 ) — (CH 2 ) 4 —NH 2 ) on the nitrogen atom as crosslinkable reagents having ˜4 amino groups, ie diaminoalkanes And derivatives in which the amino group is substituted with a lysyl group to form a lysylamino group, and specific examples include diaminoethane, diaminopropane, N- (lysyl) -diaminoethane, and the like.
When the curing agent is contained, it is desirable that the content of the curing agent is appropriately adjusted according to the viscosity of the composition of the present invention. The content of the curing agent in the composition of the present invention is, for example, 0.1 wt% to 10 wt%, preferably 1 wt% to 3 wt%.
本発明のいくつかの態様の組成物は、アルギン酸の１価金属塩の硬化剤を含有しない。硬化剤を含有しない組成物の形態は、例えば、流動性を有する液体状、凍結乾燥体（特に、凍結乾燥粉体）などの固形物である。本発明の別のいくつかの態様の組成物は、アルギン酸の１価金属塩の硬化剤を含有する。硬化剤を含有する組成物の形態は、例えば、凍結乾燥体（特に、凍結乾燥粉体）、シート状、スポンジ状などの固形物；またはゲル状である。
硬化剤は、アルギン酸の１価金属塩の溶液を架橋することにより、固定化するものである。硬化剤は、例えば、Ｃａ２＋、Ｍｇ２＋、Ｂａ２＋、Ｓｒ２＋などの２価以上の金属イオン化合物、分子内に２～４個のアミノ基を有する架橋性試薬などが挙げられる。より具体的には、２価以上の金属イオン化合物として、ＣａＣｌ２、ＭｇＣｌ２、ＣａＳＯ４、ＢａＣｌ２、ＳｒＣｌ２等（好ましくは、ＣａＣｌ２、ＣａＳＯ４、ＢａＣｌ２等）を、分子内に２～４個のアミノ基を有する架橋性試薬として、窒素原子上にリジル（ｌｙｓｙｌ）基（－ＣＯＣＨ（ＮＨ２）－（ＣＨ２）４－ＮＨ２）を有することもあるジアミノアルカン、すなわちジアミノアルカンおよびそのアミノ基がリジル基で置換されてリジルアミノ基を形成している誘導体が包含され、具体的にはジアミノエタン、ジアミノプロパン、Ｎ－（リジル）－ジアミノエタン等を挙げることができる。
硬化剤を含有する場合、硬化剤の含有量は、本発明の組成物の粘度に応じて適宜調節するのが望ましい。本発明の組成物における、硬化剤の含有量は、例えば、０．１ｗｔ％～１０ｗｔ％、好ましくは、１ｗｔ％～３ｗｔ％である。
6). Hardener
The composition of some embodiments of the present invention does not contain a curing agent of a monovalent metal salt of alginic acid. The form of the composition that does not contain a curing agent is, for example, a liquid having fluidity, a solid such as a lyophilized product (particularly lyophilized powder). The composition of some other embodiments of the present invention contains a curing agent of a monovalent metal salt of alginic acid. The form of the composition containing the curing agent is, for example, a freeze-dried product (particularly freeze-dried powder), a solid material such as a sheet or sponge; or a gel.
The curing agent is immobilized by crosslinking a solution of a monovalent metal salt of alginic acid. Examples of the curing agent include divalent or higher valent metal ion compounds such as Ca 2+ , Mg 2+ , Ba 2+ , and Sr 2+, and crosslinkable reagents having 2 to 4 amino groups in the molecule. More specifically, CaCl 2 , MgCl 2 , CaSO 4 , BaCl 2 , SrCl 2, etc. (preferably CaCl 2 , CaSO 4 , BaCl 2, etc.) are used as the divalent or higher valent metal ion compound in the molecule. Diaminoalkanes that may have a lysyl group (—COCH (NH 2 ) — (CH 2 ) 4 —NH 2 ) on the nitrogen atom as crosslinkable reagents having ˜4 amino groups, ie diaminoalkanes And derivatives in which the amino group is substituted with a lysyl group to form a lysylamino group, and specific examples include diaminoethane, diaminopropane, N- (lysyl) -diaminoethane, and the like.
When the curing agent is contained, it is desirable that the content of the curing agent is appropriately adjusted according to the viscosity of the composition of the present invention. The content of the curing agent in the composition of the present invention is, for example, 0.1 wt% to 10 wt%, preferably 1 wt% to 3 wt%.
７．低エンドトキシン処理
本発明で用いるアルギン酸の１価金属塩は、低エンドトキシンアルギン酸の１価金属塩である。本発明で用いるカルボキシメチルセルロースまたはその薬学的に許容される塩も、低エンドトキシンカルボキシメチルセルロースまたはその薬学的に許容される塩であるのが好ましい。低エンドトキシンとは、実質的に炎症、または発熱を惹起しない程度にまでエンドトキシンレベルを低下させたものである。すなわち、低エンドトキシン処理に供されたものである。 7). Low endotoxin treatment The monovalent metal salt of alginic acid used in the present invention is a monovalent metal salt of low endotoxin alginic acid. Carboxymethylcellulose or a pharmaceutically acceptable salt thereof used in the present invention is also preferably a low endotoxin carboxymethylcellulose or a pharmaceutically acceptable salt thereof. Low endotoxins are those in which endotoxin levels are reduced to such an extent that they do not substantially cause inflammation or fever. That is, it has been subjected to low endotoxin treatment.
本発明で用いるアルギン酸の１価金属塩は、低エンドトキシンアルギン酸の１価金属塩である。本発明で用いるカルボキシメチルセルロースまたはその薬学的に許容される塩も、低エンドトキシンカルボキシメチルセルロースまたはその薬学的に許容される塩であるのが好ましい。低エンドトキシンとは、実質的に炎症、または発熱を惹起しない程度にまでエンドトキシンレベルを低下させたものである。すなわち、低エンドトキシン処理に供されたものである。 7). Low endotoxin treatment The monovalent metal salt of alginic acid used in the present invention is a monovalent metal salt of low endotoxin alginic acid. Carboxymethylcellulose or a pharmaceutically acceptable salt thereof used in the present invention is also preferably a low endotoxin carboxymethylcellulose or a pharmaceutically acceptable salt thereof. Low endotoxins are those in which endotoxin levels are reduced to such an extent that they do not substantially cause inflammation or fever. That is, it has been subjected to low endotoxin treatment.
低エンドトキシン処理は、公知の方法またはそれに準じる方法によって行うことができる。例えば、ヒアルロン酸ナトリウムを精製する、菅らの方法（例えば、特開平９－３２４００１号公報など参照）、β１，３－グルカンを精製する、吉田らの方法（例えば、特開平８－２６９１０２号公報など参照）、アルギネート、ゲランガム等の生体高分子塩を精製する、ウィリアムらの方法（例えば、特表２００２－５３０４４０号公報など参照）、ポリサッカライドを精製する、ジェームスらの方法（例えば、国際公開第９３／１３１３６号パンフレットなど参照）、ルイスらの方法（例えば、米国特許第５５８９５９１号明細書など参照）、アルギネートを精製する、ハーマンフランクらの方法（例えば、Ａｐｐｌ Ｍｉｃｒｏｂｉｏｌ Ｂｉｏｔｅｃｈｎｏｌ （１９９４）４０：６３８－６４３など参照）等またはこれらに準じる方法によって実施することができる。本発明の低エンドトキシン処理は、それらに限らず、洗浄、フィルター（エンドトキシン除去フィルターや帯電したフィルターなど）によるろ過、限外ろ過、カラム（エンドトキシン吸着アフィニティーカラム、ゲルろ過カラム、イオン交換樹脂によるカラムなど）を用いた精製、疎水性物質、樹脂または活性炭などへの吸着、有機溶媒処理（有機溶媒による抽出、有機溶剤添加による析出・沈降など）、界面活性剤処理（例えば、特開２００５－０３６０３６号公報など参照）など公知の方法によって、あるいはこれらを適宜組合せて実施することができる。これらの処理の工程に、遠心分離など公知の方法を適宜組み合わせてもよい。アルギン酸の種類、カルボキシメチルセルロースまたはその薬学的に許容される塩の種類などに合わせて適宜選択するのが望ましい。
The low endotoxin treatment can be performed by a known method or a method analogous thereto. For example, the method of Takada et al. (See, for example, JP-A-9-32001) for purifying sodium hyaluronate, and the method of Yoshida et al. (Eg, JP-A-8-269102) for purifying β1,3-glucan. Etc.), a method of William et al. (For example, see JP-T-2002-530440, etc.) for purifying biopolymer salts such as alginate, gellan gum, etc., a method of James et al. (For example, international publication) for purifying polysaccharides, etc. 93/13136 pamphlet), the method of Lewis et al. (For example, see US Pat. No. 5,585,591 etc.), the method of Herman Frank et al. (For example, Appl Microbiol Biotechnol (1994) 40: 638) for purifying alginate. -Refer to -643 etc.) or similar It can be carried out by methods. The low endotoxin treatment of the present invention is not limited thereto, but is washed, filtered with a filter (such as an endotoxin removal filter or a charged filter), ultrafiltration, a column (an endotoxin adsorption affinity column, a gel filtration column, a column with an ion exchange resin, etc.) ), Adsorption to hydrophobic substances, resin or activated carbon, organic solvent treatment (extraction with organic solvent, precipitation / precipitation by addition of organic solvent, etc.), surfactant treatment (for example, JP-A-2005-036036) It can be carried out by a known method such as a gazette) or a combination thereof. These processing steps may be appropriately combined with known methods such as centrifugation. It is desirable to select appropriately according to the type of alginic acid, the type of carboxymethylcellulose or a pharmaceutically acceptable salt thereof.
エンドトキシンレベルは、公知の方法で確認することができ、例えば、リムルス試薬（ＬＡＬ）による方法、エントスペシー（登録商標）ＥＳ－２４Ｓセット（生化学工業株式会社）を用いる方法などによって測定することができる。
The endotoxin level can be confirmed by a known method, and can be measured, for example, by a method using Limulus reagent (LAL), a method using Enspercy (registered trademark) ES-24S set (Seikagaku Corporation), or the like. .
本発明の組成物に含有されるアルギン酸の１価金属塩のエンドトキシンの処理方法は特に限定されないが、その結果として、アルギン酸の１価金属塩のエンドトキシン含有量が、リムルス試薬（ＬＡＬ）によるエンドトキシン測定を行った場合に、５００エンドトキシン単位（ＥＵ）／ｇ以下であることが好ましく、さらに好ましくは、１００ＥＵ／ｇ以下、とりわけ好ましくは５０ＥＵ／ｇ以下、特には３０ＥＵ／ｇ以下である。低エンドトキシン処理されたアルギン酸ナトリウムは、例えば、Ｓｅａ Ｍａｔｒｉｘ（滅菌）（（株）キミカ－（株）持田インターナショナル）、ＰＲＯＮＯＶＡＴＭＵＰ ＬＶＧ（ＦＭＣ）など市販品により入手可能である。
The method for treating the endotoxin of the monovalent metal salt of alginic acid contained in the composition of the present invention is not particularly limited. As a result, the endotoxin content of the monovalent metal salt of alginic acid is measured by the endorsin assay using the Limulus reagent (LAL) Is preferably 500 endotoxin units (EU) / g or less, more preferably 100 EU / g or less, particularly preferably 50 EU / g or less, and particularly preferably 30 EU / g or less. Low endotoxin-treated sodium alginate is commercially available, for example, Sea Matrix (sterilized) (Kimika Co., Ltd. Mochida International), PRONOVA ™ UP LVG (FMC), and the like.
また、本発明の組成物に含有されるカルボキシメチルセルロースまたはその薬学的に許容される塩のエンドトキシン含有量も限定されないが、その結果として、カルボキシメチルセルロースまたはその薬学的に許容される塩のエンドトキシン含有量が、リムルス試薬（ＬＡＬ）によるエンドトキシン測定を行った場合に、５００エンドトキシン単位（ＥＵ）／ｇ以下であることが好ましく、さらに好ましくは、１００ＥＵ／ｇ以下、とりわけ好ましくは５０ＥＵ／ｇ以下、特には３０ＥＵ／ｇ以下である。このようなカルボキシメチルセルロースまたはその薬学的に許容される塩は、上記エンドトキシン処理に供することにより入手することができる。
Further, the endotoxin content of carboxymethylcellulose or a pharmaceutically acceptable salt thereof contained in the composition of the present invention is not limited. As a result, the endotoxin content of carboxymethylcellulose or a pharmaceutically acceptable salt thereof is not limited. Is preferably 500 endotoxin units (EU) / g or less, more preferably 100 EU / g or less, particularly preferably 50 EU / g or less, particularly when endotoxin measurement using Limulus reagent (LAL) is performed. 30 EU / g or less. Such carboxymethylcellulose or a pharmaceutically acceptable salt thereof can be obtained by subjecting it to the above endotoxin treatment.
