KR101437471B1 - A novel lipid-substance-P conjugate - Google Patents
A novel lipid-substance-P conjugate Download PDFInfo
- Publication number
- KR101437471B1 KR101437471B1 KR1020110089415A KR20110089415A KR101437471B1 KR 101437471 B1 KR101437471 B1 KR 101437471B1 KR 1020110089415 A KR1020110089415 A KR 1020110089415A KR 20110089415 A KR20110089415 A KR 20110089415A KR 101437471 B1 KR101437471 B1 KR 101437471B1
- Authority
- KR
- South Korea
- Prior art keywords
- lipid
- substance
- conjugate
- disease
- present
- Prior art date
Links
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 12
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 claims description 12
- 201000010099 disease Diseases 0.000 claims description 10
- 239000008194 pharmaceutical composition Substances 0.000 claims description 10
- 150000002632 lipids Chemical class 0.000 claims description 7
- 208000032382 Ischaemic stroke Diseases 0.000 claims description 4
- 125000000637 arginyl group Chemical group N[C@@H](CCCNC(N)=N)C(=O)* 0.000 claims description 4
- 206010008118 cerebral infarction Diseases 0.000 claims description 4
- 208000026106 cerebrovascular disease Diseases 0.000 claims description 4
- IPCSVZSSVZVIGE-UHFFFAOYSA-M hexadecanoate Chemical compound CCCCCCCCCCCCCCCC([O-])=O IPCSVZSSVZVIGE-UHFFFAOYSA-M 0.000 claims description 4
- 208000027418 Wounds and injury Diseases 0.000 claims description 3
- 230000006378 damage Effects 0.000 claims description 3
- 208000014674 injury Diseases 0.000 claims description 3
- 208000015122 neurodegenerative disease Diseases 0.000 claims description 3
- 208000020431 spinal cord injury Diseases 0.000 claims description 3
- 206010002383 Angina Pectoris Diseases 0.000 claims description 2
- 208000024172 Cardiovascular disease Diseases 0.000 claims description 2
- 208000022306 Cerebral injury Diseases 0.000 claims description 2
- 206010011878 Deafness Diseases 0.000 claims description 2
- 206010012438 Dermatitis atopic Diseases 0.000 claims description 2
- 206010040047 Sepsis Diseases 0.000 claims description 2
- 206010003246 arthritis Diseases 0.000 claims description 2
- 208000006673 asthma Diseases 0.000 claims description 2
- 201000008937 atopic dermatitis Diseases 0.000 claims description 2
- 208000013677 cerebrovascular dementia Diseases 0.000 claims description 2
- 230000003412 degenerative effect Effects 0.000 claims description 2
- 206010012601 diabetes mellitus Diseases 0.000 claims description 2
- 208000035475 disorder Diseases 0.000 claims description 2
- 230000010370 hearing loss Effects 0.000 claims description 2
- 231100000888 hearing loss Toxicity 0.000 claims description 2
- 208000016354 hearing loss disease Diseases 0.000 claims description 2
- 208000010125 myocardial infarction Diseases 0.000 claims description 2
- 208000033808 peripheral neuropathy Diseases 0.000 claims description 2
- 201000010041 presbyopia Diseases 0.000 claims description 2
- 230000000241 respiratory effect Effects 0.000 claims description 2
- 208000023504 respiratory system disease Diseases 0.000 claims description 2
- 230000008733 trauma Effects 0.000 claims description 2
- 206010008129 cerebral palsy Diseases 0.000 claims 1
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 230000004770 neurodegeneration Effects 0.000 claims 1
- 210000000278 spinal cord Anatomy 0.000 claims 1
- 238000001727 in vivo Methods 0.000 abstract description 10
- 238000000338 in vitro Methods 0.000 abstract description 5
- 239000000203 mixture Substances 0.000 description 13
- QDZOEBFLNHCSSF-PFFBOGFISA-N (2S)-2-[[(2R)-2-[[(2S)-1-[(2S)-6-amino-2-[[(2S)-1-[(2R)-2-amino-5-carbamimidamidopentanoyl]pyrrolidine-2-carbonyl]amino]hexanoyl]pyrrolidine-2-carbonyl]amino]-3-(1H-indol-3-yl)propanoyl]amino]-N-[(2R)-1-[[(2S)-1-[[(2R)-1-[[(2S)-1-[[(2S)-1-amino-4-methyl-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]amino]-3-(1H-indol-3-yl)-1-oxopropan-2-yl]amino]-1-oxo-3-phenylpropan-2-yl]amino]-3-(1H-indol-3-yl)-1-oxopropan-2-yl]pentanediamide Chemical compound C([C@@H](C(=O)N[C@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(N)=O)NC(=O)[C@@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CCCCN)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](N)CCCNC(N)=N)C1=CC=CC=C1 QDZOEBFLNHCSSF-PFFBOGFISA-N 0.000 description 12
- 102400000096 Substance P Human genes 0.000 description 12
- 101800003906 Substance P Proteins 0.000 description 12
- 238000000034 method Methods 0.000 description 11
- 108090000765 processed proteins & peptides Proteins 0.000 description 11
- 150000001413 amino acids Chemical class 0.000 description 9
- 210000002901 mesenchymal stem cell Anatomy 0.000 description 9
- 239000011324 bead Substances 0.000 description 8
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 description 7
- 235000001014 amino acid Nutrition 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 7
- 102000004196 processed proteins & peptides Human genes 0.000 description 7
- 210000001185 bone marrow Anatomy 0.000 description 6
- 230000004663 cell proliferation Effects 0.000 description 6
- 230000000694 effects Effects 0.000 description 5
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 5
- 210000000987 immune system Anatomy 0.000 description 5
- 238000004949 mass spectrometry Methods 0.000 description 5
- CMWYAOXYQATXSI-UHFFFAOYSA-N n,n-dimethylformamide;piperidine Chemical compound CN(C)C=O.C1CCNCC1 CMWYAOXYQATXSI-UHFFFAOYSA-N 0.000 description 5
- 235000018102 proteins Nutrition 0.000 description 5
- 102000004169 proteins and genes Human genes 0.000 description 5
- 108090000623 proteins and genes Proteins 0.000 description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
- 235000021314 Palmitic acid Nutrition 0.000 description 4
- 230000037396 body weight Effects 0.000 description 4
- 239000003814 drug Substances 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 230000035876 healing Effects 0.000 description 4
- 238000004128 high performance liquid chromatography Methods 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 230000001404 mediated effect Effects 0.000 description 4
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 230000035755 proliferation Effects 0.000 description 4
- 230000008929 regeneration Effects 0.000 description 4
- 238000011069 regeneration method Methods 0.000 description 4
- 239000011347 resin Substances 0.000 description 4
- 229920005989 resin Polymers 0.000 description 4
- 210000003491 skin Anatomy 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 239000002904 solvent Substances 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- YEDUAINPPJYDJZ-UHFFFAOYSA-N 2-hydroxybenzothiazole Chemical compound C1=CC=C2SC(O)=NC2=C1 YEDUAINPPJYDJZ-UHFFFAOYSA-N 0.000 description 3
- 241000282414 Homo sapiens Species 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 230000008878 coupling Effects 0.000 description 3
- 238000010168 coupling process Methods 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- 230000006870 function Effects 0.000 description 3
- 125000000524 functional group Chemical group 0.000 description 3
- NPZTUJOABDZTLV-UHFFFAOYSA-N hydroxybenzotriazole Substances O=C1C=CC=C2NNN=C12 NPZTUJOABDZTLV-UHFFFAOYSA-N 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- CWWARWOPSKGELM-SARDKLJWSA-N methyl (2s)-2-[[(2s)-2-[[2-[[(2s)-2-[[(2s)-2-[[(2s)-5-amino-2-[[(2s)-5-amino-2-[[(2s)-1-[(2s)-6-amino-2-[[(2s)-1-[(2s)-2-amino-5-(diaminomethylideneamino)pentanoyl]pyrrolidine-2-carbonyl]amino]hexanoyl]pyrrolidine-2-carbonyl]amino]-5-oxopentanoyl]amino]-5 Chemical compound C([C@@H](C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(=O)OC)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CCCCN)NC(=O)[C@H]1N(CCC1)C(=O)[C@@H](N)CCCN=C(N)N)C1=CC=CC=C1 CWWARWOPSKGELM-SARDKLJWSA-N 0.000 description 3
- 230000001737 promoting effect Effects 0.000 description 3
- 210000001519 tissue Anatomy 0.000 description 3
- 239000004475 Arginine Substances 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 102000015735 Beta-catenin Human genes 0.000 description 2
- 108060000903 Beta-catenin Proteins 0.000 description 2
- 208000028006 Corneal injury Diseases 0.000 description 2
- 229920001917 Ficoll Polymers 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 102000003797 Neuropeptides Human genes 0.000 description 2
- 108090000189 Neuropeptides Proteins 0.000 description 2
- 241000700159 Rattus Species 0.000 description 2
- 102100037346 Substance-P receptor Human genes 0.000 description 2
- 206010052428 Wound Diseases 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 150000001408 amides Chemical class 0.000 description 2
- 239000003963 antioxidant agent Substances 0.000 description 2
- 235000006708 antioxidants Nutrition 0.000 description 2
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 2
- 235000009697 arginine Nutrition 0.000 description 2
- 230000036765 blood level Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 238000000354 decomposition reaction Methods 0.000 description 2
- 238000010511 deprotection reaction Methods 0.000 description 2
- 235000005911 diet Nutrition 0.000 description 2
