WO2017022981A9 - 합성 펩타이드를 이용한 유도만능줄기세포의 제조방법 - Google Patents
합성 펩타이드를 이용한 유도만능줄기세포의 제조방법 Download PDFInfo
- Publication number
- WO2017022981A9 WO2017022981A9 PCT/KR2016/007857 KR2016007857W WO2017022981A9 WO 2017022981 A9 WO2017022981 A9 WO 2017022981A9 KR 2016007857 W KR2016007857 W KR 2016007857W WO 2017022981 A9 WO2017022981 A9 WO 2017022981A9
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- cells
- peptide
- pluripotent stem
- stem cells
- induced pluripotent
- Prior art date
Links
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 74
- 210000004263 induced pluripotent stem cell Anatomy 0.000 title claims abstract description 25
- 238000004519 manufacturing process Methods 0.000 title abstract description 11
- 210000004027 cell Anatomy 0.000 claims abstract description 62
- 230000001737 promoting effect Effects 0.000 claims abstract description 11
- 108010057466 NF-kappa B Proteins 0.000 claims abstract description 10
- 102000003945 NF-kappa B Human genes 0.000 claims abstract description 10
- 230000007704 transition Effects 0.000 claims abstract description 9
- 210000000130 stem cell Anatomy 0.000 claims abstract description 8
- 230000000694 effects Effects 0.000 claims abstract description 6
- 230000004069 differentiation Effects 0.000 claims description 40
- 238000000034 method Methods 0.000 claims description 31
- 235000018417 cysteine Nutrition 0.000 claims description 20
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 claims description 20
- 229930182817 methionine Natural products 0.000 claims description 20
- 230000032459 dedifferentiation Effects 0.000 claims description 19
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 claims description 18
- 210000001082 somatic cell Anatomy 0.000 claims description 17
- 239000000411 inducer Substances 0.000 claims description 13
- 150000001413 amino acids Chemical group 0.000 claims description 11
- 230000006698 induction Effects 0.000 claims description 10
- 108010031318 Vitronectin Proteins 0.000 claims description 8
- 102100035140 Vitronectin Human genes 0.000 claims description 8
- 239000000203 mixture Substances 0.000 claims description 7
- 101710135898 Myc proto-oncogene protein Proteins 0.000 claims description 6
- 102100038895 Myc proto-oncogene protein Human genes 0.000 claims description 6
- 101100247004 Rattus norvegicus Qsox1 gene Proteins 0.000 claims description 6
- 101710150448 Transcriptional regulator Myc Proteins 0.000 claims description 6
- 239000012620 biological material Substances 0.000 claims description 6
- 108700021430 Kruppel-Like Factor 4 Proteins 0.000 claims description 5
- 229920005615 natural polymer Polymers 0.000 claims description 5
- 101150086694 SLC22A3 gene Proteins 0.000 claims description 4
- 239000004480 active ingredient Substances 0.000 claims description 4
- 101150111214 lin-28 gene Proteins 0.000 claims description 4
- 229920001059 synthetic polymer Polymers 0.000 claims description 4
- 241000282412 Homo Species 0.000 claims description 3
- SQDAZGGFXASXDW-UHFFFAOYSA-N 5-bromo-2-(trifluoromethoxy)pyridine Chemical compound FC(F)(F)OC1=CC=C(Br)C=N1 SQDAZGGFXASXDW-UHFFFAOYSA-N 0.000 claims description 2
- 229920001287 Chondroitin sulfate Polymers 0.000 claims description 2
- 108010035532 Collagen Proteins 0.000 claims description 2
- 102000008186 Collagen Human genes 0.000 claims description 2
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 claims description 2
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 claims description 2
- 108010067306 Fibronectins Proteins 0.000 claims description 2
- 102000016359 Fibronectins Human genes 0.000 claims description 2
- 108010010803 Gelatin Proteins 0.000 claims description 2
- RVGRUAULSDPKGF-UHFFFAOYSA-N Poloxamer Chemical compound C1CO1.CC1CO1 RVGRUAULSDPKGF-UHFFFAOYSA-N 0.000 claims description 2
- 235000010443 alginic acid Nutrition 0.000 claims description 2
- 229920000615 alginic acid Polymers 0.000 claims description 2
- 229960001126 alginic acid Drugs 0.000 claims description 2
- 239000000783 alginic acid Substances 0.000 claims description 2
- 150000004781 alginic acids Chemical class 0.000 claims description 2
- 229940059329 chondroitin sulfate Drugs 0.000 claims description 2
- 229920001436 collagen Polymers 0.000 claims description 2
- 229960005188 collagen Drugs 0.000 claims description 2
- 229920000159 gelatin Polymers 0.000 claims description 2
- 239000008273 gelatin Substances 0.000 claims description 2
- 229940014259 gelatin Drugs 0.000 claims description 2
- 235000019322 gelatine Nutrition 0.000 claims description 2
- 235000011852 gelatine desserts Nutrition 0.000 claims description 2
- 229920001983 poloxamer Polymers 0.000 claims description 2
- 229960000502 poloxamer Drugs 0.000 claims description 2
- -1 polyethylene Polymers 0.000 claims description 2
- 229920001451 polypropylene glycol Polymers 0.000 claims description 2
- RMZNXRYIFGTWPF-UHFFFAOYSA-N 2-nitrosoacetic acid Chemical compound OC(=O)CN=O RMZNXRYIFGTWPF-UHFFFAOYSA-N 0.000 claims 1