８．本発明の組成物の粘度
本発明のいくつかの態様の組成物は、流動性のある液体状、すなわち、溶液状である。この態様の本発明の組成物の粘度は、本発明の効果が得られれば、特に限定されないが、例えば、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓ、好ましくは、１０ｍＰａ・ｓ～２００００ｍＰａ・ｓである。本発明のいくつかの態様の組成物は、シリンジ等で対象に適用することもできる粘度である。 8). Viscosity of Compositions of the Invention The composition of some embodiments of the invention is a fluid liquid, ie, a solution. The viscosity of the composition of the present invention of this embodiment is not particularly limited as long as the effects of the present invention can be obtained. For example, the viscosity is 5 mPa · s to 100,000 mPa · s, preferably 10 mPa · s to 20000 mPa · s. The composition of some embodiments of the present invention is a viscosity that can also be applied to a subject, such as with a syringe.
本発明のいくつかの態様の組成物は、流動性のある液体状、すなわち、溶液状である。この態様の本発明の組成物の粘度は、本発明の効果が得られれば、特に限定されないが、例えば、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓ、好ましくは、１０ｍＰａ・ｓ～２００００ｍＰａ・ｓである。本発明のいくつかの態様の組成物は、シリンジ等で対象に適用することもできる粘度である。 8). Viscosity of Compositions of the Invention The composition of some embodiments of the invention is a fluid liquid, ie, a solution. The viscosity of the composition of the present invention of this embodiment is not particularly limited as long as the effects of the present invention can be obtained. For example, the viscosity is 5 mPa · s to 100,000 mPa · s, preferably 10 mPa · s to 20000 mPa · s. The composition of some embodiments of the present invention is a viscosity that can also be applied to a subject, such as with a syringe.
本発明の組成物の粘度は、付着性の観点から１０ｍＰａ・ｓ程度以上、組成物の取り扱いやすさの観点から、２００００ｍＰａ・ｓ程度以下であることが好ましく、より好ましくは、２０００ｍＰａ・ｓ～１００００ｍＰａ・ｓ、１０００ｍＰａ・ｓ～５０００ｍＰａ・ｓ、または１０ｍＰａ・ｓ～１００００ｍＰａ・ｓである。
The viscosity of the composition of the present invention is preferably about 10 mPa · s or more from the viewpoint of adhesion, and preferably about 20000 mPa · s or less from the viewpoint of easy handling of the composition, and more preferably 2000 mPa · s to 10,000 mPa. S, 1000 mPa · s to 5000 mPa · s, or 10 mPa · s to 10000 mPa · s.
本発明の組成物の粘度は、例えば、アルギン酸濃度、アルギン酸の分子量、アルギン酸のＭ／Ｇ比、カルボキシメチルセルロースまたはその薬学的に許容される塩の濃度、カルボキシメチルセルロースまたはその薬学的に許容される塩の分子量、アルギン酸の１価金属塩とカルボキシメチルセルロースまたはその薬学的に許容される塩との混合比等を制御することにより調整することができる。
The viscosity of the composition of the present invention includes, for example, alginic acid concentration, alginic acid molecular weight, alginic acid M / G ratio, carboxymethylcellulose or pharmaceutically acceptable salt concentration, carboxymethylcellulose or pharmaceutically acceptable salt thereof. By adjusting the molecular weight, the mixing ratio of the monovalent metal salt of alginic acid and carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and the like.
アルギン酸の１価金属塩の溶液の粘度は、溶液中のアルギン酸濃度が高い場合に高く、溶液中のアルギン酸濃度が低い場合に低くなる。またアルギン酸の分子量が大きい場合に高く、小さい場合に低くなる。例として、分子量約２０～５０万Ｄａのアルギン酸を用いて、１０ｍＰａ・ｓ～２００００ｍＰａ・ｓの粘度を得るには、約１％ｗ／ｖ～３％ｗ／ｖのアルギン酸水溶液とすればよい。より分子量が小さいアルギン酸を用いる場合は、これよりアルギン酸の濃度を高める必要がある。アルギン酸水溶液の粘度は、例えば、回転粘度測定器（コーンプレートタイプ）（ＴＶＥ－２０ＬＴ，ＴＯＫＩ ＳＡＮＧＹＯ ＣＯ．，ＬＴＤ．ＪＡＰＡＮ）などを用いて、公知の方法で測定することができる。公知の方法は、例えば、第１６改正日本薬局方 一般試験法粘度測定法（円すい－平板形回転粘度計）である。
The viscosity of the monovalent metal salt solution of alginic acid is high when the concentration of alginic acid in the solution is high, and is low when the concentration of alginic acid in the solution is low. Also, it is high when the molecular weight of alginic acid is large, and low when it is small. For example, in order to obtain a viscosity of 10 mPa · s to 20000 mPa · s using alginic acid having a molecular weight of about 200 to 500,000 Da, an aqueous alginate solution of about 1% w / v to 3% w / v may be used. When alginic acid having a smaller molecular weight is used, it is necessary to increase the concentration of alginic acid. The viscosity of the aqueous alginate solution can be measured by a known method using, for example, a rotational viscometer (cone plate type) (TVE-20LT, TOKI SANGYO CO., LTD. JAPAN). A known method is, for example, the 16th revision Japanese Pharmacopoeia, General Test Method Viscosity Measurement Method (cone-plate rotational viscometer).
アルギン酸の１価金属塩の溶液の粘度は、Ｍ／Ｇ比によって影響を受けるため、例えば、溶液の粘度等により好ましいＭ／Ｇ比を有するアルギン酸を適宜選択することができる。本発明に用いるアルギン酸のＭ／Ｇ比は、約０．４～４．０であり、好ましくは約０．８～３．０、より好ましくは約１．０～１．６である。
Since the viscosity of a solution of a monovalent metal salt of alginic acid is affected by the M / G ratio, for example, an alginic acid having a preferable M / G ratio can be selected depending on the viscosity of the solution. The M / G ratio of alginic acid used in the present invention is about 0.4 to 4.0, preferably about 0.8 to 3.0, more preferably about 1.0 to 1.6.
前述のように、Ｍ／Ｇ比が主に海藻の種類によって決まることなどから、原料として用いられる褐藻類の種類はアルギン酸の１価金属塩の溶液の粘度に影響を及ぼす。本発明で用いられるアルギン酸としては、好ましくは、レッソニア属、マクロシスティス属、ラミナリア属、アスコフィラム属、ダービリア属の褐藻由来であり、より好ましくはレッソニア属の褐藻由来であり、特に好ましくはレッソニア・ニグレッセンズ（Ｌｅｓｓｏｎｉａ ｎｉｇｒｅｓｃｅｎｓ）由来である。
As described above, since the M / G ratio is mainly determined by the type of seaweed, the type of brown algae used as a raw material affects the viscosity of a monovalent metal salt solution of alginic acid. The alginic acid used in the present invention is preferably derived from a brown algae of the genus Lessonia, Macrocystis, Laminaria, Ascofilum, Davilia, more preferably a brown alga of the genus Lessonia, particularly preferably Lessonia It is derived from Niscens (Lessonia nigrescens).
カルボキシメチルセルロースまたはその薬学的に許容される塩の溶液の粘度は、溶液中のカルボキシメチルセルロース濃度が高い場合に高く、溶液中のカルボキシメチルセルロース濃度が低い場合に低くなる。またカルボキシメチルセルロースの分子量が大きい場合に高く、小さい場合に低くなる。例として、分子量約１０万Ｄａのカルボキシメチルセルロースナトリウムを用いて、４００ｍＰａ・ｓ～２００００ｍＰａ・ｓの粘度を得るには、約１％ｗ／ｖ～２％ｗ／ｖのカルボキシメチルセルロース水溶液とすればよい。より分子量が小さいカルボキシメチルセルロースを用いる場合は、これよりカルボキシメチルセルロースの濃度を高める必要がある。カルボキシメチルセルロース水溶液の粘度は、例えば、回転粘度測定器（コーンプレートタイプ）（ＴＶＥ－２０ＬＴ，ＴＯＫＩ ＳＡＮＧＹＯ ＣＯ．，ＬＴＤ．ＪＡＰＡＮ）などを用いて、公知の方法で測定することができる。公知の方法は、例えば、第１６改正日本薬局方 一般試験法粘度測定法（円すい－平板形回転粘度計）である。
The viscosity of the solution of carboxymethylcellulose or a pharmaceutically acceptable salt thereof is high when the carboxymethylcellulose concentration in the solution is high, and is low when the carboxymethylcellulose concentration in the solution is low. Moreover, it is high when the molecular weight of carboxymethyl cellulose is large, and low when it is small. As an example, in order to obtain a viscosity of 400 mPa · s to 20000 mPa · s using sodium carboxymethylcellulose having a molecular weight of about 100,000 Da, an aqueous carboxymethyl cellulose solution of about 1% w / v to 2% w / v may be used. . When carboxymethylcellulose having a smaller molecular weight is used, it is necessary to increase the concentration of carboxymethylcellulose. The viscosity of the aqueous carboxymethyl cellulose solution can be measured by a known method using, for example, a rotational viscosity measuring device (corn plate type) (TVE-20LT, TOKI SANGYO CO., LTD. JAPAN). A known method is, for example, the 16th revision Japanese Pharmacopoeia, General Test Method Viscosity Measurement Method (cone-plate rotational viscometer).
アルギン酸の１価金属塩とカルボキシメチルセルロースまたはその薬学的に許容される塩とを混合して得られる組成物の粘度は、混合前の各溶液の粘度が同じであれば、混合前とほぼ同じになる。混合前の一方の溶液の粘度が高い場合には、これに、低い粘度の他方の溶液を混合することで、得られる組成物の粘度を下げることができる。混合前の一方の粘度の溶液が低い場合には、これに、高い粘度の他方の溶液を混合することで、得られる組成物の粘度を上げることができる。このようにして、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓの粘度を得ることができる。
The viscosity of the composition obtained by mixing the monovalent metal salt of alginic acid with carboxymethylcellulose or a pharmaceutically acceptable salt thereof is almost the same as before mixing if the viscosity of each solution before mixing is the same. Become. When the viscosity of one solution before mixing is high, the viscosity of the composition obtained can be lowered | hung by mixing the other solution of low viscosity with this. When the solution of one viscosity before mixing is low, the viscosity of the composition obtained can be raised by mixing the other solution of high viscosity with this. In this way, a viscosity of 5 mPa · s to 100,000 mPa · s can be obtained.
９．混合方法
本発明の組成物において、(ａ)成分と（ｂ）成分の混合方法は、(ａ)成分と（ｂ）成分が均一に混合可能である限り特に限定されない。混合方法としては、例えば、スタティックミキサーを使用する方法、ダブルシリンジを使用する方法などが、挙げられる。 9. Mixing Method In the composition of the present invention, the mixing method of the component (a) and the component (b) is not particularly limited as long as the component (a) and the component (b) can be mixed uniformly. Examples of the mixing method include a method using a static mixer and a method using a double syringe.
本発明の組成物において、(ａ)成分と（ｂ）成分の混合方法は、(ａ)成分と（ｂ）成分が均一に混合可能である限り特に限定されない。混合方法としては、例えば、スタティックミキサーを使用する方法、ダブルシリンジを使用する方法などが、挙げられる。 9. Mixing Method In the composition of the present invention, the mixing method of the component (a) and the component (b) is not particularly limited as long as the component (a) and the component (b) can be mixed uniformly. Examples of the mixing method include a method using a static mixer and a method using a double syringe.
スタティックミキサーは、駆動部のない静的混合装置のことを言う。より具体的には、スタティックミキサーは、通常、管と管内に固定された駆動部の無いミキシング・エレメントからなり、これにより、流れを分割し、かつ流れ方向を転換または反転させ、流れを縦方向、横方向に分割、転換および反転を繰り返すことにより流体を混合する混合装置のことである。スタティックミキサーの種類によっては、管外周部に熱交換の為のジャケットが備えられているものもあり、またミキシング・エレメント自体に熱媒体を通す熱交換の為のチューブが備えられているものもある。スタティックミキサーについての詳細は、例えば、Ｔｈａｋｕｒ ｅｔ ａｌ．，（２００３）Ｔｒａｎｓ ＩＣｈｅｍＥ， ８１， Ｐａｒｔ Ａ：７８７－８２６に記載されており、これを参照することができる。
スタティックミキサーとしては、公知のものを使用することができる、公知のスタティックミキサーとしては、例えば、ケニックス型（例えば、ノードソン社製、およびノリタケ社製）；Ｋｅｎｉｃｓ（Ｃｈｅｍｉｎｅｅｒ Ｉｎｃ．）；ｌｏｗ ｐｒｅｓｓｕｒｅ ｄｒｏｐ（Ｒｏｓｓ Ｅｎｇｉｎｅｅｒｉｎｇ Ｉｎｃ．）；ＳＭＶ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；ＳＭＸ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；ＳＭＸＬ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；Ｉｎｔｅｒｆａｃｉａｌ Ｓｕｒｆａｃｅ Ｇｅｎｅｒａｔｏｒ－ＩＳＧ（Ｒｏｓｓ Ｅｎｇｉｎｅｅｒｉｎｇ Ｉｎｃ．）；ＨＥＶ（Ｃｈｅｍｉｎｅｅｒ Ｉｎｃ．）；Ｉｎｌｉｎｅｒ ｓｅｒｉｅｓ ５０（Ｌｉｇｈｔｎｉｎ Ｉｎｃ．）；Ｉｎｌｉｎｅｒ ｓｅｒｉｅｓ ４５（Ｌｉｇｈｔｎｉｎ Ｉｎｃ．）；Ｃｕｓｔｏｄｙ ｔｒａｎｓｆｅｒ ｍｉｘｅｒ（Ｋｏｍａｘ ｓｙｓｔｅｍｓ Ｉｎｃ．）；およびＳＭＲ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ，Ｉｎｃ．）が挙げられる。 A static mixer refers to a static mixing device without a drive unit. More specifically, a static mixer usually consists of a tube and a mixing element without a drive fixed in the tube, thereby splitting the flow and diverting or reversing the flow direction, so that the flow is longitudinal. It is a mixing device that mixes fluids by repeating division, conversion and inversion in the horizontal direction. Some types of static mixers are equipped with a jacket for heat exchange on the outer periphery of the tube, and others are equipped with a tube for heat exchange through which the heat medium passes through the mixing element itself. . Details on the static mixer can be found in, for example, Thakur et al. (2003) Trans IC ChemE, 81, Part A: 787-826, which can be referred to.