- 230000037213 diet Effects 0.000 description 2
- 230000003511 endothelial effect Effects 0.000 description 2
- 210000002919 epithelial cell Anatomy 0.000 description 2
- 239000003889 eye drop Substances 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 230000012010 growth Effects 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 238000001990 intravenous administration Methods 0.000 description 2
- 238000001840 matrix-assisted laser desorption--ionisation time-of-flight mass spectrometry Methods 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 210000005087 mononuclear cell Anatomy 0.000 description 2
- VMGAPWLDMVPYIA-HIDZBRGKSA-N n'-amino-n-iminomethanimidamide Chemical compound N\N=C\N=N VMGAPWLDMVPYIA-HIDZBRGKSA-N 0.000 description 2
- 210000000653 nervous system Anatomy 0.000 description 2
- 230000001537 neural effect Effects 0.000 description 2
- 239000002858 neurotransmitter agent Substances 0.000 description 2
- -1 organic acid salts Chemical class 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 238000001228 spectrum Methods 0.000 description 2
- 210000000130 stem cell Anatomy 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 239000003981 vehicle Substances 0.000 description 2
- 230000029663 wound healing Effects 0.000 description 2
- JDDWRLPTKIOUOF-UHFFFAOYSA-N 9h-fluoren-9-ylmethyl n-[[4-[2-[bis(4-methylphenyl)methylamino]-2-oxoethoxy]phenyl]-(2,4-dimethoxyphenyl)methyl]carbamate Chemical compound COC1=CC(OC)=CC=C1C(C=1C=CC(OCC(=O)NC(C=2C=CC(C)=CC=2)C=2C=CC(C)=CC=2)=CC=1)NC(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21 JDDWRLPTKIOUOF-UHFFFAOYSA-N 0.000 description 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-M Acetate Chemical compound CC([O-])=O QTBSBXVTEAMEQO-UHFFFAOYSA-M 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- 206010011985 Decubitus ulcer Diseases 0.000 description 1
- 206010012289 Dementia Diseases 0.000 description 1
- 229920001353 Dextrin Polymers 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 206010056340 Diabetic ulcer Diseases 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- LVGKNOAMLMIIKO-UHFFFAOYSA-N Elaidinsaeure-aethylester Natural products CCCCCCCCC=CCCCCCCCC(=O)OCC LVGKNOAMLMIIKO-UHFFFAOYSA-N 0.000 description 1
- 102000016359 Fibronectins Human genes 0.000 description 1
- 108010067306 Fibronectins Proteins 0.000 description 1
- 208000005577 Gastroenteritis Diseases 0.000 description 1
- 108010010803 Gelatin Proteins 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
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101000600903 Homo sapiens Substance-P receptor Proteins 0.000 description 1
- 108060003951 Immunoglobulin Proteins 0.000 description 1
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 1
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 1
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 102000003729 Neprilysin Human genes 0.000 description 1
- 108090000028 Neprilysin Proteins 0.000 description 1
- 108010040718 Neurokinin-1 Receptors Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 235000019483 Peanut oil Nutrition 0.000 description 1
- 208000004210 Pressure Ulcer Diseases 0.000 description 1
- 102000007562 Serum Albumin Human genes 0.000 description 1
- 108010071390 Serum Albumin Proteins 0.000 description 1
- 206010072170 Skin wound Diseases 0.000 description 1
- 208000020339 Spinal injury Diseases 0.000 description 1
- 208000007107 Stomach Ulcer Diseases 0.000 description 1
- 102000040945 Transcription factor Human genes 0.000 description 1
- 108091023040 Transcription factor Proteins 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 239000008186 active pharmaceutical agent Substances 0.000 description 1
- 239000000443 aerosol Substances 0.000 description 1
- 239000000556 agonist Substances 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000005557 antagonist Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000010323 ascorbic acid Nutrition 0.000 description 1
- 229960005070 ascorbic acid Drugs 0.000 description 1
- 239000011668 ascorbic acid Substances 0.000 description 1
- 235000003704 aspartic acid Nutrition 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004271 bone marrow stromal cell Anatomy 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 230000009702 cancer cell proliferation Effects 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000003093 cationic surfactant Substances 0.000 description 1
- 230000012292 cell migration Effects 0.000 description 1
- 230000005754 cellular signaling Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000002738 chelating agent Substances 0.000 description 1
- 238000003889 chemical engineering Methods 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000005757 colony formation Effects 0.000 description 1
- 229940124301 concurrent medication Drugs 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 230000001268 conjugating effect Effects 0.000 description 1
- 230000021615 conjugation Effects 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 210000004087 cornea Anatomy 0.000 description 1
- 235000012343 cottonseed oil Nutrition 0.000 description 1
- 239000002385 cottonseed oil Substances 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 235000019425 dextrin Nutrition 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 210000002889 endothelial cell Anatomy 0.000 description 1
- LVGKNOAMLMIIKO-QXMHVHEDSA-N ethyl oleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OCC LVGKNOAMLMIIKO-QXMHVHEDSA-N 0.000 description 1
- 229940093471 ethyl oleate Drugs 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 229940012356 eye drops Drugs 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- 201000005917 gastric ulcer Diseases 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- 230000013595 glycosylation Effects 0.000 description 1
- 238000006206 glycosylation reaction Methods 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 230000003394 haemopoietic effect Effects 0.000 description 1
- 230000013632 homeostatic process Effects 0.000 description 1
- AKPUJVVHYUHGKY-UHFFFAOYSA-N hydron;propan-2-ol;chloride Chemical compound Cl.CC(C)O AKPUJVVHYUHGKY-UHFFFAOYSA-N 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
- 102000018358 immunoglobulin Human genes 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- 210000002510 keratinocyte Anatomy 0.000 description 1
- 125000003473 lipid group Chemical group 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 210000002540 macrophage Anatomy 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 210000000663 muscle cell Anatomy 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 206010028537 myelofibrosis Diseases 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 210000002569 neuron Anatomy 0.000 description 1
- 210000000440 neutrophil Anatomy 0.000 description 1
- 239000002736 nonionic surfactant Substances 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 239000000312 peanut oil Substances 0.000 description 1
- 230000006320 pegylation Effects 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 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
- 229920001983 poloxamer Polymers 0.000 description 1
- 229920000724 poly(L-arginine) polymer Polymers 0.000 description 1
- 108010011110 polyarginine Proteins 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 102000005962 receptors Human genes 0.000 description 1
- 108020003175 receptors Proteins 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 150000004671 saturated fatty acids Chemical class 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000008159 sesame oil Substances 0.000 description 1
- 235000011803 sesame oil Nutrition 0.000 description 1
- 230000008054 signal transmission Effects 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- ADNPLDHMAVUMIW-CUZNLEPHSA-N substance P Chemical compound C([C@@H](C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(N)=O)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CCCCN)NC(=O)[C@H]1N(CCC1)C(=O)[C@@H](N)CCCN=C(N)N)C1=CC=CC=C1 ADNPLDHMAVUMIW-CUZNLEPHSA-N 0.000 description 1
- 150000005846 sugar alcohols Chemical class 0.000 description 1
- 230000003319 supportive effect Effects 0.000 description 1
- 239000000829 suppository Substances 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 239000003826 tablet Substances 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
- 230000024883 vasodilation Effects 0.000 description 1
- 235000015112 vegetable and seed oil Nutrition 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/08—Peptides having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
- A61K47/543—Lipids, e.g. triglycerides; Polyamines, e.g. spermine or spermidine
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
Abstract
본 발명은 신규한 지질-서브스턴스-P 접합체를 제공한다. 본 발명의 지질-서브스턴스-P 접합체는 시험관내 또는 생체내에서 향상된 안정성을 가지며, 특히, 증가된 생체내 반감기를 나타낸다. The present invention provides a novel lipid-substance-P conjugate. The lipid-substance-P conjugates of the present invention have improved stability in vitro or in vivo, and in particular, exhibit increased in vivo half-life.