- 239000004698 Polyethylene Substances 0.000 claims 1
- 239000001963 growth medium Substances 0.000 claims 1
- 229920000573 polyethylene Polymers 0.000 claims 1
- 210000001778 pluripotent stem cell Anatomy 0.000 abstract description 13
- 239000003814 drug Substances 0.000 abstract description 6
- 230000002401 inhibitory effect Effects 0.000 abstract description 5
- 241001465754 Metazoa Species 0.000 abstract description 4
- 238000012258 culturing Methods 0.000 abstract description 2
- 210000002966 serum Anatomy 0.000 abstract description 2
- 229940124597 therapeutic agent Drugs 0.000 abstract 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 17
- 210000002950 fibroblast Anatomy 0.000 description 13
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 12
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 12
- 238000003786 synthesis reaction Methods 0.000 description 10
- 230000002500 effect on skin Effects 0.000 description 9
- 210000001671 embryonic stem cell Anatomy 0.000 description 9
- 101710126211 POU domain, class 5, transcription factor 1 Proteins 0.000 description 8
- 102100035423 POU domain, class 5, transcription factor 1 Human genes 0.000 description 8
- 101800004193 Peptide P3 Chemical group 0.000 description 8
- 238000010186 staining Methods 0.000 description 8
- 230000015572 biosynthetic process Effects 0.000 description 7
- 239000002609 medium Substances 0.000 description 7
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 6
- 102000018651 Epithelial Cell Adhesion Molecule Human genes 0.000 description 6
- 108010066687 Epithelial Cell Adhesion Molecule Proteins 0.000 description 6
- 108010068647 P2 peptide Proteins 0.000 description 6
- 238000010166 immunofluorescence Methods 0.000 description 6
- 239000011347 resin Substances 0.000 description 6
- 229920005989 resin Polymers 0.000 description 6
- 230000007705 epithelial mesenchymal transition Effects 0.000 description 5
- 230000014509 gene expression Effects 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 108090000623 proteins and genes Proteins 0.000 description 5
- 239000007790 solid phase Substances 0.000 description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
- 239000006145 Eagle's minimal essential medium Substances 0.000 description 4
- 101000800116 Homo sapiens Thy-1 membrane glycoprotein Proteins 0.000 description 4
- 102100033523 Thy-1 membrane glycoprotein Human genes 0.000 description 4
- 229940098773 bovine serum albumin Drugs 0.000 description 4
- 238000002659 cell therapy Methods 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 230000018109 developmental process Effects 0.000 description 4
- 238000004520 electroporation Methods 0.000 description 4
- 238000000684 flow cytometry Methods 0.000 description 4
- 238000001943 fluorescence-activated cell sorting Methods 0.000 description 4
- 239000003550 marker Substances 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 241000700605 Viruses Species 0.000 description 3
- 239000000872 buffer Substances 0.000 description 3
- 239000002771 cell marker Substances 0.000 description 3
- 238000012790 confirmation Methods 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
- 238000010511 deprotection reaction Methods 0.000 description 3
- 210000002919 epithelial cell Anatomy 0.000 description 3
- 230000008672 reprogramming Effects 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 108010003545 C1 peptide Proteins 0.000 description 2
- 241000282326 Felis catus Species 0.000 description 2
- 101000655352 Homo sapiens Telomerase reverse transcriptase Proteins 0.000 description 2
- 108090000144 Human Proteins Proteins 0.000 description 2
- 102000003839 Human Proteins Human genes 0.000 description 2
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 description 2
- 229930040373 Paraformaldehyde Natural products 0.000 description 2
- 108091027967 Small hairpin RNA Proteins 0.000 description 2
- 235000001014 amino acid Nutrition 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 210000004899 c-terminal region Anatomy 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000003776 cleavage reaction Methods 0.000 description 2
- 239000011248 coating agent Substances 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 2
- 230000001939 inductive effect Effects 0.000 description 2
- 230000003834 intracellular effect Effects 0.000 description 2
- 125000001360 methionine group Chemical group N[C@@H](CCSC)C(=O)* 0.000 description 2
- 210000000056 organ Anatomy 0.000 description 2
- 229920002866 paraformaldehyde Polymers 0.000 description 2
- 230000007017 scission Effects 0.000 description 2
- 239000004055 small Interfering RNA Substances 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 239000013598 vector Substances 0.000 description 2
- OZFAFGSSMRRTDW-UHFFFAOYSA-N (2,4-dichlorophenyl) benzenesulfonate Chemical compound ClC1=CC(Cl)=CC=C1OS(=O)(=O)C1=CC=CC=C1 OZFAFGSSMRRTDW-UHFFFAOYSA-N 0.000 description 1
- KJCVRFUGPWSIIH-UHFFFAOYSA-N 1-naphthol Chemical compound C1=CC=C2C(O)=CC=CC2=C1 KJCVRFUGPWSIIH-UHFFFAOYSA-N 0.000 description 1