As the static mixer, a known mixer can be used. Examples of the known static mixer include, for example, Kenix type (for example, manufactured by Nordson and Noritake); Kenics (Chemineer Inc.); low pressure drop ( Ross Engineering Inc.); SMV (Koch-Glitsch Inc.); SMXL (Koch-Glitsch Inc.); Interfacial Surface Generator. InC (R) Eng. ); Inliner series 50 (Lightnin Inc.); Inliner series 45 . Lightnin Inc); Custody transfer mixer (Komax systems Inc.);. And SMR (Koch-Glitsch, Inc) and the like.
スタティックミキサーとしては、公知のものを使用することができる、公知のスタティックミキサーとしては、例えば、ケニックス型（例えば、ノードソン社製、およびノリタケ社製）；Ｋｅｎｉｃｓ（Ｃｈｅｍｉｎｅｅｒ Ｉｎｃ．）；ｌｏｗ ｐｒｅｓｓｕｒｅ ｄｒｏｐ（Ｒｏｓｓ Ｅｎｇｉｎｅｅｒｉｎｇ Ｉｎｃ．）；ＳＭＶ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；ＳＭＸ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；ＳＭＸＬ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ Ｉｎｃ．）；Ｉｎｔｅｒｆａｃｉａｌ Ｓｕｒｆａｃｅ Ｇｅｎｅｒａｔｏｒ－ＩＳＧ（Ｒｏｓｓ Ｅｎｇｉｎｅｅｒｉｎｇ Ｉｎｃ．）；ＨＥＶ（Ｃｈｅｍｉｎｅｅｒ Ｉｎｃ．）；Ｉｎｌｉｎｅｒ ｓｅｒｉｅｓ ５０（Ｌｉｇｈｔｎｉｎ Ｉｎｃ．）；Ｉｎｌｉｎｅｒ ｓｅｒｉｅｓ ４５（Ｌｉｇｈｔｎｉｎ Ｉｎｃ．）；Ｃｕｓｔｏｄｙ ｔｒａｎｓｆｅｒ ｍｉｘｅｒ（Ｋｏｍａｘ ｓｙｓｔｅｍｓ Ｉｎｃ．）；およびＳＭＲ（Ｋｏｃｈ－Ｇｌｉｔｓｃｈ，Ｉｎｃ．）が挙げられる。 A static mixer refers to a static mixing device without a drive unit. More specifically, a static mixer usually consists of a tube and a mixing element without a drive fixed in the tube, thereby splitting the flow and diverting or reversing the flow direction, so that the flow is longitudinal. It is a mixing device that mixes fluids by repeating division, conversion and inversion in the horizontal direction. Some types of static mixers are equipped with a jacket for heat exchange on the outer periphery of the tube, and others are equipped with a tube for heat exchange through which the heat medium passes through the mixing element itself. . Details on the static mixer can be found in, for example, Thakur et al. (2003) Trans IC ChemE, 81, Part A: 787-826, which can be referred to.
As the static mixer, a known mixer can be used. Examples of the known static mixer include, for example, Kenix type (for example, manufactured by Nordson and Noritake); Kenics (Chemineer Inc.); low pressure drop ( Ross Engineering Inc.); SMV (Koch-Glitsch Inc.); SMXL (Koch-Glitsch Inc.); Interfacial Surface Generator. InC (R) Eng. ); Inliner series 50 (Lightnin Inc.); Inliner series 45 . Lightnin Inc); Custody transfer mixer (Komax systems Inc.);. And SMR (Koch-Glitsch, Inc) and the like.
ダブルシリンジは、混合する成分を別々に充填するための容器を有し、成分がダブルシリンジから放出されるとき、均一に混ぜ合わされて放出される装置である。
ダブルシリンジとしては、公知のものを使用することができる。公知のダブルシリンジとしては、例えば、バクスター社製のものが挙げられる。
あるいは、スタティックミキサーおよびダブルシリンンジは、本発明の組成物を混合するために作製された特注品であってもよい。 The double syringe is a device that has containers for separately filling the components to be mixed, and is uniformly mixed and released when the components are released from the double syringe.
A well-known thing can be used as a double syringe. Examples of known double syringes include those manufactured by Baxter.
Alternatively, the static mixer and double syringe can be custom made to mix the compositions of the present invention.
ダブルシリンジとしては、公知のものを使用することができる。公知のダブルシリンジとしては、例えば、バクスター社製のものが挙げられる。
あるいは、スタティックミキサーおよびダブルシリンンジは、本発明の組成物を混合するために作製された特注品であってもよい。 The double syringe is a device that has containers for separately filling the components to be mixed, and is uniformly mixed and released when the components are released from the double syringe.
A well-known thing can be used as a double syringe. Examples of known double syringes include those manufactured by Baxter.
Alternatively, the static mixer and double syringe can be custom made to mix the compositions of the present invention.
１０．併用薬
本発明の組成物を外科手術に関連した組織に適用する前に、あるいは同時に、あるいは後で、ストレプトマイシン、ペニシリン、トブラマイシン、アミカシン、ゲンタマイシン、ネオマイシン、およびアンホテリシンＢ等の抗生物質、アスピリン、非ステロイド性解熱鎮痛剤（ＮＳＡＩＤｓ）、アセトアミノフェン等の抗炎症薬等の併用薬を投与するようにしてもよい。これらの薬剤は本発明の組成物に混入して用いてもよい。 10. Concomitant medications Before, simultaneously with, or after applying the compositions of the present invention to the tissue associated with surgery, antibiotics such as streptomycin, penicillin, tobramycin, amikacin, gentamicin, neomycin, and amphotericin B, aspirin, non- Concomitant drugs such as steroidal antipyretic analgesics (NSAIDs) and anti-inflammatory drugs such as acetaminophen may be administered. These agents may be used by mixing in the composition of the present invention.
本発明の組成物を外科手術に関連した組織に適用する前に、あるいは同時に、あるいは後で、ストレプトマイシン、ペニシリン、トブラマイシン、アミカシン、ゲンタマイシン、ネオマイシン、およびアンホテリシンＢ等の抗生物質、アスピリン、非ステロイド性解熱鎮痛剤（ＮＳＡＩＤｓ）、アセトアミノフェン等の抗炎症薬等の併用薬を投与するようにしてもよい。これらの薬剤は本発明の組成物に混入して用いてもよい。 10. Concomitant medications Before, simultaneously with, or after applying the compositions of the present invention to the tissue associated with surgery, antibiotics such as streptomycin, penicillin, tobramycin, amikacin, gentamicin, neomycin, and amphotericin B, aspirin, non- Concomitant drugs such as steroidal antipyretic analgesics (NSAIDs) and anti-inflammatory drugs such as acetaminophen may be administered. These agents may be used by mixing in the composition of the present invention.
なお、本明細書において引用した全ての刊行物、例えば、先行技術文献および公開公報、特許公報その他の特許文献は、その全体が本明細書において参照として組み込まれる。また、本明細書は、本願の優先権主張の基礎となる日本国特許出願である特願2015-006248号(2015年1月15日出願)の特許請求の範囲、明細書、および図面の開示内容を包含する。
It should be noted that all publications cited in the present specification, for example, prior art documents and publications, patent publications and other patent documents, are incorporated herein by reference in their entirety. In addition, this specification is a disclosure of claims, description, and drawings of Japanese Patent Application No. 2015-006248 (filed on Jan. 15, 2015), which is a Japanese patent application that is the basis of the priority claim of the present application. Includes content.
It should be noted that all publications cited in the present specification, for example, prior art documents and publications, patent publications and other patent documents, are incorporated herein by reference in their entirety. In addition, this specification is a disclosure of claims, description, and drawings of Japanese Patent Application No. 2015-006248 (filed on Jan. 15, 2015), which is a Japanese patent application that is the basis of the priority claim of the present application. Includes content.
以下の実施例により本発明を更に詳述するが、本発明はこれら実施例に限定して理解されるべきものではない。
The present invention will be described in further detail with reference to the following examples, but the present invention should not be understood as being limited to these examples.
ラット一部肝切除モデル
ラット一部肝切除モデルを用いて、癒着の形成を評価した。ラット一部肝切除モデルは、重篤な炎症を惹起し高い強度を有する癒着の形成を再現性高く観察できるモデルである（Ｓｈｉｍｉｚｕ Ａ ｅｔ ａｌ．，（２０１４）Ｓｕｒｇ Ｔｏｄａｙ．（４４）：３１４－３２３）。具体的には、以下のように癒着の形成を評価した。 Rat partial hepatectomy model A rat partial hepatectomy model was used to evaluate the formation of adhesions. The rat partial hepatectomy model is a model capable of reproducibly observing the formation of adhesions that cause severe inflammation and have high strength (Shimizu A et al., (2014) Surg Today. (44): 314-314. 323). Specifically, the formation of adhesions was evaluated as follows.
ラット一部肝切除モデルを用いて、癒着の形成を評価した。ラット一部肝切除モデルは、重篤な炎症を惹起し高い強度を有する癒着の形成を再現性高く観察できるモデルである（Ｓｈｉｍｉｚｕ Ａ ｅｔ ａｌ．，（２０１４）Ｓｕｒｇ Ｔｏｄａｙ．（４４）：３１４－３２３）。具体的には、以下のように癒着の形成を評価した。 Rat partial hepatectomy model A rat partial hepatectomy model was used to evaluate the formation of adhesions. The rat partial hepatectomy model is a model capable of reproducibly observing the formation of adhesions that cause severe inflammation and have high strength (Shimizu A et al., (2014) Surg Today. (44): 314-314. 323). Specifically, the formation of adhesions was evaluated as follows.
［材料］
間葉系幹細胞（ＭＳＣ）は、ＳＤラット骨髄由来ＭＳＣを単離し、培養して用いた。
低エンドトキシンアルギン酸ナトリウムは、持田製薬株式会社から入手した。（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ２０（Ｌｏｔ ＮＯ．３Ｈ１９），エンドトキシン量５００ＥＵ／ｇ）
カルボキシメチルセルロース（ＣＭＣ）は、Ｓｉｇｍａ－Ａｌｄｒｉｃｈから入手した。（Ｓｏｄｉｕｍ Ｃａｒｂｏｘｙｍｅｔｈｙｌ ｃｅｌｌｕｌｏｓｅ（Ｓｉｇｍａ）４１９３３８－１００Ｇ，重量平均分子量：７００，０００，置換度０．９，カウンターイオンＮａ型）
Ｓｅｐｒａｆｉｌｍ（商品名）は、カルボキシメチルセルロース（ＣＭＣ）とヒアルロン酸を混合したシート状材料であり、Ｇｅｎｚｙｍｅ ＧｍｂＨより入手した。
Ｉｎｔｅｒｃｅｅｄ（商品名）は、再生酸化セルロースシートであり、Ｊｏｈｎｓｏｎ＆Ｊｏｈｎｓｏｎより入手した。
ダブルシリンジは、バクスター株式会社のＤＵＰＬＯＪＥＣＴ Ｓｙｓｔｅｍを使用した。
アルギン酸ナトリウムおよびＣＭＣは、リン酸緩衝生理食塩水（ＰＢＳ）や生理食塩水（０．９ｗ／ｖ％ ＮａＣｌ）に溶かして溶液として用いた。
ＰＢＳの組成は、以下の通りである。 [material]
As mesenchymal stem cells (MSC), SD rat bone marrow-derived MSCs were isolated and cultured.
Low endotoxin sodium alginate was obtained from Mochida Pharmaceutical. (Low endotoxin sodium alginate (sterile) AL20 (Lot NO. 3H19), endotoxin amount 500 EU / g)
Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich. (Sodium Carboxymethyl Cellulose (Sigma) 419338-100G, weight average molecular weight: 700,000, substitution degree 0.9, counter ion Na type)
Seprafilm (trade name) is a sheet-like material obtained by mixing carboxymethylcellulose (CMC) and hyaluronic acid, and was obtained from Genzyme GmbH.
Interlaced (trade name) is a regenerated oxidized cellulose sheet and was obtained from Johnson & Johnson.
The double syringe used was DUPLOJECT System manufactured by Baxter Corporation.
Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
The composition of PBS is as follows.