Description
본 발명은 신규한 지질-서브스턴스-P 접합체에 관한 것이다. The present invention relates to a novel lipid-substance-P conjugate.
서브스턴스-P (Substance-P; 또한 ‘SP’라 함)는 면역, 신경계를 조절하는 신경 펩타이드로써, 종래에는 신경 전달 물질로만 알려져 왔지만 최근 들어서는 면역계와 신경계를 연결해주는 역할을 하며 상처치유과정에 중요한 역할을 하는 것으로 밝혀졌다. SP는 골수에서 분비될 수 있는 뉴로펩타이드로 다른 성장인자들처럼 특정 수용체 (receptor)에 결합하여 신호를 전달하고, 골수와 림프구의 기능을 조절한다. Substance-P (also called 'SP') is a neuropeptide that regulates the immune system and the nervous system. It has been known as a neurotransmitter in the past but recently it has been linked to the immune system and the nervous system. It has been found to play an important role. SP is a neuropeptide that can be secreted in the bone marrow. Like other growth factors, it binds to a specific receptor and regulates the function of bone marrow and lymphocytes.
SP는 11개의 아미노산으로 구성된 펩타이드로 사람, 쥐, 토끼에서 동일한 아미노산 서열을 가지고 있어 종간 구별이 없는 인자이다. 뉴로키닌 리셉터 (neurokinin receptor)-1을 통해 시그널을 전달하고 이 리셉터는 각막 (Watanabe M et al Jpn J Ophthalmol. 46, 616-20, 2002), 피부 (Leu J Y et al. Br J Dermatol. 155, 657-62 2006), 면역세포 등에서 발현된다. SP의 기능은 통증 유도, 혈관 확장 등이 있다. 하지만 SP는 신경 세포뿐만 아니라 비신경 조직에서도 발현되는데, 상피세포, 내피세포 (Watanabe M et al Jpn J Ophthalmol. 46, 616-20, 2002), 대식세포, 호중구 (Ho W. Z. J Immunol. 159, 5654-60 1997), 암세포 (Singh D et al. PNAS 97, 388-393, 2000) 등에서 발현이 확인되었다. 이러한 결과들을 통해 SP가 기존에 알려진 기능인, 신경 전달 물질로서의 역할 외에도 신경-면역계 조절, 골수 섬유증, 암세포 증식, 상처치유 등에서도 중요한 역할을 할 수 있음을 알 수 있다. SP is a peptide composed of 11 amino acids and has the same amino acid sequence in human, rat, and rabbit, and is a species-free factor. The neurokinin receptor-1 signal is transmitted through the cornea (Watanabe M et al Jpn J Ophthalmol. 46, 616-20, 2002), skin (Leu JY et al., Br J Dermatol. , 657-62 2006), and are expressed in immune cells and the like. The functions of SP include pain induction and vasodilation. However, SP is expressed not only in neurons but also in non-neuronal tissues, but epithelial cells and endothelial cells (Watanabe M et al Jpn J Ophthalmol. 46, 616-20, 2002), macrophages, neutrophils (Ho WZ J Immunol. -60 1997), and cancer cells (Singh D et al. PNAS 97, 388-393, 2000). These results indicate that SP plays an important role in neuronal immune system regulation, myelofibrosis, cancer cell proliferation and wound healing as well as a known function of neurotransmitter.
최근 SP는 상처 치유에 이용되고 있으며, 그 중 하나로, 각막 손상 시 상피세포의 재생을 촉진시키는 안약으로의 개발이 진행되어 왔으며 각막손상 동물 모델에서 SP에 의한 치유 효과가 보고되었다. (Nakamura M. et al. Diabetologia. 46,839-42, 2003), Yamada N et al. Invest Ophthalmol Vis Sci.45, 1125-31, 2004) 현재 안약으로서의 개발이 일본에서 진행 중이다. Recently, SP has been used for wound healing. One of them has been developed as an eyedrop for promoting the regeneration of epithelial cells in corneal injury, and the healing effect by SP in corneal injured animal models has been reported. (Nakamura M. et al., Diabetologia, 46, 839-42, 2003), Yamada N et al. Invest Ophthalmol Vis Sci. 45, 1125-31, 2004) Development of eye drops is currently underway in Japan.
또한, SP는 피부 상처에도 적용되고 있는데, 인트라사이트 겔과 SP를 혼합하여 상처에 적용하였을 때 SP를 처리한 그룹에서 치유가 빨라지는 것이 보고되었고 (Rook JM. Biochemical pharmacology. 74, 752 757, 2007), 피부 상처 동물 모델에서 SP를 직접 상처 주변 조직에 주사시 치유 효과가 있음이 확인되었다(Delgado AV.Exp Biol Med. 230, 271-270, 2005). In addition, SP has also been applied to skin wounds, and it has been reported that when applied to wounds by mixing intrasite gel and SP, the healing is accelerated in the group treated with SP (Rook JM. Biochemical pharmacology. 74, 752 757, 2007 ), It was confirmed that SP directly injected into the surrounding tissues of wounded skin had a healing effect (Delgado AV. Exp Biol Med. 230, 271-270, 2005).
또한, 중배엽 줄기세포에 의한 조혈세포의 지지 작용이 SP에 의해 촉진됨을 보고되었다 (Pranela Rameshwar. et al. Journal of Neuroimmunology 121, 22-31, 2001). 상기 논문에서는 시험관내(In vitro) 내에서 조혈세포와 중배엽 줄기세포를 공동 배양시 SP의 처리에 의한 조혈 세포의 콜로니 형성 증가가 보고되었다. 즉, SP가 골수의 조혈 작용을 조절하는 골수 기질의 세포와 줄기세포에서 일어나는 일련의 반응들을 개시함을 보고하였다. 이는 생체내(in vivo)에서 골수의 재생에 SP의 역할의 중요성을 강조한다(Pranela Rameshwar. et al. Journal of Neuroimmunology 121, 22-31, 2001).In addition, supportive actions of hematopoietic cells by mesenchymal stem cells have been reported to be promoted by SP (Pranela Rameshwar et al., Journal of Neuroimmunology 121, 22-31, 2001). In this article, it has been reported that the colony formation of hematopoietic cells by SP treatment when co-culturing hematopoietic cells and mesenchymal stem cells in vitro has been reported. In other words, it has been reported that SP initiates a series of reactions in bone marrow stromal cells and stem cells that regulate the hematopoietic action of the bone marrow. This emphasizes the importance of the role of SP in bone marrow regeneration in vivo (Pranela Rameshwar, et al. Journal of Neuroimmunology 121, 22-31, 2001).