- PRDFBSVERLRRMY-UHFFFAOYSA-N 2'-(4-ethoxyphenyl)-5-(4-methylpiperazin-1-yl)-2,5'-bibenzimidazole Chemical compound C1=CC(OCC)=CC=C1C1=NC2=CC=C(C=3NC4=CC(=CC=C4N=3)N3CCN(C)CC3)C=C2N1 PRDFBSVERLRRMY-UHFFFAOYSA-N 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 239000012591 Dulbecco’s Phosphate Buffered Saline Substances 0.000 description 1
- 101000687905 Homo sapiens Transcription factor SOX-2 Proteins 0.000 description 1
- 206010027476 Metastases Diseases 0.000 description 1
- 101100510267 Mus musculus Klf4 gene Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 108091023040 Transcription factor Proteins 0.000 description 1
- 102000040945 Transcription factor Human genes 0.000 description 1
- 102100024270 Transcription factor SOX-2 Human genes 0.000 description 1
- 150000001408 amides Chemical class 0.000 description 1
- 238000009412 basement excavation Methods 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 239000007975 buffered saline Substances 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 239000006143 cell culture medium Substances 0.000 description 1
- 230000007348 cell dedifferentiation Effects 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000005757 colony formation Effects 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000001973 epigenetic effect Effects 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 239000012212 insulator Substances 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 230000009401 metastasis Effects 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- CMWYAOXYQATXSI-UHFFFAOYSA-N n,n-dimethylformamide;piperidine Chemical compound CN(C)C=O.C1CCNCC1 CMWYAOXYQATXSI-UHFFFAOYSA-N 0.000 description 1
- FEMOMIGRRWSMCU-UHFFFAOYSA-N ninhydrin Chemical compound C1=CC=C2C(=O)C(O)(O)C(=O)C2=C1 FEMOMIGRRWSMCU-UHFFFAOYSA-N 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 230000017111 nuclear migration Effects 0.000 description 1
- 229920002113 octoxynol Polymers 0.000 description 1
- 230000002018 overexpression Effects 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 239000002245 particle 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
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 230000001172 regenerating effect Effects 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000007727 signaling mechanism Effects 0.000 description 1
- 230000000087 stabilizing effect Effects 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 239000013603 viral vector Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0696—Artificially induced pluripotent stem cells, e.g. iPS
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/001—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof by chemical synthesis
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
- C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
- C07K14/4702—Regulators; Modulating activity
- C07K14/4703—Inhibitors; Suppressors
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
- C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
- C07K14/4723—Cationic antimicrobial peptides, e.g. defensins
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0607—Non-embryonic pluripotent stem cells, e.g. MASC
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
- C12N5/0662—Stem cells
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
- C07K14/485—Epidermal growth factor [EGF], i.e. urogastrone
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
- C07K14/50—Fibroblast growth factor [FGF]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/998—Proteins not provided for elsewhere
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2506/00—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells
- C12N2506/13—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells from connective tissue cells, from mesenchymal cells
- C12N2506/1307—Differentiation of animal cells from one lineage to another; Differentiation of pluripotent cells from connective tissue cells, from mesenchymal cells from adult fibroblasts
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2510/00—Genetically modified cells
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/0018—Culture media for cell or tissue culture
- C12N5/0037—Serum-free medium, which may still contain naturally-sourced components
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/0018—Culture media for cell or tissue culture
- C12N5/0043—Medium free of human- or animal-derived components
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/0018—Culture media for cell or tissue culture
- C12N5/005—Protein-free medium
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/0018—Culture media for cell or tissue culture
- C12N5/0056—Xeno-free medium
Definitions
- the present invention relates to a method for producing induced pluripotent stem cells using a synthetic peptide, and more particularly, a peptide that inhibits the activity of NF- ⁇ B and promotes mesenchymal-Epithelial transition (MET). It relates to a method for producing induced pluripotent stem cells.