間葉系幹細胞（ＭＳＣ）は、ＳＤラット骨髄由来ＭＳＣを単離し、培養して用いた。
低エンドトキシンアルギン酸ナトリウムは、持田製薬株式会社から入手した。（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ２０（Ｌｏｔ ＮＯ．３Ｈ１９），エンドトキシン量５００ＥＵ／ｇ）
カルボキシメチルセルロース（ＣＭＣ）は、Ｓｉｇｍａ－Ａｌｄｒｉｃｈから入手した。（Ｓｏｄｉｕｍ Ｃａｒｂｏｘｙｍｅｔｈｙｌ ｃｅｌｌｕｌｏｓｅ（Ｓｉｇｍａ）４１９３３８－１００Ｇ，重量平均分子量：７００，０００，置換度０．９，カウンターイオンＮａ型）
Ｓｅｐｒａｆｉｌｍ（商品名）は、カルボキシメチルセルロース（ＣＭＣ）とヒアルロン酸を混合したシート状材料であり、Ｇｅｎｚｙｍｅ ＧｍｂＨより入手した。
Ｉｎｔｅｒｃｅｅｄ（商品名）は、再生酸化セルロースシートであり、Ｊｏｈｎｓｏｎ＆Ｊｏｈｎｓｏｎより入手した。
ダブルシリンジは、バクスター株式会社のＤＵＰＬＯＪＥＣＴ Ｓｙｓｔｅｍを使用した。
アルギン酸ナトリウムおよびＣＭＣは、リン酸緩衝生理食塩水（ＰＢＳ）や生理食塩水（０．９ｗ／ｖ％ ＮａＣｌ）に溶かして溶液として用いた。
ＰＢＳの組成は、以下の通りである。 [material]
As mesenchymal stem cells (MSC), SD rat bone marrow-derived MSCs were isolated and cultured.
Low endotoxin sodium alginate was obtained from Mochida Pharmaceutical. (Low endotoxin sodium alginate (sterile) AL20 (Lot NO. 3H19), endotoxin amount 500 EU / g)
Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich. (Sodium Carboxymethyl Cellulose (Sigma) 419338-100G, weight average molecular weight: 700,000, substitution degree 0.9, counter ion Na type)
Seprafilm (trade name) is a sheet-like material obtained by mixing carboxymethylcellulose (CMC) and hyaluronic acid, and was obtained from Genzyme GmbH.
Interlaced (trade name) is a regenerated oxidized cellulose sheet and was obtained from Johnson & Johnson.
The double syringe used was DUPLOJECT System manufactured by Baxter Corporation.
Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
The composition of PBS is as follows.
［実験群］
Ｃｏｎｔｒｏｌ群（ｎ＝４）：ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅの辺縁を３ｃｍ計測し切離し凝固止血した（無処置対照群）。
Ａｌｇｉｎａｔｅ／ＣＭＣ群（ｎ＝４）：低エンドトキシンアルギン酸ナトリウム３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は３．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
Ａｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群（ｎ＝４）：低エンドトキシンアルギン酸３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬに間葉系幹細胞５．０×１０６ｃｅｌｌｓを封入し癒着防止材として適用した。
ＭＳＣ ｉ．ｐ．群（ｎ＝４）：肝切除施術後すみやかに生理食塩水１ｍＬに懸濁した間葉系幹細胞５．０×１０６ｃｅｌｌｓを腹腔投与し閉腹した。
ＭＳＣ ｉ．ｖ．群（ｎ＝４）：肝切除施術後すみやかに生理食塩水１ｍＬに懸濁した間葉系幹細胞５．０×１０６ｃｅｌｌｓを尾静脈注射により投与し閉腹した。
Ｓｅｐｒａｆｉｌｍ群（ｎ＝１０）：２×３ｃｍのＳｅｐｒａｆｉｌｍを癒着防止材として適用した。
Ｉｎｔｅｒｃｅｅｄ群（ｎ＝１０）：２×３ｃｍのＩｎｔｅｒｃｅｅｄを癒着防止材として適用した。 [Experimental group]
Control group (n = 4): The edge of the left lateral lobe was measured 3 cm, separated and coagulated and hemostatic (untreated control group).
Alginate / CMC group (n = 4): Prevention adhesion and low endotoxinalginic acid sodium 3 wt% solution 0.5mL, the Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL Applied as a material. The content of sodium alginate in the Alginate / CMC gel was 3.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
Alginate / CMC + MSC group (n = 4): a lowendotoxin alginic acid 3 wt% solution 0.5mL, mesenchymal in Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL Stem cells 5.0 × 10 6 cells were encapsulated and applied as an adhesion preventive material.
MSC i. p. Group (n = 4): Immediately after hepatectomy, mesenchymal stem cells 5.0 × 10 6 cells suspended in 1 mL of physiological saline were intraperitoneally administered and the abdomen was closed.
MSC i. v. Group (n = 4): Immediately after the hepatectomy, mesenchymal stem cells 5.0 × 10 6 cells suspended in 1 mL of physiological saline were administered by tail vein injection and the abdomen was closed.
Seprafilm group (n = 10): 2 × 3 cm of Seprafilm was applied as an adhesion prevention material.
Interleaved group (n = 10): 2 × 3 cm of Interceed was applied as an adhesion preventing material.
Ｃｏｎｔｒｏｌ群（ｎ＝４）：ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅの辺縁を３ｃｍ計測し切離し凝固止血した（無処置対照群）。
Ａｌｇｉｎａｔｅ／ＣＭＣ群（ｎ＝４）：低エンドトキシンアルギン酸ナトリウム３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は３．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
Ａｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群（ｎ＝４）：低エンドトキシンアルギン酸３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬに間葉系幹細胞５．０×１０６ｃｅｌｌｓを封入し癒着防止材として適用した。
ＭＳＣ ｉ．ｐ．群（ｎ＝４）：肝切除施術後すみやかに生理食塩水１ｍＬに懸濁した間葉系幹細胞５．０×１０６ｃｅｌｌｓを腹腔投与し閉腹した。
ＭＳＣ ｉ．ｖ．群（ｎ＝４）：肝切除施術後すみやかに生理食塩水１ｍＬに懸濁した間葉系幹細胞５．０×１０６ｃｅｌｌｓを尾静脈注射により投与し閉腹した。
Ｓｅｐｒａｆｉｌｍ群（ｎ＝１０）：２×３ｃｍのＳｅｐｒａｆｉｌｍを癒着防止材として適用した。
Ｉｎｔｅｒｃｅｅｄ群（ｎ＝１０）：２×３ｃｍのＩｎｔｅｒｃｅｅｄを癒着防止材として適用した。 [Experimental group]
Control group (n = 4): The edge of the left lateral lobe was measured 3 cm, separated and coagulated and hemostatic (untreated control group).
Alginate / CMC group (n = 4): Prevention adhesion and low endotoxin
Alginate / CMC + MSC group (n = 4): a low
MSC i. p. Group (n = 4): Immediately after hepatectomy, mesenchymal stem cells 5.0 × 10 6 cells suspended in 1 mL of physiological saline were intraperitoneally administered and the abdomen was closed.
MSC i. v. Group (n = 4): Immediately after the hepatectomy, mesenchymal stem cells 5.0 × 10 6 cells suspended in 1 mL of physiological saline were administered by tail vein injection and the abdomen was closed.
Seprafilm group (n = 10): 2 × 3 cm of Seprafilm was applied as an adhesion prevention material.
Interleaved group (n = 10): 2 × 3 cm of Interceed was applied as an adhesion preventing material.
［手順］
ペントバルビタール３５ｍｇ／ｋｇ相当の腹腔投与によりラットに麻酔をかけ、体重を電子天秤で測定した。その後、ラットを、正中切開で開腹した。次に、腹壁をピンセットでつまんで持ち上げ、腹壁を切断した。肝切除を行う前準備として、ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅを腹腔の奥から引き出し、その下にガーゼを敷いた。次に、実際の肝切除に移った。具体的には、ものさしを肝臓にあて、離断面が３ｃｍになる位置を探し、その両端に、バイポーラで焼灼し、印をつけた。印をつけた２点の間を直線的に切除していった。Ｃｏｎｔｒｏｌ群では、この後すぐ閉腹して処置を終了した。癒着防止材を適用する群では、この後、ガーゼを除去したのちに癒着防止材を適用した。ＭＳＣ ｉ．ｐ．群およびＭＳＣ ｉ．ｖ．群では、この後、すみやかに間葉系幹細胞を注射により投与した。次に、腹壁と皮膚を二回に分けて縫合し閉腹した。腹壁を縫合する際には生分解糸を用い、皮膚を縫合する際には非吸収糸を用いた。閉腹から一週間後に、ラットに過剰量の麻酔として約２ｍＬのソムノペンチル原液を投与して屠殺し、体重を電子天秤で測定した。その後、再び開腹し、以下の通り癒着を評価した。 [procedure]
Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points. In the Control group, the abdomen was immediately closed and the treatment was terminated. In the group to which the anti-adhesive material was applied, the anti-adhesive material was applied after removing the gauze. MSC i. p. Group and MSC i. v. In the group, the mesenchymal stem cells were then immediately administered by injection. Next, the abdominal wall and skin were sutured twice and closed. A biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin. One week after laparotomy, rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance. Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
ペントバルビタール３５ｍｇ／ｋｇ相当の腹腔投与によりラットに麻酔をかけ、体重を電子天秤で測定した。その後、ラットを、正中切開で開腹した。次に、腹壁をピンセットでつまんで持ち上げ、腹壁を切断した。肝切除を行う前準備として、ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅを腹腔の奥から引き出し、その下にガーゼを敷いた。次に、実際の肝切除に移った。具体的には、ものさしを肝臓にあて、離断面が３ｃｍになる位置を探し、その両端に、バイポーラで焼灼し、印をつけた。印をつけた２点の間を直線的に切除していった。Ｃｏｎｔｒｏｌ群では、この後すぐ閉腹して処置を終了した。癒着防止材を適用する群では、この後、ガーゼを除去したのちに癒着防止材を適用した。ＭＳＣ ｉ．ｐ．群およびＭＳＣ ｉ．ｖ．群では、この後、すみやかに間葉系幹細胞を注射により投与した。次に、腹壁と皮膚を二回に分けて縫合し閉腹した。腹壁を縫合する際には生分解糸を用い、皮膚を縫合する際には非吸収糸を用いた。閉腹から一週間後に、ラットに過剰量の麻酔として約２ｍＬのソムノペンチル原液を投与して屠殺し、体重を電子天秤で測定した。その後、再び開腹し、以下の通り癒着を評価した。 [procedure]
Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points. In the Control group, the abdomen was immediately closed and the treatment was terminated. In the group to which the anti-adhesive material was applied, the anti-adhesive material was applied after removing the gauze. MSC i. p. Group and MSC i. v. In the group, the mesenchymal stem cells were then immediately administered by injection. Next, the abdominal wall and skin were sutured twice and closed. A biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin. One week after laparotomy, rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance. Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
［癒着の評価］
癒着の評価は、目視により行った。腹腔内の以下の各部位について下記スコアリング法に基づき癒着スコアをつけた。
部位：肝離断面、大網、腹膜、小腸、正中創直下
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着） [Evaluation of adhesions]
Adhesion was evaluated visually. An adhesion score was assigned to each of the following sites in the abdominal cavity based on the following scoring method.
Site: Hepatic section, omentum, peritoneum, small intestine, directly under the median wound Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
癒着の評価は、目視により行った。腹腔内の以下の各部位について下記スコアリング法に基づき癒着スコアをつけた。
部位：肝離断面、大網、腹膜、小腸、正中創直下
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着） [Evaluation of adhesions]
Adhesion was evaluated visually. An adhesion score was assigned to each of the following sites in the abdominal cavity based on the following scoring method.
Site: Hepatic section, omentum, peritoneum, small intestine, directly under the median wound Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
［結果］
癒着の評価の結果を図１および２に示し、体重測定の結果を図３に示す。
癒着の評価については、Ａｌｇｉｎａｔｅ／ＣＭＣ群が一部肝切除モデルにおいて高い癒着防止効果を示した他、このゲルにＭＳＣを封入したＡｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群で、ほぼ同等の高い癒着防止効果を示した（図１および２）。
体重については、Ａｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群では、他の群と比べ高い体重増加率を示し、ＭＳＣの投与により肝切除後の術後ストレスが軽減されたことが示唆された（図３）。また、Ａｌｇｉｎａｔｅ／ＣＭＣ群では、体重の変化があまり見られず、このゲルの適用による悪影響はないことが示唆された。 [result]
The results of adhesion evaluation are shown in FIGS. 1 and 2, and the results of body weight measurement are shown in FIG.
Regarding the evaluation of adhesion, the Alginate / CMC group showed a high adhesion prevention effect in a partial hepatectomy model, and the Alginate / CMC + MSC group in which MSC was encapsulated in this gel showed almost the same high adhesion prevention effect ( Figures 1 and 2).
Regarding the body weight, the Alginate / CMC + MSC group showed a higher rate of weight gain than other groups, suggesting that postoperative hepatic resection stress was reduced by the administration of MSC (FIG. 3). In the Alginate / CMC group, the change in body weight was not so much, suggesting that there was no adverse effect by the application of this gel.