한편, 골수 재생 및 유지에서 SP의 역할이 보고된 바 있다. 실제로 중배엽 줄기세포를 시험관내(in vitro)에서 배양하며 SP를 처리한 결과 세포의 증식이 빨라진다는 결과가 확인되었으며, 이런 결과들은 중배엽 줄기세포뿐만이 아니라 피부의 섬유 아세포나 각질 세포, 근육 세포에서도 증명되었다. 또한, 이와 관련한 전반적인 신호 전달 체계도 많은 연구가 진행되어있다. 이러한 SP의 증식 촉진 효과가 분자생물학적 수준으로 많은 세포에서 연구되었다(Chuen Mao Yang et. al. Cellular signalling. 14, 913-923, 2002).On the other hand, the role of SP in bone marrow regeneration and maintenance has been reported. In fact, mesenchymal stem cells were cultured in vitro and treated with SP, indicating that cell proliferation was accelerated. These results were confirmed not only in mesenchymal stem cells but also in fibroblasts, keratinocytes, and muscle cells of the skin . In addition, the overall signal transmission system related to this is undergoing much research. These SP promoting effects on proliferation have been studied in many cells at the molecular level (Chuen Mao Yang et al., Cellular signaling 14, 913-923, 2002).
또한, SP에 의한 중배엽 줄기세포 증식 촉진 기작을 밝히기 위한 연구 결과로써, SP를 중배엽 줄기세포에 처리하면 세포 증식에 관련된 전사 인자인 베타 카테닌 (Beta catenin)이 세포질에서 핵으로 이동하는 것이 밝혀졌다. 베타 카테닌이 관여하는 Wnt 시그널은 중배엽 줄기세포와 조혈 모세포의 증식에 중요한 역할을 하는 것으로 보고되었다(Reya T Nature 423, 409-414, 2003, De Boer J, Tissue eng. 10, 393-401, 2004). 중배엽 줄기세포의 증식이 빨라지면 중배엽 줄기세포에 의해 지지되고 있는 조혈 세포의 회복도 빨라지게 되어 면역계의 재생 또한 앞당겨지게 된다. 이렇듯, SP는 상처 부분으로 세포의 이동을 촉진하여 치유를 촉진할 뿐 아니라 골수 환경을 유지함으로써 전체적인 면역계의 항상성 유지에도 관여하는 것으로 알려져 있다. In addition, as a result of studying SP promoting mesenchymal stem cell proliferation, it has been found that when SP is treated in mesenchymal stem cells, beta catenin, a transcription factor involved in cell proliferation, migrates from the cytoplasm to the nucleus. The beta-catenin-mediated Wnt signal has been reported to play an important role in the proliferation of mesenchymal stem cells and hematopoietic stem cells (Reya T Nature 423, 409-414, 2003, De Boer J, Tissue eng. 10, 393-401, 2004 ). When the proliferation of mesenchymal stem cells is accelerated, the recovery of the hematopoietic cells supported by mesenchymal stem cells is accelerated and the regeneration of the immune system is accelerated. Thus, it is known that SP promotes cell migration to the wound site to promote healing as well as to maintain the immune system homeostasis by maintaining the marrow environment.
그러나, SP는 혈청 내에 있는 중성 엔도펩티다제(Neutral endopeptidase)에 의해 분해되기 때문에, 실제로 체내에서 머무르는 시간은 매우 짧다는 한계가 있다. However, since SP is degraded by the neutral endopeptidase in serum, there is a limit in that the time for staying in the body is very short.
이와 관련하여, SP 변이체가 항체의 종양 세포 내 전달을 촉진한다는 보고 (Rizk SS 등, Proc. Natl. Acad. Sci. USA; 106(27);11011-5; 2009 July 7), C-말단에서 변형된 SP의 유사체가 2차 메신저 경로에 종속하여 NK-1 수용체의 효현제로도, 길항제로도 작용할 수 있다는 보고는 있었으나, 생체내 안정성이 증가된 SP 변이체에 관하여는 현재까지 보고된 바 없었다. In this regard, it has been reported that the SP variant promotes the delivery of the antibody into tumor cells (Rizk SS et al., Proc. Natl. Acad. Sci. USA; 106 (27) It has been reported that modified SP analogues can act as agonists and antagonists of the NK-1 receptor depending on the secondary messenger pathway, but no SP mutants with increased in vivo stability have been reported to date.
이와 관련하여, 단백질의 안정성을 증가시키기 위한 기술로써, PEG화 (Pegylation), 시알화 (sialation), 당화 (glycosylation), 타 단백질과의 접합(conjugation), 점 돌연변이 (point mutation) 등과 같은 기술이 알려져 있으나 현재까지 SP에 대하여는 보고된 바 없었다.In this regard, techniques such as pegylation, sialation, glycosylation, conjugation with other proteins, point mutation, and the like have been used as techniques for increasing protein stability However, no SP has been reported to date.
한편, 단백질에 팔미트산의 접합이 세포 표면의 항체에 부착을 촉진한다는 보고(Stanley A. Kim 등, J. Immunol. Methods; 158(1):57-65; 1993 Jan 14)가 있었으나, 이는 단백질의 안정성 향상과는 무관한 것이었다. On the other hand, there has been a report that the binding of palmitic acid to proteins promotes adhesion to antibody on the surface of cells (Stanley A. Kim et al., J. Immunol. Methods; 158 (1): 57-65; It was not related to the improvement of protein stability.
본 발명은 안정성이 향상된 신규한 지질-서브스턴스-P 접합체를 제공하고자 한다. The present invention seeks to provide a novel lipid-substance-P conjugate with improved stability.
본 발명자들은 서브스턴스-P 의 분해속도를 전임상실험에서 관찰한 결과, 서브스턴스-P를 정맥에 주사한 결과 주사 후 1시간이 지나면 혈역 내 수치가 정성수준으로 떨어지는 것을 관찰하고, 서브스턴스-P의 시험관내 또는 생체내 안정성, 보다 구체적으로는 생체내 반감기를 증가시키기 위한 다양한 변형방법을 모색한 결과, 본 발명을 완성하였다. As a result of observing the decomposition rate of Substance-P in the pre-clinical experiment, the present inventors observed that the intravenous blood level decreased to the qualitative level after 1 hour from the injection of Substance-P, In vitro or in vivo stability, more specifically, various modifications for increasing the half-life in vivo.
이에, 본 발명은 안정성이 증가된 신규한 지질-서브스턴스-P 접합체를 제공한다. Thus, the present invention provides a novel lipid-substrance-P conjugate with increased stability.
본 발명은 서열번호 1의 아미노산 서열 상의 하나 이상의 위치에 지질이 접합된 지질-서브스턴스-P 접합체를 제공한다. The present invention provides a lipid-substratum-P conjugate that is lipid conjugated at one or more positions on the amino acid sequence of SEQ ID NO: 1.
본 발명의 바람직한 구현예로써, 본 발명은 서열번호 1의 1번 위치의 아르기닌 잔기에 지질이 접합된 지질-서브스턴스-P 접합체를 제공한다. As a preferred embodiment of the present invention, the present invention provides a lipid-substrute-P conjugate lipid conjugated to an arginine residue at
본 발명에서 사용되는 지질은 C12 내지 C16의 지방산이 바람직하며, 가장 바람직하게는 팔미테이트이다. The lipid used in the present invention is preferably C12 to C16 fatty acid, and most preferably palmitate.
본 발명의 가장 바람직한 구현예로써, 본 발명은 서브스턴스-P의 N-말단의 아르기닌 잔기에 팔미트산이 접합된 형태로, C79H127N17O15S 의 화학식을 갖고, 1586.06 의 분자질량을 가지며, 구체적으로는 하기 식의 구조를 가지는 지질-서브스턴스-P 접합체를 제공한다. As a most preferred embodiment of the present invention, the present invention relates to a composition comprising a palmitic acid conjugated to an arginine residue at the N-terminus of Substance-P and having the formula C 79 H 127 N 17 O 15 S, Specifically, a lipid-substance-P conjugate having a structure of the following formula.
[식 1][Formula 1]
본 발명의 지질-서브스턴스-P 접합체는 당 분야에 알려진 통상의 화학적, 유전공학적 변형 방법을 이용하여 제조될 수 있으며, 서브스턴스-P의 아미노산 서열 중 적절한 잔기, 바람직하게는 N-말단의 아르기닌 서열의 아민그룹에 지질, 바람직하게는 C12 내지 C16 지방산, 가장 바람직하게는 팔미테이트(또한 ‘팔미트산’이라고 함)를 접합하여 제조할 수 있다. The Lipid-Substance-P conjugates of the present invention can be prepared using conventional chemical and genetic engineering methods known in the art and can be prepared by adding appropriate residues of Substance-P, preferably N-terminal arginine May be prepared by conjugating a lipid, preferably a C12 to C16 fatty acid, and most preferably palmitate (also referred to as palmitic acid) to the amine group of the sequence.