- a synthetic peptide and more particularly, a peptide that inhibits the activity of NF- ⁇ B and promotes mesenchymal-Epithelial transition (MET). It relates to a method for producing induced pluripotent stem cells.
- MET mesenchymal-Epithelial transition
- Stem cells are undifferentiated cells capable of infinitely self-renewing and differentiating into cells of all tissues of the body. They are used for regenerative medicine, the development of new medicines for cell therapy, the causes and treatment of human diseases. It is an important object of research. Embryonic stem cells can differentiate into various cells and make whole organs, but there are ethical problems and immune rejection reactions that require the use of an egg as a cell therapy.
- somatic cells which have been differentiated in 2006, during in vitro culture, induces differentiation-induced pluripotent stem cells with autologous regeneration and pluripotency, which are characteristic of embryonic stem cells.
- the technology for producing cells was the first in the world to be succeeded by Yamanaka's team at Kyoto University, Japan (Takahashi K et al., Cell 126 (4): 663-676, 2006; Takahashi K et al., Cell 131 (5). ): 861-72, 2007).
- Reverse differentiation refers to a state in which already differentiated cells return to their initial undifferentiated state.
- iPSC refers to cells that have acquired pluripotency similar to embryonic stem cells capable of giving artificially stimulated externally differentiated somatic cells to cells of all organs forming our bodies. .
- dedifferentiation factors are most effective with the use of viruses, but the use of viruses for the production of dedifferentiated pluripotent stem cells for therapeutic purposes has the potential risk, and the randomization of intracellular genomes is very stable.
- problems such as mutations in genes.
- xenopathogens such as fetal bovine serum (FBS) and mouse embryonic fibroblasts (MEF), which are animal-derived feeder cells, are needed in the process of inducing differentiation.
- FBS fetal bovine serum
- MEF mouse embryonic fibroblasts
- the ultimate purpose of dedifferentiated pluripotent stem cells is to produce tissue that can be implanted into the human body, there is a limit to the clinical application due to the above risk.
- NF- ⁇ B promotes Epithelial-Mesenchymal Transition (EMT) (Huber MA et al., J Clin Invest. 114 (4): 569-81, 2004), and mesenchyme.
- EMT Epithelial-Mesenchymal Transition
- MET Mesenchymal-Epithelial Transition
- the present inventors have made efforts to increase the efficiency of induction of dedifferentiation of iPSCs in order to develop cell therapeutics that are safe for clinical application.
- the present inventors have found a functional peptide that inhibits the activity of intracellular NF- ⁇ B protein.
- Excavation and confirmed that the peptide induces mesenchymal-Epithelial transition (MET) to promote differentiation into pluripotent stem cells (iPSC), and completed the present invention.
- MET mesenchymal-Epithelial transition
- An object of the present invention is to provide a composition for promoting reverse differentiation into induced pluripotent stem cells from differentiated cells containing the peptide represented by the amino acid sequence of the general formula (I) as an active ingredient.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- Another object of the invention is the step of (a) introducing a differentiation inducer into the differentiated cells; And (b) treating and incubating a peptide having the amino acid sequence represented by the general formula I in a cell into which the reverse differentiation inducer has been introduced, thereby preparing a reverse differentiation induced pluripotent stem cell from the differentiated cells. .