癒着の評価の結果を図１および２に示し、体重測定の結果を図３に示す。
癒着の評価については、Ａｌｇｉｎａｔｅ／ＣＭＣ群が一部肝切除モデルにおいて高い癒着防止効果を示した他、このゲルにＭＳＣを封入したＡｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群で、ほぼ同等の高い癒着防止効果を示した（図１および２）。
体重については、Ａｌｇｉｎａｔｅ／ＣＭＣ＋ＭＳＣ群では、他の群と比べ高い体重増加率を示し、ＭＳＣの投与により肝切除後の術後ストレスが軽減されたことが示唆された（図３）。また、Ａｌｇｉｎａｔｅ／ＣＭＣ群では、体重の変化があまり見られず、このゲルの適用による悪影響はないことが示唆された。 [result]
The results of adhesion evaluation are shown in FIGS. 1 and 2, and the results of body weight measurement are shown in FIG.
Regarding the evaluation of adhesion, the Alginate / CMC group showed a high adhesion prevention effect in a partial hepatectomy model, and the Alginate / CMC + MSC group in which MSC was encapsulated in this gel showed almost the same high adhesion prevention effect ( Figures 1 and 2).
Regarding the body weight, the Alginate / CMC + MSC group showed a higher rate of weight gain than other groups, suggesting that postoperative hepatic resection stress was reduced by the administration of MSC (FIG. 3). In the Alginate / CMC group, the change in body weight was not so much, suggesting that there was no adverse effect by the application of this gel.
ラット一部肝切除モデル
実施例１に記載のラット一部肝切除モデルを用いて、癒着の形成を評価した。具体的には、以下のように癒着の形成を評価した。 Rat partial hepatectomy model The rat partial hepatectomy model described in Example 1 was used to evaluate the formation of adhesions. Specifically, the formation of adhesions was evaluated as follows.
実施例１に記載のラット一部肝切除モデルを用いて、癒着の形成を評価した。具体的には、以下のように癒着の形成を評価した。 Rat partial hepatectomy model The rat partial hepatectomy model described in Example 1 was used to evaluate the formation of adhesions. Specifically, the formation of adhesions was evaluated as follows.
［材料］
低エンドトキシンアルギン酸ナトリウムは、持田製薬株式会社から入手した。（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ２０（Ｌｏｔ ＮＯ．３Ｈ１９），エンドトキシン量７ＥＵ／ｇ）（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ１００（Ｌｏｔ ＮＯ．５Ｈ０８），エンドトキシン量７ＥＵ／ｇ）（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ５００（Ｌｏｔ ＮＯ．３Ｈ１７），エンドトキシン量１５ＥＵ／ｇ）
カルボキシメチルセルロース（ＣＭＣ）は、Ｓｉｇｍａ－Ａｌｄｒｉｃｈから入手した。（Ｓｏｄｉｕｍ Ｃａｒｂｏｘｙｍｅｔｈｙｌ ｃｅｌｌｕｌｏｓｅ（Ｓｉｇｍａ）４１９３３８－１００Ｇ，重量平均分子量：７００，０００，置換度０．９，カウンターイオンＮａ型）
低エンドトキシンＣＭＣは、持田製薬株式会社から入手した。（低エンドトキシンＣＭＣ（非滅菌）（Ｌｏｔ ＮＯ．５Ｃ０４２５１，エンドトキシン量２４ＥＵ／ｇ）
Ｓｅｐｒａｆｉｌｍ（商品名）およびＩｎｔｅｒｃｅｅｄ（商品名）は、実施例１に記載の方法で入手したものを用いた。
ダブルシリンジは、実施例１に記載したものを使用した。
アルギン酸ナトリウムおよびＣＭＣは、リン酸緩衝生理食塩水（ＰＢＳ）や生理食塩水（０．９ｗ／ｖ％ ＮａＣｌ）に溶かして溶液として用いた。
ＰＢＳの組成は、実施例１に記載したものを使用した。
なお、本発明に用いたアルギン酸について、以下の各方法により測定した重量平均分子量を記載した。
［前処理方法］
試料に溶離液を加え溶解後、０．４５μｍメンブランフィルター濾過したものを測定溶液とした。
（１）ゲル浸透クロマトグラフィー（ＧＰＣ）測定
［測定条件（相対分子量分布測定）］
カラム：ＴＳＫｇｅｌ ＧＭＰＷ－ＸＬ×２＋Ｇ２５００ＰＷ－ＸＬ（７．８ｍｍ Ｉ．Ｄ．×３００ｍｍ×３本）
溶離液：２００ｍＭ硝酸ナトリウム水溶液
流量：１．０ｍＬ／ｍｉｎ．
濃度：０．０５％
検出器：ＲＩ検出器
カラム温度：４０℃
注入量：２００μＬ
分子量標準：標準プルラン、グルコース
［結果］
ＡＬ２０ ： ３７０，０００
ＡＬ１００： ８６０，０００
ＡＬ５００：１，５００，０００
（２）ＧＰＣ－ＭＡＬＳ測定
［測定条件（屈折率増分（ｄｎ／ｄｃ）測定）］
示差屈折率計：Ｏｐｔｉｌａｂ Ｔ－ｒＥＸ
測定波長：６５８ｎｍ
測定温度：４０℃
溶媒：２００ｍＭ硝酸ナトリウム水溶液
試料濃度：０．５～２．５ｍｇ／ｍＬ（５濃度）
［測定条件（絶対分子量分布測定）］
カラム：ＴＳＫｇｅｌ ＧＭＰＷ－ＸＬ×２＋Ｇ２５００ＰＷ－ＸＬ（７．８ｍｍ Ｉ．Ｄ．×３００ｍｍ×３本）
溶離液：２００ｍＭ硝酸ナトリウム水溶液
流量：１．０ｍＬ／ｍｉｎ．
濃度：０．０５％
検出器：ＲＩ検出器、光散乱検出器（ＭＡＬＳ）
カラム温度：４０℃
注入量：２００μＬ
［結果］
ＡＬ２０ ： ７２，０００
ＡＬ１００： １５０，０００
ＡＬ５００： ２５０，０００
[material]
Low endotoxin sodium alginate was obtained from Mochida Pharmaceutical. (Low endotoxin sodium alginate (sterile product) AL20 (Lot NO. 3H19), endotoxin amount 7 EU / g) (Low endotoxin sodium alginate (sterile product) AL100 (Lot NO. 5H08), endotoxin amount 7 EU / g) (low endotoxin alginic acid Sodium (sterilized product) AL500 (Lot NO. 3H17),endotoxin amount 15 EU / g)
Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich. (Sodium Carboxymethyl Cellulose (Sigma) 419338-100G, weight average molecular weight: 700,000, substitution degree 0.9, counter ion Na type)
Low endotoxin CMC was obtained from Mochida Pharmaceutical. (Low endotoxin CMC (non-sterile) (Lot NO. 5C04251, endotoxin amount 24 EU / g)
As Seprafilm (trade name) and Interceed (trade name), those obtained by the method described in Example 1 were used.
The double syringe described in Example 1 was used.
Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
The composition of the PBS described in Example 1 was used.
In addition, about the alginic acid used for this invention, the weight average molecular weight measured by each following method was described.
[Pre-processing method]
An eluent was added to the sample and dissolved, and then filtered through a 0.45 μm membrane filter to obtain a measurement solution.
(1) Gel permeation chromatography (GPC) measurement [Measurement conditions (relative molecular weight distribution measurement)]
Column: TSKgel GMPW-XL × 2 + G2500PW-XL (7.8 mm ID × 300 mm × 3)
Eluent: 200 mM sodium nitrate aqueous solution Flow rate: 1.0 mL / min.
Concentration: 0.05%
Detector: RI detector Column temperature: 40 ° C
Injection volume: 200 μL
Molecular weight standards: Standard pullulan, glucose [Result]
AL20: 370,000
AL100: 860,000
AL500: 1,500,000
(2) GPC-MALS measurement [Measurement conditions (refractive index increment (dn / dc) measurement)]
Differential refractometer: Optilab T-rEX
Measurement wavelength: 658 nm
Measurement temperature: 40 ° C
Solvent: 200 mM sodium nitrate aqueous solution Sample concentration: 0.5 to 2.5 mg / mL (5 concentrations)
[Measurement conditions (absolute molecular weight distribution measurement)]
Column: TSKgel GMPW-XL × 2 + G2500PW-XL (7.8 mm ID × 300 mm × 3)
Eluent: 200 mM sodium nitrate aqueous solution Flow rate: 1.0 mL / min.
Concentration: 0.05%
Detector: RI detector, light scattering detector (MALS)
Column temperature: 40 ° C
Injection volume: 200 μL
[result]
AL20: 72,000
AL100: 150,000
AL500: 250,000
低エンドトキシンアルギン酸ナトリウムは、持田製薬株式会社から入手した。（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ２０（Ｌｏｔ ＮＯ．３Ｈ１９），エンドトキシン量７ＥＵ／ｇ）（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ１００（Ｌｏｔ ＮＯ．５Ｈ０８），エンドトキシン量７ＥＵ／ｇ）（低エンドトキシンアルギン酸ナトリウム（滅菌品）ＡＬ５００（Ｌｏｔ ＮＯ．３Ｈ１７），エンドトキシン量１５ＥＵ／ｇ）
カルボキシメチルセルロース（ＣＭＣ）は、Ｓｉｇｍａ－Ａｌｄｒｉｃｈから入手した。（Ｓｏｄｉｕｍ Ｃａｒｂｏｘｙｍｅｔｈｙｌ ｃｅｌｌｕｌｏｓｅ（Ｓｉｇｍａ）４１９３３８－１００Ｇ，重量平均分子量：７００，０００，置換度０．９，カウンターイオンＮａ型）
低エンドトキシンＣＭＣは、持田製薬株式会社から入手した。（低エンドトキシンＣＭＣ（非滅菌）（Ｌｏｔ ＮＯ．５Ｃ０４２５１，エンドトキシン量２４ＥＵ／ｇ）
Ｓｅｐｒａｆｉｌｍ（商品名）およびＩｎｔｅｒｃｅｅｄ（商品名）は、実施例１に記載の方法で入手したものを用いた。
ダブルシリンジは、実施例１に記載したものを使用した。
アルギン酸ナトリウムおよびＣＭＣは、リン酸緩衝生理食塩水（ＰＢＳ）や生理食塩水（０．９ｗ／ｖ％ ＮａＣｌ）に溶かして溶液として用いた。
ＰＢＳの組成は、実施例１に記載したものを使用した。
なお、本発明に用いたアルギン酸について、以下の各方法により測定した重量平均分子量を記載した。
［前処理方法］
試料に溶離液を加え溶解後、０．４５μｍメンブランフィルター濾過したものを測定溶液とした。
（１）ゲル浸透クロマトグラフィー（ＧＰＣ）測定
［測定条件（相対分子量分布測定）］
カラム：ＴＳＫｇｅｌ ＧＭＰＷ－ＸＬ×２＋Ｇ２５００ＰＷ－ＸＬ（７．８ｍｍ Ｉ．Ｄ．×３００ｍｍ×３本）
溶離液：２００ｍＭ硝酸ナトリウム水溶液
流量：１．０ｍＬ／ｍｉｎ．
濃度：０．０５％
検出器：ＲＩ検出器
カラム温度：４０℃
注入量：２００μＬ
分子量標準：標準プルラン、グルコース
［結果］
ＡＬ２０ ： ３７０，０００
ＡＬ１００： ８６０，０００
ＡＬ５００：１，５００，０００
（２）ＧＰＣ－ＭＡＬＳ測定
［測定条件（屈折率増分（ｄｎ／ｄｃ）測定）］
示差屈折率計：Ｏｐｔｉｌａｂ Ｔ－ｒＥＸ
測定波長：６５８ｎｍ
測定温度：４０℃
溶媒：２００ｍＭ硝酸ナトリウム水溶液
試料濃度：０．５～２．５ｍｇ／ｍＬ（５濃度）
［測定条件（絶対分子量分布測定）］
カラム：ＴＳＫｇｅｌ ＧＭＰＷ－ＸＬ×２＋Ｇ２５００ＰＷ－ＸＬ（７．８ｍｍ Ｉ．Ｄ．×３００ｍｍ×３本）
溶離液：２００ｍＭ硝酸ナトリウム水溶液
流量：１．０ｍＬ／ｍｉｎ．
濃度：０．０５％
検出器：ＲＩ検出器、光散乱検出器（ＭＡＬＳ）
カラム温度：４０℃
注入量：２００μＬ
［結果］
ＡＬ２０ ： ７２，０００
ＡＬ１００： １５０，０００
ＡＬ５００： ２５０，０００
[material]
Low endotoxin sodium alginate was obtained from Mochida Pharmaceutical. (Low endotoxin sodium alginate (sterile product) AL20 (Lot NO. 3H19), endotoxin amount 7 EU / g) (Low endotoxin sodium alginate (sterile product) AL100 (Lot NO. 5H08), endotoxin amount 7 EU / g) (low endotoxin alginic acid Sodium (sterilized product) AL500 (Lot NO. 3H17),
Carboxymethylcellulose (CMC) was obtained from Sigma-Aldrich. (Sodium Carboxymethyl Cellulose (Sigma) 419338-100G, weight average molecular weight: 700,000, substitution degree 0.9, counter ion Na type)
Low endotoxin CMC was obtained from Mochida Pharmaceutical. (Low endotoxin CMC (non-sterile) (Lot NO. 5C04251, endotoxin amount 24 EU / g)
As Seprafilm (trade name) and Interceed (trade name), those obtained by the method described in Example 1 were used.