본 발명에서, 서브스턴스-P는 서열번호1의 아미노산 서열을 갖는 펩티드로써, 시판되는 것을 이용하거나, 당 분야에 알려진 방법으로 화학적, 또는 유전공학적으로 합성할 수 있다. In the present invention, Substance-P is a peptide having the amino acid sequence of SEQ ID NO: 1, which may be commercially available or synthesized chemically or genetically by methods known in the art.
본 발명에서, 지질은 당 분야에서 일반적으로 사용되는 것으로 시판되는 것을 사용할 수 있으며, 본 발명의 가장 바람직한 구현 형태인 팔미테이트는 CH3(CH2)14COOH 의 화학식을 가지는 탄소 16의 포화 직쇄 지방산으로, 시판되는 것을 이용하거나, 당 분야에 알려진 방법으로 화학적으로 합성할 수 있다. In the present invention, lipids can be those commercially available as those commonly used in the art, and the most preferred embodiment of the present invention, palmitate, is a saturated fatty acid of carbon 16 having the formula CH 3 (CH 2 ) 14 COOH , Commercially available or chemically synthesized by methods known in the art.
상기 지질-서브스턴스-P 접합체의 제조는 당 분야의 통상의 지식을 가진 자가 용이하게 실시하는 기술수준의 범위에 해당하며, 구체적인 제조방법은 본원 실시예를 참조할 수 있다. The preparation of the lipid-substance-P conjugate corresponds to a range of skill levels that a person skilled in the art can easily carry out, and specific manufacturing methods can be referred to the embodiments of the present invention.
본 발명은 또한 지질-서브스턴스-P 접합체를 포함하는 약학적 조성물을 제공한다. The present invention also provides a pharmaceutical composition comprising a lipid-substrance-P conjugate.
본 발명은 또한 상기 식 1의 지질-서브스턴스-P 접합체를 포함하는 약학적 조성물을 제공한다. The present invention also provides a pharmaceutical composition comprising the lipid-substrance-P conjugate of
본 발명의 약학적 조성물은 서브스턴스-P에 의해 매개되는 질환이나 상태의 개선, 예방 또는 치료를 위하여 사용될 수 있다. The pharmaceutical composition of the present invention can be used for improving, preventing or treating diseases or conditions mediated by Substance-P.
서브스턴스-P에 의해 매개되는 질환이나 상태의 예로는 패혈증 (sepsis), 관절염 (arthritis), 천식(asthma), 호흡기질환, 호흡기세포융합바이러스 (respiratory syncitial virus) 질환, 방사선 손상, 위궤양, 위장염, 당뇨성 궤양, 욕창, 신경척추 손상(spinal injury), 치매(dementia), 각막손상, 피부 외상, 아토피성피부염, 심혈관질환, 심근경색, 협심증, 허혈성 뇌졸중, 뇌혈관성 치매, 뇌경색, 뇌손상 후유증, 척수손상, 척수신경 후유증, 퇴행성 질환, 뇌경색 후유증, 말초신경 장애, 노안, 퇴행성 난청, 뇌수술 후유증, 디스크 손상 또는 당뇨병 등이 있다. 이러한 서브스턴스-P에 의해 매개되는 질환이나 상태는 당 분야에 널리 알려져 있다. Examples of diseases or conditions mediated by Substance-P include sepsis, arthritis, asthma, respiratory disease, respiratory syncitial virus disease, radiation damage, gastric ulcer, gastroenteritis, Corneal injury, skin trauma, atopic dermatitis, cardiovascular disease, myocardial infarction, angina pectoris, ischemic stroke, cerebrovascular dementia, cerebral infarction, cerebral injury aftereffects, diabetic ulcer, pressure sores, nerve spinal injury, dementia, Spinal cord injury, spinal cord injury, degenerative diseases, cerebral infarction, peripheral neuropathy, presbyopia, degenerative hearing loss, sequelae of brain surgery, disc damage or diabetes. Diseases and conditions mediated by such Substance-P are well known in the art.
본 발명의 약학적 조성물은, 각각 통상의 방법에 따라 산제, 과립제, 정제, 캡슐제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구형 제형, 외용제, 좌제 또는 멸균 주사용액 등의 적절한 형태로 제형화하여 사용될 수 있다.The pharmaceutical composition of the present invention may be formulated into appropriate forms such as powders, granules, tablets, capsules, suspensions, emulsions, oral formulations such as syrups and aerosols, external preparations, suppositories, Can be used.
본 발명의 약학적 조성물은 랫트, 마우스, 가축, 인간 등의 포유동물에 다양한 경로로, 예를 들면, 경구, 설하, 직장, 피부, 피하, 근육, 복강, 정맥, 동맥 등으로 투여될 수 있다. 상기 약학적 조성물은 목적하는 순도를 갖는 지질-서브스턴스-P 접합체를 약학적으로 허용되는 담체, 부형제 또는 안정화제와 혼합함으로써 저장 또는 투여용으로 제조될 수 있다. 이러한 물질은 사용되는 투여량 및 농도에서 복용자에게 무독성인 것으로, 인산염, 시트르산염, 아세트산염 및 다른 유기산염과 같은 완충제 아스코르브산과 같은 산화방지제 폴리아르기닌, 단백질, 예를 들면 혈청 알부민, 젤라틴 또는 면역글로불린과 같은 저분자량 (약 10개 미만의 잔기를 갖는) 펩티드 폴리비닐피롤리디논과 같은 친수성 중합체 글리신, 글루탐산, 아스파르트산 또는 아르기닌과 같은 아미노산 모노사카라이드, 디사카라이드, 및 셀룰로오스 또는 그의 유도체를 포함하는 다른 탄수화물, 글루코스, 만노스 또는 덱스트린 EDTA와 같은 킬레이트제 만니톨 또는 소르비톨과 같은 당 알콜 나트륨과 같은 카운터 이온 및(또는) 트윈(Tween), 플루로닉스(Pluronics) 또는 폴리에틸렌글리콜과 같은 비이온성 계면활성제를 포함한다.The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock and humans in various routes such as oral, sublingual, rectal, subcutaneous, intramuscular, intraperitoneal, intravenous, . The pharmaceutical composition may be prepared for storage or administration by mixing a lipid-Substance-P conjugate having the desired purity with a pharmaceutically acceptable carrier, excipient or stabilizer. Such materials are non-toxic to the recipient at the dosages and concentrations employed and include antioxidants polyarginine such as buffer ascorbic acid, such as phosphate, citrate, acetate and other organic acid salts, proteins such as serum albumin, gelatin or immunoglobulin (Having less than about 10 residues) peptide polyvinylpyrrolidinone, such as glycine, glutamic acid, aspartic acid or arginine, amino acid monosaccharides, disaccharides, and cellulose or derivatives thereof Other cationic surfactants such as chelating agents mannitol such as glucose, mannose or dextrin EDTA or sodium hydroxide of sugar alcohol such as sorbitol and / or non-ionic surfactants such as Tween, Pluronics or polyethylene glycols, .
본 발명의 약학적 조성물은 주사용 멸균 조성물의 형태로 당업계의 통상적 방법에 따라 제조될 수 있다. 상기 주사용 멸균 조성물은 비히클, 예를 들면, 물 또는 참깨유, 땅콩유 또는 면실유와 같은 천연 식물성 오일 또는 에틸 올레이트와 같은 합성 지방 비히클 중의 활성 화합물의 용액 또는 현택액을 포함할 수 있다. The pharmaceutical compositions of the present invention can be prepared according to conventional methods in the art in the form of injectable sterile compositions. The injectable sterile composition may contain a solution or suspension of the active compound in a vehicle, for example a synthetic vegetable oil such as water or sesame oil, peanut oil or cottonseed oil, or a synthetic fatty vehicle such as ethyl oleate.
상기 주사용 멸균 조성물에는 또한 완충제, 방부제, 산화 방지제 등이 당업계에 통상적으로 사용되는 모습에 따라 혼입될 수 있다.The injectable sterilized composition for injection may also contain a buffer, preservative, antioxidant or the like according to a state conventionally used in the art.