The double syringe described in Example 1 was used.
Sodium alginate and CMC were dissolved in phosphate buffered saline (PBS) and physiological saline (0.9 w / v% NaCl) and used as a solution.
The composition of the PBS described in Example 1 was used.
In addition, about the alginic acid used for this invention, the weight average molecular weight measured by each following method was described.
[Pre-processing method]
An eluent was added to the sample and dissolved, and then filtered through a 0.45 μm membrane filter to obtain a measurement solution.
(1) Gel permeation chromatography (GPC) measurement [Measurement conditions (relative molecular weight distribution measurement)]
Column: TSKgel GMPW-XL × 2 + G2500PW-XL (7.8 mm ID × 300 mm × 3)
Eluent: 200 mM sodium nitrate aqueous solution Flow rate: 1.0 mL / min.
Concentration: 0.05%
Detector: RI detector Column temperature: 40 ° C
Injection volume: 200 μL
Molecular weight standards: Standard pullulan, glucose [Result]
AL20: 370,000
AL100: 860,000
AL500: 1,500,000
(2) GPC-MALS measurement [Measurement conditions (refractive index increment (dn / dc) measurement)]
Differential refractometer: Optilab T-rEX
Measurement wavelength: 658 nm
Measurement temperature: 40 ° C
Solvent: 200 mM sodium nitrate aqueous solution Sample concentration: 0.5 to 2.5 mg / mL (5 concentrations)
[Measurement conditions (absolute molecular weight distribution measurement)]
Column: TSKgel GMPW-XL × 2 + G2500PW-XL (7.8 mm ID × 300 mm × 3)
Eluent: 200 mM sodium nitrate aqueous solution Flow rate: 1.0 mL / min.
Concentration: 0.05%
Detector: RI detector, light scattering detector (MALS)
Column temperature: 40 ° C
Injection volume: 200 μL
[result]
AL20: 72,000
AL100: 150,000
AL500: 250,000
［実験群］
Ｃｏｎｔｒｏｌ群（ｎ＝８）：ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅの辺縁を３ｃｍ計測し切離し凝固止血した（無処置対照群）。
Ａｌｇｉｎａｔｅ／ＣＭＣ群（ｎ＝５～９）：ＡＬ２０およびＡＬ１００については、アルギン酸ナトリウム３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液（１００ｍＭＣａＣｌ２と１ｗｔ％ＣＭＣとを含む溶液を意味する、以下同様）０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は３．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
ＡＬ５００については、アルギン酸ナトリウム１ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は１．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
Ａｌｇｉｎａｔｅ／ＣＭＣ群は、具体的には、以下の４つの群を用いた。
（１）ＡＬ２０（－）／ＣＭＣ（＋）：低エンドトキシン処理したアルギン酸（ＡＬ２０）と低エンドトキシン処理していないＣＭＣとの組合せ。
（２）ＡＬ２０（－）／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ２０）と低エンドトキシン処理したＣＭＣとの組合せ。
（３）ＡＬ１００／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ１００）と低エンドトキシン処理したＣＭＣとの組合せ
（４）ＡＬ５００－１％／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ５００；１％）と低エンドトキシン処理したＣＭＣとの組合せ
Ｓｅｐｒａｆｉｌｍ群（ｎ＝８）：２×３ｃｍのＳｅｐｒａｆｉｌｍを癒着防止材として適用した。
Ｉｎｔｅｒｃｅｅｄ群（ｎ＝８）：２×３ｃｍのＩｎｔｅｒｃｅｅｄを癒着防止材として適用した。 [Experimental group]
Control group (n = 8): The margin of the left lateral lobe was measured 3 cm, separated and coagulated and hemostatic (untreated control group).
Alginate / CMC group (n = 5 ~ 9): About AL20 and AL100 means asodium alginate 3 wt% solution 0.5 mL, a solution containing the 100mMCaCl 2 / 1wt% CMC solution (100mMCaCl 2 and 1 wt% CMC, The same applies hereinafter) 1 mL of Alginate / CMC gel prepared by mixing 0.5 mL with a double syringe was applied as an adhesion preventing material. The content of sodium alginate in the Alginate / CMC gel was 3.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
The AL500, asodium alginate 1 wt% solution 0.5mL, was applied Alginate / CMC gel 1mL prepared by mixing the double syringe 100mMCaCl 2 / 1wt% CMC solution 0.5mL as antiadhesive material. The content of sodium alginate in the Alginate / CMC gel was 1.0 wt%, the content of CMC was 1.0 wt%, and the content of CaCl 2 was 1.1 wt%.
Specifically, the following four groups were used as the Alginate / CMC group.
(1) AL20 (−) / CMC (+): a combination of alginic acid treated with low endotoxin (AL20) and CMC not treated with low endotoxin.
(2) AL20 (−) / CMC (−): Combination of low endotoxin-treated alginic acid (AL20) and low endotoxin-treated CMC.
(3) AL100 / CMC (−): Combination of low endotoxin-treated alginic acid (AL100) and low endotoxin-treated CMC (4) AL500-1% / CMC (−): Low endotoxin-treated alginic acid (AL500; 1% ) And low endotoxin-treated CMC Seprafilm group (n = 8): 2 × 3 cm of Seprafilm was applied as an anti-adhesion material.
Interleaved group (n = 8): 2 × 3 cm of Interceed was applied as an anti-adhesion material.
Ｃｏｎｔｒｏｌ群（ｎ＝８）：ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅの辺縁を３ｃｍ計測し切離し凝固止血した（無処置対照群）。
Ａｌｇｉｎａｔｅ／ＣＭＣ群（ｎ＝５～９）：ＡＬ２０およびＡＬ１００については、アルギン酸ナトリウム３ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液（１００ｍＭＣａＣｌ２と１ｗｔ％ＣＭＣとを含む溶液を意味する、以下同様）０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は３．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
ＡＬ５００については、アルギン酸ナトリウム１ｗｔ％溶液０．５ｍＬと、１００ｍＭＣａＣｌ２／１ｗｔ％ＣＭＣ溶液０．５ｍＬをダブルシリンジにより混合することにより作製したＡｌｇｉｎａｔｅ／ＣＭＣゲル１ｍＬを癒着防止材として適用した。Ａｌｇｉｎａｔｅ／ＣＭＣゲル中でのアルギン酸ナトリウムの含有量は１．０ｗｔ％であり、ＣＭＣの含有量は、１．０ｗｔ％であり、ＣａＣｌ２の含有量は１．１ｗｔ％であった。
Ａｌｇｉｎａｔｅ／ＣＭＣ群は、具体的には、以下の４つの群を用いた。
（１）ＡＬ２０（－）／ＣＭＣ（＋）：低エンドトキシン処理したアルギン酸（ＡＬ２０）と低エンドトキシン処理していないＣＭＣとの組合せ。
（２）ＡＬ２０（－）／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ２０）と低エンドトキシン処理したＣＭＣとの組合せ。
（３）ＡＬ１００／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ１００）と低エンドトキシン処理したＣＭＣとの組合せ
（４）ＡＬ５００－１％／ＣＭＣ（－）：低エンドトキシン処理したアルギン酸（ＡＬ５００；１％）と低エンドトキシン処理したＣＭＣとの組合せ
Ｓｅｐｒａｆｉｌｍ群（ｎ＝８）：２×３ｃｍのＳｅｐｒａｆｉｌｍを癒着防止材として適用した。
Ｉｎｔｅｒｃｅｅｄ群（ｎ＝８）：２×３ｃｍのＩｎｔｅｒｃｅｅｄを癒着防止材として適用した。 [Experimental group]
Control group (n = 8): The margin of the left lateral lobe was measured 3 cm, separated and coagulated and hemostatic (untreated control group).
Alginate / CMC group (n = 5 ~ 9): About AL20 and AL100 means a
The AL500, a
Specifically, the following four groups were used as the Alginate / CMC group.
(1) AL20 (−) / CMC (+): a combination of alginic acid treated with low endotoxin (AL20) and CMC not treated with low endotoxin.
(2) AL20 (−) / CMC (−): Combination of low endotoxin-treated alginic acid (AL20) and low endotoxin-treated CMC.
(3) AL100 / CMC (−): Combination of low endotoxin-treated alginic acid (AL100) and low endotoxin-treated CMC (4) AL500-1% / CMC (−): Low endotoxin-treated alginic acid (AL500; 1% ) And low endotoxin-treated CMC Seprafilm group (n = 8): 2 × 3 cm of Seprafilm was applied as an anti-adhesion material.
Interleaved group (n = 8): 2 × 3 cm of Interceed was applied as an anti-adhesion material.
［手順］
ペントバルビタール３５ｍｇ／ｋｇ相当の腹腔投与によりラットに麻酔をかけ、体重を電子天秤で測定した。その後、ラットを、正中切開で開腹した。次に、腹壁をピンセットでつまんで持ち上げ、腹壁を切断した。肝切除を行う前準備として、ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅを腹腔の奥から引き出し、その下にガーゼを敷いた。次に、実際の肝切除に移った。具体的には、ものさしを肝臓にあて、離断面が３ｃｍになる位置を探し、その両端に、バイポーラで焼灼し、印をつけた。印をつけた２点の間を直線的に切除していった。Ｃｏｎｔｒｏｌ群では、この後すぐ閉腹して処置を終了した。癒着防止材を適用する群では、この後、ガーゼを除去したのちに癒着防止材を適用した。次に、腹壁と皮膚を二回に分けて縫合し閉腹した。腹壁を縫合する際には生分解糸を用い、皮膚を縫合する際には非吸収糸を用いた。閉腹から一週間後に、ラットに過剰量の麻酔として約２ｍＬのソムノペンチル原液を投与して屠殺し、体重を電子天秤で測定した。
その後、再び開腹し、以下の通り癒着を評価した。 [procedure]
Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points. In the Control group, the abdomen was immediately closed and the treatment was terminated. In the group to which the anti-adhesive material was applied, the anti-adhesive material was applied after removing the gauze. Next, the abdominal wall and skin were sutured twice and closed. A biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin. One week after laparotomy, rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance.
Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
ペントバルビタール３５ｍｇ／ｋｇ相当の腹腔投与によりラットに麻酔をかけ、体重を電子天秤で測定した。その後、ラットを、正中切開で開腹した。次に、腹壁をピンセットでつまんで持ち上げ、腹壁を切断した。肝切除を行う前準備として、ｌｅｆｔ ｌａｔｅｒａｌ ｌｏｂｅを腹腔の奥から引き出し、その下にガーゼを敷いた。次に、実際の肝切除に移った。具体的には、ものさしを肝臓にあて、離断面が３ｃｍになる位置を探し、その両端に、バイポーラで焼灼し、印をつけた。印をつけた２点の間を直線的に切除していった。Ｃｏｎｔｒｏｌ群では、この後すぐ閉腹して処置を終了した。癒着防止材を適用する群では、この後、ガーゼを除去したのちに癒着防止材を適用した。次に、腹壁と皮膚を二回に分けて縫合し閉腹した。腹壁を縫合する際には生分解糸を用い、皮膚を縫合する際には非吸収糸を用いた。閉腹から一週間後に、ラットに過剰量の麻酔として約２ｍＬのソムノペンチル原液を投与して屠殺し、体重を電子天秤で測定した。
その後、再び開腹し、以下の通り癒着を評価した。 [procedure]
Rats were anesthetized by intraperitoneal administration equivalent to 35 mg / kg pentobarbital, and the body weight was measured with an electronic balance. The rats were then laparotomized with a midline incision. Next, the abdominal wall was pinched with tweezers and lifted to cut the abdominal wall. As a preparation prior to performing a hepatectomy, a left lateral lobe was drawn from the back of the abdominal cavity and a gauze was laid under it. Next, we moved on to actual hepatectomy. Specifically, a ruler was applied to the liver, a position where the separation cross-section was 3 cm was searched, and both ends were cauterized by bipolar and marked. A straight line was cut between the marked two points. In the Control group, the abdomen was immediately closed and the treatment was terminated. In the group to which the anti-adhesive material was applied, the anti-adhesive material was applied after removing the gauze. Next, the abdominal wall and skin were sutured twice and closed. A biodegradable thread was used for suturing the abdominal wall, and a non-absorbable thread was used for suturing the skin. One week after laparotomy, rats were sacrificed by administering about 2 mL of somnopentyl stock solution as an excessive amount of anesthesia, and the body weight was measured with an electronic balance.
Thereafter, the abdomen was opened again, and the adhesion was evaluated as follows.