본 발명에서 사용되는 지질-서브스턴스-P 접합체의 "치료적으로 유효한 양"은 연구자, 수의사, 의사 또는 기타 임상의에 의해 생각되는 동물 또는 인간에서 생물학적 또는 의학적 반응을 유도하는 활성 성분 또는 약학적 조성물의 양을 의미하는 것으로, 이는 치료되는 질환 또는 장애의 증상 완화를 유도하는 양을 포함한다. 본 발명의 유효 성분에 대한 치료상 유효 투여량 및 투여횟수는 원하는 효과에 따라 변화될 것임은 당업자에게 자명하다.A "therapeutically effective amount " of the lipid-substance-P conjugate used in the present invention refers to an amount of active ingredient or pharmaceutical agent that induces a biological or medical response in an animal or human considered by a researcher, veterinarian, Refers to the amount of the composition, which includes an amount that induces symptomatic relief of the disease or disorder being treated. It will be apparent to those skilled in the art that the therapeutically effective dose and the number of administrations of the active ingredient of the present invention will vary depending on the desired effect.
그러므로, 투여될 최적의 투여량은 당업자에 의해 쉽게 결정될 수 있으며, 질환의 종류, 질환의 중증도, 조성물에 함유된 유효 성분 및 다른 성분의 함량, 제형의 종류, 및 환자의 연령, 체중, 일반 건강 상태, 성별 및 식이, 투여 시간, 투여 경로 및 조성물의 분비율, 치료기간, 동시 사용되는 약물을 비롯한 다양한 인자에 따라 조절될 수 있다. 일례로, 성인의 경우, 본 발명의 서브스턴스-P 변이체를 1일 1회 투여시, 0.000005 - 500 mg/체중kg, 바람직하게는 0.00005-5 mg/체중 kg 의 용량으로 투여하는 것이 바람직하다. Thus, the optimal dosage to be administered can be readily determined by those skilled in the art and will vary with the nature of the disease, the severity of the disease, the amount of active and other ingredients contained in the composition, the type of formulation, and the age, The age, body weight, sex, diet, time of administration, route of administration and fraction of the composition, duration of treatment, concurrent medication, and the like. For example, in the case of an adult, administering the Substance-P variant of the present invention once per day is preferably administered in a dose of 0.000005-500 mg / kg body weight, preferably 0.00005-5 mg / kg body weight.
본 발명의 약학적 조성물의 투여용량은 상기 설명된 질병들을 치료하는 방법에 관련하여 의사에 의해 약물의 효과를 변형시킬 만한 요인들, 예를 들어 나이, 상태, 체중, 성별, 식이, 어떤 감염에 의한 중증도, 투여시점 등의 기타 임상학적인 조건에 따른 요인들에 따라서 결정되어야 한다.The dosage of the pharmaceutical composition of the present invention may be determined by a physician insofar as it relates to a method of treating the above-described diseases, by factors that may modify the effectiveness of the drug, such as age, condition, weight, sex, The severity of the disease, the time of administration, and other clinical factors.
본 발명의 약학적 조성물은 단회 또는 계속적 투여가 가능하다. 다만, 초회 투여 후 일정한 순환계 농도를 유지시키기 위해 계속적인 투여를 하는 것이 바람직하다. 효과적인 투여용량과 방법은 당업계에 공지된 일반적 기술들에 기초하여 최적화될 것이며, 반복투여 횟수는 각 환자의 약동학적인 인자들과 투여경로에 따라 달라질 것이다. 상기 투여용량, 용법 및 투여횟수는 또한 투여된 약물의 물리적인 상태, 안정성, 생체내 방출속도, 생체내 제거율 등에 따라 최적화될 수 있다. 또한, 투여경로에 따른 적절한 투여용량이 체중, 체표면적, 장기의 크기에 따라 계산될 수 있다. 적절한 용량은 적절한 용량-반응 자료와 함께 혈중농도의 자료와 함께 판단하여 확정되어야 한다. 최종의 투여법은 의사에 의해 결정될 것이며, 의사는 약물의 비활성, 환자의 중증도, 약물에 대한 환자의 반응정도, 나이, 컨디션, 체중, 성별, 그리고 식이와 기타 다른 임상학적인 요인들을 고려하여 처방해야 한다. The pharmaceutical composition of the present invention can be administered once or continuously. However, it is preferable to continue administration to maintain a constant circulating concentration after the first administration. Effective dosing dosages and methods will be optimized based on the general techniques known in the art, and the number of repeated dosing will depend on the pharmacokinetic parameters of each patient and the route of administration. The dosage, usage, and frequency of administration may also be optimized according to the physical state, stability, rate of in vivo release, rate of in vivo clearance, etc. of the administered drug. In addition, the appropriate dosage depending on the route of administration can be calculated according to body weight, body surface area, and size of organs. Appropriate doses should be determined with appropriate dose-response data along with data on blood levels. The final dosage regimen will be decided by the physician and the physician should prescribe taking into account the inactivity of the drug, the severity of the patient, the patient's response to the drug, age, condition, weight, sex, diet and other clinical factors. do.
본 발명의 신규한 지질-서브스턴스-P 접합체는 서브스턴스-P 본래의 활성은 유지하거나, 그와 동등한 효과를 나타내는 동시에, 시험관내 및 생체내에서 향상된 안정성을 가지며, 특히, 생체내에서 증가된 반감기를 가진다. The novel Lipid-Substance-P conjugate of the present invention maintains the original activity of Substance-P or exhibits its equivalent effect, while having improved stability in vitro and in vivo, in particular, increased in vivo It has a half-life.
도 1은 서브스턴스-P의 질량분석 결과를 나타내는 스펙트럼이다.
도 2는 팔미테이트-서브스턴스-P 접합체의 질량분석 결과를 나타내는 스펙트럼이다.
도 3은 팔미테이트-서브스턴스-P 접합체에 의한 EPC 증식 향상 효과를 확인한 그래프이다. Fig. 1 is a spectrum showing mass spectrometry results of Substance-P. Fig.
Fig. 2 is a spectrum showing the result of mass spectrometry of the palmitate-substrons-P conjugate.
FIG. 3 is a graph showing the effect of improving the EPC proliferation by the palmitate-Substance-P conjugate.
본 발명의 이점 및 특징, 그리고 그것들을 달성하는 방법은 후술하는 실시예들을 참조하면 명확해질 것이다. 그러나 본 발명은 이하에서 개시되는 실시예들에 한정되지 않는다. Advantages and features of the present invention and methods of achieving them will become apparent with reference to the embodiments described hereinafter. However, the present invention is not limited to the embodiments disclosed below.
[제조예][Manufacturing Example]
<제조예 1> 서브스턴스-P의 제조Production Example 1 Production of Substance-P
서브스턴스 P는 Fmoc 화학 시스템으로써 서열번호1의 11개 아미노산을 Fmoc이 결합되어 있는 링크 아미드 MBHA 레진 (Rink Amide MBHA Resin)을 이용하여 합성하였다. Substance P was synthesized by Fmoc chemical system using 11 amino acids of SEQ ID NO: 1 using Fmoc-linked Rink Amide MBHA Resin.
구체적으로, Fmoc이 결합되어 있는 링크 아미드 MBHA 레진을 반응기에 넣고 DMF 상에서 스웰링(swelling)하였다. Specifically, the link amide MBHA resin to which Fmoc was bonded was put into a reactor and swelled on DMF.
그 후, 용매(DCM, DMF)를 이용하여 레진을 세척하였다. 20% 피페리딘/DMF 이용하여 비드에 NH2를 보호하고 있는 FMOC을 제거하였다 (5분/10분). 용매를 (DCM,DMF)를 이용하여 비드 상에 잔존하는 20% 피페리딘/DMF 를 완전히 제거하였다. 아미노산을 DMF로 녹인 후, 2MHOBT/DMF와 2MDIC/DMF를 첨가하여 활성화시킨 후, 반응기에 투여하여 3 시간동안 커플링을 실시하였다. 상기 공정을 진행하여 11개의 아미노산을 반복 커플링을 실시하였다. 합성이 끝난 비드는 절개 용액을 첨가하여 비드 상의 관능기 탈보호(Functional group deprotection)를 4 시간동안 실시하였다. 관능기 탈보호가 끝난 혼합물을 에틸 에테르를 이용한 추출을 통해 생성물을 수득하였다. 수득한 생성물을 건조하여 1mg를 칭량하여 0.1%TFA/ACN 200㎕, 0.1%TFA/H2O 400㎕ 에 넣고 녹인 후 SIMADZU사의 HPLC를 이용하여 화합물 분석을 실시하였다.The resin was then washed with a solvent (DCM, DMF). The FMOC protecting the NH 2 from the beads was removed using 20% piperidine / DMF (5 min / 10 min). The solvent (DCM, DMF) was used to completely remove the remaining 20% piperidine / DMF on the beads. The amino acid was dissolved in DMF and activated by addition of 2M HOBT / DMF and 2MDIC / DMF. The mixture was then added to the reactor and coupled for 3 hours. Repeated coupling of 11 amino acids was carried out through the above process. The synthesized beads were subjected to functional group deprotection for 4 hours by adding incision solution. The mixture was extracted with ethyl ether to obtain a product. The obtained product was dried, weighed 1 mg, and dissolved in 400 μl of 0.1% TFA / ACN, 200 μl of 0.1% TFA / H2O and analyzed by SIMADZU HPLC.
원하는 피크의 물질을 이동상에서 받아내어 bruker사의 Maldi-Tof를 이용하여 질량 분석을 실시하였다. 원하는 물질을 선별하여 SIMADZU사의 HPLC를 이용하여 화합물 정제를 실시하여 99.99%의 고순도 펩티드를 생산하였다. The desired peak material was taken from the mobile phase and mass spectrometry was performed using Maldi-Tof from bruker. The desired substances were selected and the compounds were purified using SIMADZU HPLC to produce 99.99% high purity peptides.
액체화되어 있는 펩티드는 (-80℃) 동결건조기를 이용하여 3 일 동안 동결건조를 실시하였다. 건조된 펩티드를 0.1N Hcl/H2O을 이용하여 10mg당 1ml 기준으로 혼합하고 초음파분쇄기와 교반장치를 이용하여 3 시간동안 염석한 후 (-80℃) 동결건조기를 이용하여 3 일 동안 다시 동결건조하였다. 건조된 펩티드를 전자저울을 이용하여 일정량의 생성물을 얻었다. 합성된 생성물을 질량 분석기(mass spectrometry)로 확인하였다 (도 1).The liquefied peptides were lyophilized for 3 days using a (-80 ° C) freeze dryer. The dried peptides were mixed with 0.1 N HCl / H 2 O per 1 ml per 10 mg, salted for 3 hours using an ultrasonic mill and a stirrer, and then frozen (-80 ° C) for 3 days using a freeze dryer And dried. An amount of the product was obtained by using an electronic balance of the dried peptide. The synthesized product was identified by mass spectrometry (Fig. 1).
<제조예 2> 팔미테이트-서브스턴스-P 접합체의 제조PREPARATION EXAMPLE 2 Preparation of Palmitate-Substance-P conjugate
Fmoc이 결합되어 있는 링크 아미드 MBHA 레진을 반응기에 넣고 DMF 상에서 스웰링하였다. Fmoc conjugated link amide MBHA resin was added to the reactor and swelled on DMF.
용매(DCM, DMF)를 이용하여 RESIN 세척하였다. 20% 피페리딘/DMF 을 이용하여 비드에 NH2 를 보호하고 있는 FMOC을 제거하였다. (5분/10분) 용매(DCM, DMF)를 이용하여 비드 상에 잔존하는 20% 피페리딘/DMF 를 완전히 제거하였다. 아미노산을 DMF로 녹인 후, 2MHOBT/DMF와 2MDIC/DMF를 첨가하여 활성화시킨 후, 반응기에 투여하여 3 시간동안 커플링을 실시하였다. 상기 공정을 진행하여 11개의 아미노산의 반복 커플링을 실시하였다. 합성이 끝난 SP의 Fmoc을 20% 피페리딘/DMF을 이용하여(5분/10분) 제거한 후, 팔미트산에 2MHOBT/DMF와 2MDIC/DMF를 첨가하여 활성화시킨 후 SP 비드와 같이 반응기에 투여하여 3 시간동안 커플링을 실시하였다. 합성이 끝난 비드는 절개용액을 첨가하여 비드 상의 관능기 탈보호를 4시간 동안 실시하였다. 관능기 탈보호가 끝난 혼합물은 에틸 에테르를 이용한 추출을 통해 생성물을 수득하였다.Resin was washed with solvent (DCM, DMF). FMOC protecting NH 2 from beads was removed using 20% piperidine / DMF. The remaining 20% piperidine / DMF on the beads was completely removed using a solvent (DCM, DMF) (5 min / 10 min). The amino acid was dissolved in DMF and activated by addition of 2M HOBT / DMF and 2MDIC / DMF. The mixture was then added to the reactor and coupled for 3 hours. Repeated coupling of 11 amino acids was carried out by proceeding with the above process. After the Fmoc of the synthesized SP was removed by using 20% piperidine / DMF (5 min / 10 min), 2M HOBT / DMF and 2MDIC / DMF were added to the palmitic acid to activate the SP bead. And the coupling was performed for 3 hours. The synthesized beads were subjected to the deprotection of the functional groups on the beads by adding the incision solution for 4 hours. After the functional group deprotected mixture, the product was obtained through extraction with ethyl ether.
수득한 생성물을 건조하여 1mg을 칭량하여0.1% TFA/ACN 500㎕, 0.1% TFA/H2O 100㎕를 넣고 녹인 후 SIMADZU사의 HPLC를 이용하여 화합물 분석을 실시하였다.The obtained product was dried, weighed 1 mg, and dissolved in 500 μl of 0.1% TFA / ACN and 100 μl of 0.1% TFA / H2O, and analyzed by SIMADZU HPLC.
원하는 피크의 물질을 이동상에서 받아내어 bruker사의 Maldi-Tof를 이용하여 질량 분석을 실시하였다. 원하는 물질을 선별하여 SIMADZU사의 HPLC를 이용하여 화합물 정제를 실시하여 99.99%의 고순도 펩티드를 생산하였다.The desired peak material was taken from the mobile phase and mass spectrometry was performed using Maldi-Tof from bruker. The desired substances were selected and the compounds were purified using SIMADZU HPLC to produce 99.99% high purity peptides.
액체화되어 있는 펩티드는 (-80℃) 동결건조기를 이용하여 3 일동안 동결건조를 실시하였다. 건조된 펩티드는 0.1N Hcl/H2O을 이용하여 10mg당 1ml 기준으로 혼합하여 초음파분쇄기와 교반장치를 이용하여 3 시간 동안 염석한 후 (-80℃) 동결건조기를 이용하여 3 일동안 다시 동결건조하였다. 건조된 펩티드는 전자저울을 이용하여 일정량의 생성물을 얻었다. 합성된 생성물을 질량 분석기로 확인하였다 (도 2).The liquefied peptides were lyophilized for 3 days using a (-80 ° C) freeze dryer. The dried peptides were mixed in a ratio of 1 ml per 10 mg using 0.1 N HCl / H 2 O, salted for 3 hours using an ultrasonic grinder and a stirrer, and then frozen (-80 ° C.) for 3 days using a freeze dryer And dried. The dried peptides were obtained by using electronic scales to obtain a certain amount of product. The synthesized product was identified by a mass spectrometer (Fig. 2).