［癒着の評価］
以下の通り、癒着の評価を行った。
（１）離断面
上記［手順］に記載した肝離断面について、以下の（ａ）、（ｂ）の評価を行った。
（ａ）癒着グレード
癒着の評価は、目視により行った。肝離断面について、下記スコアリング法に基づき癒着スコアをつけた。
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着）
（ｂ）癒着Ｅｘｔｅｎｔ
３ｃｍの肝離断面のうち、癒着が形成された幅をものさしをあてて計測し、ｍｍで表した（従って、離断面の最大Ｅｘｔｅｎｔは３０ｍｍとなる）。
（２）非離断面
肝離断面以外、具体的には肝表面、大網、腹膜、小腸、正中創直下、等について、以下の（ａ）、（ｂ）の評価を行った。
（ａ）癒着グレード
癒着の評価は、目視により行った。肝離断面以外の部位について、下記スコアリング法に基づき癒着スコアをつけた。部位は特定せず、認められた癒着スコアの最大値を、その試験動物の癒着スコアとして記録した。
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着）
（ｂ）癒着Ｅｘｔｅｎｔ
肝離断面以外の部位について、癒着が形成された幅をものさしをあてて計測し、ｍｍで表した。上記（２）（ａ）と同様、部位は特定せず、認められた癒着が形成された幅の最大値を、その試験動物の癒着Ｅｘｔｅｎｔとして記録した。
[Evaluation of adhesions]
Adhesion was evaluated as follows.
(1) Separation Section The following (a) and (b) were evaluated for the liver separation section described in [Procedure] above.
(A) Adhesion grade Adhesion was evaluated visually. An adhesion score was assigned to the hepatic section based on the following scoring method.
Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
(B) Adhesive Extent
Of the 3 cm hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm (thus, the maximum extent of the section is 30 mm).
(2) Non-separation section Other than the hepatic section, the following evaluations (a) and (b) were performed on the liver surface, omentum, peritoneum, small intestine, directly under the midline, and the like.
(A) Adhesion grade Adhesion was evaluated visually. Adhesion scores were assigned based on the following scoring method for sites other than the hepatic section. The site was not specified, and the maximum value of the observed adhesion score was recorded as the adhesion score of the test animal.
Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
(B) Adhesive Extent
For sites other than the hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm. Similar to (2) and (a) above, the site was not specified, and the maximum value of the width at which the observed adhesion was formed was recorded as the adhesion extent of the test animal.
以下の通り、癒着の評価を行った。
（１）離断面
上記［手順］に記載した肝離断面について、以下の（ａ）、（ｂ）の評価を行った。
（ａ）癒着グレード
癒着の評価は、目視により行った。肝離断面について、下記スコアリング法に基づき癒着スコアをつけた。
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着）
（ｂ）癒着Ｅｘｔｅｎｔ
３ｃｍの肝離断面のうち、癒着が形成された幅をものさしをあてて計測し、ｍｍで表した（従って、離断面の最大Ｅｘｔｅｎｔは３０ｍｍとなる）。
（２）非離断面
肝離断面以外、具体的には肝表面、大網、腹膜、小腸、正中創直下、等について、以下の（ａ）、（ｂ）の評価を行った。
（ａ）癒着グレード
癒着の評価は、目視により行った。肝離断面以外の部位について、下記スコアリング法に基づき癒着スコアをつけた。部位は特定せず、認められた癒着スコアの最大値を、その試験動物の癒着スコアとして記録した。
癒着スコア：
Ｇｒａｄｅ０：癒着が全く見られない。
Ｇｒａｄｅ１：自重により剥離する程度の癒着（生理的な癒着）
Ｇｒａｄｅ２：ピンセットによる剥離が可能な癒着（鈍的な癒着）
Ｇｒａｄｅ３：ハサミ、メスを使わないと剥離できない癒着（鋭的な癒着）
（ｂ）癒着Ｅｘｔｅｎｔ
肝離断面以外の部位について、癒着が形成された幅をものさしをあてて計測し、ｍｍで表した。上記（２）（ａ）と同様、部位は特定せず、認められた癒着が形成された幅の最大値を、その試験動物の癒着Ｅｘｔｅｎｔとして記録した。
[Evaluation of adhesions]
Adhesion was evaluated as follows.
(1) Separation Section The following (a) and (b) were evaluated for the liver separation section described in [Procedure] above.
(A) Adhesion grade Adhesion was evaluated visually. An adhesion score was assigned to the hepatic section based on the following scoring method.
Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
(B) Adhesive Extent
Of the 3 cm hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm (thus, the maximum extent of the section is 30 mm).
(2) Non-separation section Other than the hepatic section, the following evaluations (a) and (b) were performed on the liver surface, omentum, peritoneum, small intestine, directly under the midline, and the like.
(A) Adhesion grade Adhesion was evaluated visually. Adhesion scores were assigned based on the following scoring method for sites other than the hepatic section. The site was not specified, and the maximum value of the observed adhesion score was recorded as the adhesion score of the test animal.
Adhesion score:
Grade 0: No adhesion is seen at all.
Grade 1: Adhesion to the extent that it peels off due to its own weight (physiological adhesion)
Grade 2: Adhesion that can be peeled off with tweezers (blunt adhesion)
Grade 3: Adhesion that cannot be removed without using scissors and scalpel (sharp adhesion)
(B) Adhesive Extent
For sites other than the hepatic section, the width at which adhesions were formed was measured with a ruler and expressed in mm. Similar to (2) and (a) above, the site was not specified, and the maximum value of the width at which the observed adhesion was formed was recorded as the adhesion extent of the test animal.
［結果］
癒着の評価の結果を図４（離断面）および図５（非離断面）に示した。
離断面においては、各群とも、Ｃｏｎｔｒｏｌ群に比べて癒着が抑制される傾向が認められた（図４（ａ）および（ｂ））。
非離断面において、本発明のＡｌｇｉｎａｔｅ／ＣＭＣゲルによる、顕著な癒着防止効果が確認された（図５（ａ）および（ｂ））。陽性対照として用いたＳｅｐｒａｆｉｌｍ（商品名）およびＩｎｔｅｒｃｅｅｄ（商品名）では癒着防止効果は認められず、Ｃｏｎｔｒｏｌ群に比べて癒着が増悪する傾向が認められた。一方、Ａｌｇｉｎａｔｅ／ＣＭＣゲルでは、各群とも優位な癒着防止効果が確認された。また、低エンドトキシン処理していないＣＭＣに比べ、低エンドトキシン処理したＣＭＣを用いることにより、癒着防止効果が高まる傾向が認められた。
体重については、Ｃｏｎｔｒｏｌ群、陽性対照群、およびＡｌｇｉｎａｔｅ／ＣＭＣ群間で有意差はなく、本発明のＡｌｇｉｎａｔｅ／ＣＭＣゲルの適用による悪影響はないことが示唆された。
[result]
The results of adhesion evaluation are shown in FIG. 4 (separated section) and FIG. 5 (non-separated section).
In the separation plane, in each group, there was a tendency for adhesion to be suppressed as compared to the Control group (FIGS. 4A and 4B).
In the non-separated cross section, the remarkable adhesion preventing effect by the Alginate / CMC gel of the present invention was confirmed (FIGS. 5A and 5B). In the case of Seprafilm (trade name) and Interceed (trade name) used as positive controls, no adhesion prevention effect was observed, and a tendency for adhesion to worsen was observed compared to the Control group. On the other hand, with the Alginate / CMC gel, a superior adhesion prevention effect was confirmed in each group. Moreover, the tendency for the adhesion prevention effect to increase was recognized by using CMC treated with low endotoxin as compared with CMC not treated with low endotoxin.
Regarding body weight, there was no significant difference among the Control group, the positive control group, and the Alginate / CMC group, suggesting that there was no adverse effect due to the application of the Alginate / CMC gel of the present invention.
癒着の評価の結果を図４（離断面）および図５（非離断面）に示した。
離断面においては、各群とも、Ｃｏｎｔｒｏｌ群に比べて癒着が抑制される傾向が認められた（図４（ａ）および（ｂ））。
非離断面において、本発明のＡｌｇｉｎａｔｅ／ＣＭＣゲルによる、顕著な癒着防止効果が確認された（図５（ａ）および（ｂ））。陽性対照として用いたＳｅｐｒａｆｉｌｍ（商品名）およびＩｎｔｅｒｃｅｅｄ（商品名）では癒着防止効果は認められず、Ｃｏｎｔｒｏｌ群に比べて癒着が増悪する傾向が認められた。一方、Ａｌｇｉｎａｔｅ／ＣＭＣゲルでは、各群とも優位な癒着防止効果が確認された。また、低エンドトキシン処理していないＣＭＣに比べ、低エンドトキシン処理したＣＭＣを用いることにより、癒着防止効果が高まる傾向が認められた。
体重については、Ｃｏｎｔｒｏｌ群、陽性対照群、およびＡｌｇｉｎａｔｅ／ＣＭＣ群間で有意差はなく、本発明のＡｌｇｉｎａｔｅ／ＣＭＣゲルの適用による悪影響はないことが示唆された。
[result]
The results of adhesion evaluation are shown in FIG. 4 (separated section) and FIG. 5 (non-separated section).
In the separation plane, in each group, there was a tendency for adhesion to be suppressed as compared to the Control group (FIGS. 4A and 4B).
In the non-separated cross section, the remarkable adhesion preventing effect by the Alginate / CMC gel of the present invention was confirmed (FIGS. 5A and 5B). In the case of Seprafilm (trade name) and Interceed (trade name) used as positive controls, no adhesion prevention effect was observed, and a tendency for adhesion to worsen was observed compared to the Control group. On the other hand, with the Alginate / CMC gel, a superior adhesion prevention effect was confirmed in each group. Moreover, the tendency for the adhesion prevention effect to increase was recognized by using CMC treated with low endotoxin as compared with CMC not treated with low endotoxin.
Regarding body weight, there was no significant difference among the Control group, the positive control group, and the Alginate / CMC group, suggesting that there was no adverse effect due to the application of the Alginate / CMC gel of the present invention.
Claims (22)
- （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる癒着防止用組成物。 (A) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) an anti-adhesion composition using a combination of cells.
- 前記細胞が、間葉系幹細胞である、請求項１に記載の組成物。 The composition according to claim 1, wherein the cells are mesenchymal stem cells.
- 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、請求項１または２に記載の組成物。 The composition according to claim 1 or 2, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
- 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、請求項１～３のいずれか１項に記載の組成物。 The composition according to any one of claims 1 to 3, wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
- 前記組成物が、流動性を有する液体状である、請求項１～４のいずれか１項に記載の組成物。 The composition according to any one of claims 1 to 4, wherein the composition is in a liquid form having fluidity.
- 前記組成物の粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、請求項５に記載の組成物。 The composition according to claim 5, wherein the viscosity of the composition is 5 mPa · s to 100,000 mPa · s.
- さらに、アルギン酸の１価金属塩の硬化剤を含有する、請求項１～４のいずれか１項に記載の組成物。 The composition according to any one of claims 1 to 4, further comprising a curing agent of a monovalent metal salt of alginic acid.
- （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースおよびその薬学的に許容される塩を組合せて用い、かつ（ｃ）細胞を用いない、癒着防止用組成物。 (A) A composition for preventing adhesion, which uses a combination of a monovalent metal salt of low endotoxin alginic acid, and (b) carboxymethylcellulose and a pharmaceutically acceptable salt thereof, and (c) does not use cells.
- 前記アルギン酸の１価金属塩のエンドトキシン含有量が、５００ＥＵ／ｇ以下である、請求項８に記載の組成物。 The composition according to claim 8, wherein the endotoxin content of the monovalent metal salt of alginic acid is 500 EU / g or less.
- 前記アルギン酸の１価金属塩が、アルギン酸ナトリウムまたはアルギン酸カリウムである、請求項８または９に記載の組成物。 The composition according to claim 8 or 9, wherein the monovalent metal salt of alginic acid is sodium alginate or potassium alginate.
- 前記組成物が、流動性を有する液体状；シート状、スポンジ状、粉体およびパウダー状からなる群から選択される固形物状；半固形状；またはゲル状である、請求項８～１０のいずれか１項に記載の組成物。 The composition according to any one of claims 8 to 10, wherein the composition is in a liquid form having fluidity; a solid form selected from the group consisting of a sheet form, a sponge form, a powder form and a powder form; a semi-solid form; or a gel form. The composition according to any one of the above.
- 前記組成物が、流動性を有する液体状であり、その粘度が、５ｍＰａ・ｓ～１０００００ｍＰａ・ｓである、請求項１１に記載の組成物。 The composition according to claim 11, wherein the composition is a fluid liquid and has a viscosity of 5 mPa · s to 100,000 mPa · s.
- さらに、アルギン酸の１価金属塩の硬化剤を含有する、請求項８～１１のいずれか１項に記載の組成物。 The composition according to any one of claims 8 to 11, further comprising a curing agent of a monovalent metal salt of alginic acid.
- （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞を組合せて用いる組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。 A composition using a combination of (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell is applied to a subject in need of adhesion prevention. A method for preventing adhesions.
- （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩から選択される増粘剤を組合せて用い、かつ（ｃ）細胞を用いない組成物を、癒着防止を必要とする対象に適用することを含む、癒着防止方法。 A combination of (a) a monovalent metal salt of low endotoxin alginic acid, and (b) a thickener selected from carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a cell-free composition. An anti-adhesion method comprising applying to a subject in need of anti-adhesion.
- （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および（ｃ）細胞と組合せて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。 (B) A composition for preventing adhesion containing carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) a monovalent metal salt of low endotoxin alginic acid used in combination with cells.
- （ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩と組合せて用いられ、かつ（ｃ）細胞を用いない、（ａ）低エンドトキシンアルギン酸の１価金属塩を含有する癒着防止用組成物。 (B) An anti-adhesion composition containing a monovalent metal salt of low endotoxin alginic acid, which is used in combination with carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and (c) does not use cells.