[실시예][Example]
<실시예 1> 팔미테이트-서브스턴스-P 접합체의 세포 증식 효과 확인Example 1 Confirmation of Cell Proliferation Effect of Palmitate-Substance-P Conjugate
(1) 혈관내피전구세포(Endothelail progenitor cell; EPC) 배양 (1) culture of endothela lil progenitor cells (EPC)
골수천자액 (bone marrow aspirates)을 PBS로 2배 희석한 후 미리 튜브에 담아둔 Ficoll 위에 조심스럽게 얹었다(Ficoll: 말초혈액= 1:1 v/v). 그 후 2200rpm에서 25분간 원심분리하고, 최상층인 혈장층 제거하고 단핵세포층 만을 분리하여 새로운 튜브로 옮겨 담았다. 얻어진 단핵세포층에 PBS 30ml을 넣고 1500rpm, 5분간 원심분리하였다(세척 단계). EGM-2 (Endothelial growth medium-2, Lonza) 배지로 현탁한 세포를 피브로넥틴이 코팅된 100mm 디쉬 (Falcon)에 접종하고, EGM-2 (Endothelial growth medium-2, Lonza)에서 2주일간 배양하였다(5% CO2, 37도).Bone marrow aspirates were diluted 2-fold with PBS and carefully placed on Ficoll (Ficoll: peripheral blood = 1: 1 v / v) pre-incubated in tubes. Thereafter, centrifugation was carried out at 2200 rpm for 25 minutes, the plasma layer as the uppermost layer was removed, and only the mononuclear cell layer was separated and transferred to a new tube. 30 ml of PBS was added to the obtained mononuclear cell layer and centrifuged at 1500 rpm for 5 minutes (washing step). Cells suspended in EGM-2 (Endothelial growth medium-2, Lonza) medium were inoculated into fibronectin coated 100 mm dish (Falcon) and cultured in EGM-2 (Endothelial growth medium-2, Lonza) for 2 weeks % CO 2, 37 degree).
(2) 세포증식률 확인(2) Confirm cell proliferation rate
상기 배양한 PEC 를 96-웰 플레이트 (Corning)에 2x104 세포/웰의 농도로 각각 SP, 팔미테이트-SP와 함께 배양하였다. 48 시간 동안 배양한 후 MTT 용액을 3시간 동안 처리하였다. 포르마잔(Formazan) 형성을 확인 한 후 웰내 배양액을 모두 흡입 제거하였다. 바닥에 붙어있는 포르마잔을 0.04N HCl-이소프로파놀로 현탁한 뒤 ELISA 리더기 (Biorad) 570nm 파장에 O.D 값을 측정하였다. 그 결과를 도 3에 나타내었다. The cultured PECs were cultured in 96-well plates (Corning) at a concentration of 2 × 10 4 cells / well together with SP, palmitate-SP. After 48 hours of incubation, the MTT solution was treated for 3 hours. After formazan formation was confirmed, all of the cells in the wells were inhaled and removed. The formazan attached to the bottom was suspended in 0.04 N HCl-isopropanol and the OD value was measured at 570 nm wavelength in an ELISA reader (Biorad). The results are shown in Fig.
도 3에서 확인되는 바와 같이, 팔미테이트-SP 접합체를 처리한 그룹의 경우 서브스턴스-P 만을 처리한 그룹에 비하여 세포 증식이 증가하였으며, 이는 팔미테이트-SP 접합체가 서브스턴스-P에 비하여 분해속도가 느려졌기 때문인 것으로 보인다. As shown in Fig. 3, in the group treated with the palmitate-SP conjugate, cell proliferation was increased as compared with the group treated only with Substance-P, indicating that the palmitate-SP conjugate had a decomposition rate It seems to be because of the slowing down.
<110> CELL & BIO CO., LTD.
<120> A NOVEL LIPID-SUBSTANCE-P CONJUGATE
<130> P11-076-CNB
<160> 1
<170> KopatentIn 1.71
<210> 1
<211> 11
<212> PRT
<213> Homo sapiens
<400> 1
Arg Pro Lys Pro Gln Gln Phe Phe Gly Leu Met
1 5 10
<110> CELL & BIO CO., LTD.
<120> A NOVEL LIPID-SUBSTANCE-P CONJUGATE
<130> P11-076-CNB
<160> 1
<170> Kopatentin 1.71
<210> 1
<211> 11
<212> PRT
<213> Homo sapiens
<400> 1
Arg Pro Lys Pro Gln Gln Phe Phe
Claims (6)
[식]
[expression]
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020110089415A KR101437471B1 (en) | 2011-09-05 | 2011-09-05 | A novel lipid-substance-P conjugate |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020110089415A KR101437471B1 (en) | 2011-09-05 | 2011-09-05 | A novel lipid-substance-P conjugate |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20130026059A KR20130026059A (en) | 2013-03-13 |
| KR101437471B1 true KR101437471B1 (en) | 2014-09-03 |
Family
ID=48177461
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1020110089415A KR101437471B1 (en) | 2011-09-05 | 2011-09-05 | A novel lipid-substance-P conjugate |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR101437471B1 (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100326938B1 (en) | 1999-06-22 | 2002-03-13 | 박호군 | Compound prepared by mixing synthetic peptide conjugated with fatty acid and phospholipid, and preparation process thereof |
| KR20110018273A (en) * | 2009-08-17 | 2011-02-23 | 경희대학교 산학협력단 | A composition for preventing or treating inflammation |
-
2011
- 2011-09-05 KR KR1020110089415A patent/KR101437471B1/en active IP Right Grant
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100326938B1 (en) | 1999-06-22 | 2002-03-13 | 박호군 | Compound prepared by mixing synthetic peptide conjugated with fatty acid and phospholipid, and preparation process thereof |
| KR20110018273A (en) * | 2009-08-17 | 2011-02-23 | 경희대학교 산학협력단 | A composition for preventing or treating inflammation |
Also Published As
| Publication number | Publication date |
|---|---|
| KR20130026059A (en) | 2013-03-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP3248584B2 (en) | Fibronectin adhesion inhibitor | |
| EP2409988B1 (en) | Peptide fragments for inducing synthesis of extracellular matrix proteins | |
| JP3445269B2 (en) | Treatment of retinal neuronal disorders by application of insulin-like growth factors and analogs | |
| JPH05506030A (en) | How to predispose mammals to promote tissue repair | |
| EP2552470B1 (en) | Peptides for promoting angiogenesis and an use thereof | |
| CN104159911A (en) | Peptidomimetic compounds as immunomodulators | |
| JPH06510305A (en) | Treatment of diseases by application of insulin-like growth factors and analogs | |
| KR20100014596A (en) | Template-fixed peptidomimetics | |
| KR20010015711A (en) | POLYPEPTIDE, cDNA ENCODING THE POLYPEPTIDE, AND USE OF THE BOTH | |
| KR20240025721A (en) | Methods and compositions for treating age-associated conditions | |
| CN102470156A (en) | Polypeptides selective for av ss3 integrin conjugated with a variant of human serum albumin (HSA) and pharmaceutical uses thereof | |
| US9352024B2 (en) | Uses of interleukin-22(IL-22) in treating and preventing nerve damage diseases or neurodegenerative diseases | |
| EP2588491A2 (en) | Novel peptide and use thereof | |
| US9480727B2 (en) | Synthetic peptide for inhibiting expression of type 2 TNF receptor and use thereof | |
| WO2009131191A1 (en) | Metastin derivative and use thereof | |
| EA006423B1 (en) | Oligopeptide with the biological activity of a thrombopoietin receptor modulator and method for using thereof | |
| KR20180050755A (en) | CYCLIC POLYPEPTIDE, PROCESS FOR PRODUCING THE SAME AND THERAPEUTIC USE | |
| EP2566494B1 (en) | Cxcr4 receptor compounds | |
| WO2021200259A1 (en) | Vipr2 antagonist peptide | |
| KR101437471B1 (en) | A novel lipid-substance-P conjugate | |
| WO2001064742A1 (en) | Modified bdnf | |
| CN115028681A (en) | Chimeric compound for degrading cyclophilin A and preparation method and application thereof | |
| RU2317097C2 (en) | Plant-originated heterocarpine protein having anti-cancer properties | |
| WO2011119008A2 (en) | Peptides for promoting angiogenesis and an use thereof | |
| WO1995015765A1 (en) | Peptides to overcome inhibition of nerve growth |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A201 | Request for examination | ||
| E902 | Notification of reason for refusal | ||
| E701 | Decision to grant or registration of patent right | ||
| GRNT | Written decision to grant | ||
| FPAY | Annual fee payment |
Payment date: 20170816 Year of fee payment: 4 |
|
| FPAY | Annual fee payment |
Payment date: 20180827 Year of fee payment: 5 |
|
| FPAY | Annual fee payment |
Payment date: 20190828 Year of fee payment: 6 |