- 請求項１～１３、１６および１７のいずれか１項に記載の組成物を製造するために用いられる、低エンドトキシンアルギン酸の１価金属塩を含有する製剤。 A preparation containing a monovalent metal salt of low endotoxin alginic acid, which is used for producing the composition according to any one of claims 1 to 13, 16 and 17.
- （ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。 An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent.
- （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、（ｂ）カルボキシメチルセルロースまたはその薬学的に許容される塩、および硬化剤を含む癒着防止用キット。 (C) An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof, and a curing agent, used in combination with cells.
- （ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。 An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid, and (b) carboxymethylcellulose containing a curing agent or a pharmaceutically acceptable salt thereof.
- （ｃ）細胞と組み合わせて用いられる、（ａ）低エンドトキシンアルギン酸の１価金属塩、および（ｂ）硬化剤を含むカルボキシメチルセルロースまたはその薬学的に許容される塩を含む癒着防止用キット。
(C) An anti-adhesion kit comprising (a) a monovalent metal salt of low endotoxin alginic acid used in combination with cells, and (b) carboxymethylcellulose or a pharmaceutically acceptable salt thereof containing a curing agent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2016569507A JPWO2016114355A1 (en) | 2015-01-15 | 2016-01-14 | Anti-adhesion composition |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2015-006248 | 2015-01-15 | ||
| JP2015006248 | 2015-01-15 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| WO2016114355A1 true WO2016114355A1 (en) | 2016-07-21 |
Family
ID=56405895
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/JP2016/050998 WO2016114355A1 (en) | 2015-01-15 | 2016-01-14 | Composition for preventing adhesion |
Country Status (2)
| Country | Link |
|---|---|
| JP (1) | JPWO2016114355A1 (en) |
| WO (1) | WO2016114355A1 (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018164128A1 (en) * | 2017-03-07 | 2018-09-13 | 持田製薬株式会社 | Alginate liquid formulation |
| WO2019093505A1 (en) * | 2017-11-13 | 2019-05-16 | 持田製薬株式会社 | Composition for preventing adhesion |
| WO2019138583A1 (en) * | 2018-01-15 | 2019-07-18 | 持田製薬株式会社 | Anti-adhesion composition |
| CN110520165A (en) * | 2017-03-02 | 2019-11-29 | 持田制药株式会社 | Fibrocartilage tissue injury in treating composition |
| US11464597B2 (en) | 2016-07-13 | 2022-10-11 | The University Of Tokyo | Adhesion-preventing composition |
| WO2023282247A1 (en) * | 2021-07-05 | 2023-01-12 | 国立大学法人滋賀医科大学 | Tissue formation agent |
| TWI837085B (en) | 2017-03-07 | 2024-04-01 | 日商持田製藥股份有限公司 | Alginic acid liquid preparation |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001005370A1 (en) * | 1999-07-16 | 2001-01-25 | Samyang Corporation | Antiadhesion barriers containing water-soluble alginate and carboxymethyl cellulose as major components and preparation method thereof |
| JP2005000608A (en) * | 2003-06-11 | 2005-01-06 | Mitsuo Okano | Cultured cell sheet having high take capability and method for manufacturing and using the same |
| JP2007075425A (en) * | 2005-09-15 | 2007-03-29 | Mochida Pharmaceut Co Ltd | Wound covering material including alginic acid |
| JP2008515927A (en) * | 2004-10-12 | 2008-05-15 | エフエムシー バイオポリマー エイエス | Self-gelling alginate system and use thereof |
| WO2014115776A1 (en) * | 2013-01-22 | 2014-07-31 | 国立大学法人東京大学 | Adhesion preventing material |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU3065192A (en) * | 1991-12-20 | 1993-07-28 | Howmedica Inc. | Ultra-pure polysaccharide materials for medical use |
| WO2000029449A1 (en) * | 1998-11-13 | 2000-05-25 | Cp Kelco U.S. Inc. | Biopolymer salts with low endotoxin levels, biopolymer compositions thereof and methods of making the same |
| JP4198394B2 (en) * | 2001-06-15 | 2008-12-17 | グンゼ株式会社 | Anti-adhesive material |
-
2016
- 2016-01-14 JP JP2016569507A patent/JPWO2016114355A1/en active Pending
- 2016-01-14 WO PCT/JP2016/050998 patent/WO2016114355A1/en active Application Filing
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001005370A1 (en) * | 1999-07-16 | 2001-01-25 | Samyang Corporation | Antiadhesion barriers containing water-soluble alginate and carboxymethyl cellulose as major components and preparation method thereof |
| JP2005000608A (en) * | 2003-06-11 | 2005-01-06 | Mitsuo Okano | Cultured cell sheet having high take capability and method for manufacturing and using the same |
| JP2008515927A (en) * | 2004-10-12 | 2008-05-15 | エフエムシー バイオポリマー エイエス | Self-gelling alginate system and use thereof |
| JP2007075425A (en) * | 2005-09-15 | 2007-03-29 | Mochida Pharmaceut Co Ltd | Wound covering material including alginic acid |
| WO2014115776A1 (en) * | 2013-01-22 | 2014-07-31 | 国立大学法人東京大学 | Adhesion preventing material |
Non-Patent Citations (7)
| Title |
|---|
| CHATURVEDI A.A. ET AL.: "Prevention of postsurgical adhesions using an ultrapure alginate-based gel.", BRITISH JOURNAL OF SURGERY, vol. 100, no. 7, 2013, pages 904 - 910, ISSN: 0007-1323 * |
| HIROYOSHI MIZOTE ET AL.: "POSTOPERATIVE ADHESIVE ILEUS", THE JOURNAL OF THE JAPAN SURGICAL ASSOCIATION, vol. 40, no. 1, 1979, pages 131 - 138, ISSN: 0386-9776 * |
| KIM T. ET AL.: "A randomized, multi-center, clinical trial to assess the efficacy and safety of alginate carboxymethylcellulose hyaluronic acid compared to carboxymethylcellulose hyaluronic acid to prevent postoperative intrauterine adhesion.", JOURNAL OF MINIMALLY INVASIVE GYNECOLOGY, vol. 19, no. 6, 2012, pages 731 - 736, ISSN: 1553-4650 * |
| KIM Y.D. ET AL.: "Effects of human adipose- derived stem cells on the regeneration of damaged visceral pleural mesothelial cells: a morphological study in a rabbit model.", INTERACTIVE CARDIOVASCULAR AND THORACIC SURGERY, vol. 19, no. 3, 2014, pages 363 - 367, ISSN: 1569-9293 * |
| KUNIO KAWANISHI ET AL., FUKUMAKU SAIBO SHEET NO KAIHATSU TO JUTSUGO CHOKAN YUCHAKU MODEL NI OKERU YUCHAKU BOSHI KOKA NO KENTO, vol. 32, 2012, pages 72 - 73, ISSN: 0911-4491 * |
| KUNIO KAWANISHI ET AL.: "Fukumaku Saibo Sheet no Kaihatsu to Jutsugo Chokan Yuchaku Model ni Okeru Yuchaku Boshi Koka no Kento", JOURNAL OF THE JAPANESE SOCIETY FOR REGENERATIVE MEDICINE, vol. 11, 2012, pages 199, ISSN: 1347-7919 * |
| NAMBA J. ET AL.: "Modulation of peritendinous adhesion formation by alginate solution in a rabbit flexor tendon model.", JOURNAL OF BIOMEDICAL MATERIALS RESEARCH. PART B, APPLIED BIOMATERIALS ., vol. 80, no. 1, 2007, pages 273 - 279, ISSN: 1552-4973 * |
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US11464597B2 (en) | 2016-07-13 | 2022-10-11 | The University Of Tokyo | Adhesion-preventing composition |
| US11890145B2 (en) | 2016-07-13 | 2024-02-06 | The University Of Tokyo | Adhesion-preventing composition |
| CN110520165A (en) * | 2017-03-02 | 2019-11-29 | 持田制药株式会社 | Fibrocartilage tissue injury in treating composition |
| US11969437B2 (en) | 2017-03-07 | 2024-04-30 | Mochida Pharmaceutical Co., Ltd. | Alginate liquid preparation |
| TWI837085B (en) | 2017-03-07 | 2024-04-01 | 日商持田製藥股份有限公司 | Alginic acid liquid preparation |
| JPWO2018164128A1 (en) * | 2017-03-07 | 2020-04-16 | 持田製薬株式会社 | Alginic acid solution |
| WO2018164128A1 (en) * | 2017-03-07 | 2018-09-13 | 持田製薬株式会社 | Alginate liquid formulation |
| JPWO2019093505A1 (en) * | 2017-11-13 | 2020-11-19 | 持田製薬株式会社 | Adhesion prevention composition |
| JP7267204B2 (en) | 2017-11-13 | 2023-05-01 | 持田製薬株式会社 | Anti-adhesion composition |
| WO2019093505A1 (en) * | 2017-11-13 | 2019-05-16 | 持田製薬株式会社 | Composition for preventing adhesion |
| JPWO2019138583A1 (en) * | 2018-01-15 | 2021-01-28 | 持田製薬株式会社 | Adhesion prevention composition |
| JP7161496B2 (en) | 2018-01-15 | 2022-10-26 | 持田製薬株式会社 | Anti-adhesion composition |
| CN111936175A (en) * | 2018-01-15 | 2020-11-13 | 持田制药株式会社 | Anti-adhesion composition |
| WO2019138583A1 (en) * | 2018-01-15 | 2019-07-18 | 持田製薬株式会社 | Anti-adhesion composition |
| WO2023282247A1 (en) * | 2021-07-05 | 2023-01-12 | 国立大学法人滋賀医科大学 | Tissue formation agent |
Also Published As
| Publication number | Publication date |
|---|---|
| JPWO2016114355A1 (en) | 2017-04-27 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| WO2016114355A1 (en) | Composition for preventing adhesion | |
| Zhao et al. | Injectable dry cryogels with excellent blood-sucking expansion and blood clotting to cease hemorrhage for lethal deep-wounds, coagulopathy and tissue regeneration | |
| US6869938B1 (en) | Compositions of polyacids and polyethers and methods for their use in reducing adhesions | |
| JP5721386B2 (en) | Surgical composition | |
| JP5746617B2 (en) | Injectable hydrogel forming a chitosan mixture | |
| CA2583149C (en) | Biomaterials consisting of sulphated hyaluronic acid and gellan to be used in the prevention of spinal adhesions | |
| JP6907254B2 (en) | Intervertebral disc therapeutic composition | |
| Songkroh et al. | Injectable in situ forming chitosan-based hydrogels for curcumin delivery | |
| JP2017538751A (en) | Fluid hemostatic composition | |
| CA2366880C (en) | Compositions of polyacids and polyethers and methods for their use in reducing adhesions | |
| TW200824726A (en) | Rapidly acting dry sealant and methods for use and manufacture | |
| KR20100063744A (en) | Delayed self-gelling alginate systems and uses thereof | |
| US20220017646A1 (en) | Methods and compositions for achieving hemostasis and stable blood clot formation | |
| Somekawa et al. | Effect of thermoresponsive poly (L-lactic acid)–poly (ethylene glycol) gel injection on left ventricular remodeling in a rat myocardial infarction model | |
| TW201737925A (en) | Pharmaceutical composition useful for adhesion prevention or hemostasis | |
| Zhao et al. | Biomimetic, highly elastic conductive and hemostatic gelatin/rGO-based nanocomposite cryogel to improve 3D myogenic differentiation and guide in vivo skeletal muscle regeneration | |
| AU2015399459A1 (en) | Highly efficacious hemostatic adhesive polymer scaffold | |
| Cai et al. | Physically cross-linked hyaluronan-based ultrasoft cryogel prepared by freeze–thaw technique as a barrier for prevention of postoperative adhesions | |
| CN110740761A (en) | Liquid composition containing alginic acid or pharmaceutically acceptable salt thereof and colloidal polysaccharides | |
| Chen et al. | Adhesive submucosal injection material based on the nonanal group-modified poly (vinyl alcohol)/α-cyclodextrin inclusion complex for endoscopic submucosal dissection | |
| KR102580370B1 (en) | Adhesion barrier containing natural-polymer with biocompatibility and DNA fragment mixture, and process for producing the same | |
| WO2023157150A1 (en) | Use of polysaccharide derivative | |
| Salmasi et al. | Polysaccharide-based (kappa carrageenan/carboxymethyl chitosan) nanofibrous membrane loaded with antifibrinolytic drug for rapid hemostasis-in vitro and in vivo evaluation | |
| JPWO2018012605A1 (en) | Adhesion preventing composition | |
| WO2022034889A1 (en) | Polysaccharide derivative, polysaccharide derivative-drug conjugate, and method for producing same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| 121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 16737426 Country of ref document: EP Kind code of ref document: A1 |
|
| ENP | Entry into the national phase |
Ref document number: 2016569507 Country of ref document: JP Kind code of ref document: A |
|
| NENP | Non-entry into the national phase |
Ref country code: DE |
|
| 122 | Ep: pct application non-entry in european phase |
Ref document number: 16737426 Country of ref document: EP Kind code of ref document: A1 |