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- the present invention provides a composition for promoting reverse differentiation into induced pluripotent stem cells from differentiated cells containing the peptide represented by the amino acid sequence of the general formula (I) as an active ingredient.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- the invention also comprises the steps of (a) introducing a differentiation inducer into differentiated cells; And (b) treating and incubating the peptide into which the reverse differentiation inducer is introduced, the peptide represented by the amino acid sequence of the general formula I below.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- Figure 1 is treated with functional peptides (P1, P2, P3) and control peptides (C1, C2) in human dermal fibroblasts introduced into the differentiation factor at 24 hours intervals for 10 days, alkaline force Colony staining was confirmed by Alkaline phosphatase staining (AP).
- P1, P2, P3 functional peptides
- C1, C2 control peptides
- FIG. 2 is a result of confirming Oct4 expression of an embryonic stem cell (ES cell) marker through immunofluorescence (IF) after treatment of the peptide in the same manner as in FIG. 1.
- ES cell embryonic stem cell
- IF immunofluorescence
- FIG. 3 shows the degree of mesenchymal-Epithelial transition (MET), which is a process of dedifferentiating somatic cells differentiated through flow cytometry (FACS) after treating peptides in the same manner as in FIG. 1.
- MET mesenchymal-Epithelial transition
- FACS flow cytometry
- a functional peptide that inhibits the activity of NF- ⁇ B protein is identified, and the peptide inhibits epithelial-mesenchymal transition (EMT) and further mesenchymal-epithelial metastasis (Mesenchymal-Epithelial). Induced transition, MET) to promote dedifferentiation.
- EMT epithelial-mesenchymal transition
- Mesenchymal-Epithelial mesenchymal-Epithelial metastasis
- the present invention relates to a composition for promoting reverse differentiation into induced pluripotent stem cells from differentiated cells containing the peptide represented by the amino acid sequence of the general formula (I) as an active ingredient at a consistent point.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- the present invention relates to a novel use of a peptide represented by the amino acid sequence of the general formula (I) below for promoting reverse differentiation from differentiated cells into induced pluripotent stem cells.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- the present invention provides a method for preparing a cell including: (a) introducing a differentiation inducer into differentiated cells; And (b) treating and incubating a peptide into which a reverse differentiation inducer is introduced, the peptide represented by the amino acid sequence of the general formula (I) below.
- X 1 and X 2 are cysteine (C: cysteine) or methionine (M: methionine).
- the peptide is preferably any one selected from the group consisting of SEQ ID NOs: 1 to 3, but is not limited thereto.
- P1 peptide GKCSTRGRKCCRRKK (SEQ ID NO: 1)
- P3 Cectide GKCSTRGRKMCRRKK (SEQ ID NO: 3)
- the peptide is preferably 0.01 to 100 ⁇ M concentration, more preferably 10 ⁇ M, but is not limited thereto.
- the peptide is preferably treated for 10 days at intervals of 24 hours, but is not limited thereto.
- the differentiated cells are preferably somatic cells or progenitor cells, preferably derived from humans, but are not limited thereto.
- Somatic cell of the present invention refers to human dermal fibroblasts having mesenchymal characteristics.
- EMEM Eagle's Minimum Essential Medium, ATCC 30-2003
- Essential 8 Medium Gibco, A15169-01
- the reverse differentiation inducer is preferably at least one selected from the group consisting of Oct3 / 4, Sox2, c-Myc, Klf4 and Lin28, but is not limited thereto.
- Oct3 / 4, Sox2, Klf4, c-Myc or Lin28 reprogramming genes refer to genes that can reprogram differentiated cells, in particular Oct4, Sox2, Klf4 and c-Myc are Yamanaka factors It is called.
- the dedifferentiation factor is prepared by dividing the three different episomal vectors and expressing p53 shRNA together (pCXLE-hOCT3 / 4-shp53, pCXLE-hSK, pCXLE-hUL). Increased cell establishment efficiency (Okita K et al., Nat Methods. 8 (5): 409-12, 2011).
- a method for delivering a retrodifferentiation factor to differentiated cells may be administered by injecting a retrodifferentiation inducer into a culture of differentiated cells, by direct infusion, or by transfection with a viral vector containing a retrodifferentiation factor. It is preferable to use a method of infecting cells differentiated with viruses obtained from packaging cells, but is not limited thereto. It is preferable to use microinjection, electroporation, insulator, particle bombardment, etc. as a method of directly injecting the dedifferentiation factor into differentiated cells, but is not limited thereto. It is not.
- induction of differentiation factor by electroporation using a nucleofector (Amaxa, US / Nucleofector, Electroporation Gene Transfer, Lonza) device of Lonza, somatic cells into which the differentiation inducer was introduced is 3 ⁇ in a culture dish. It is preferable to treat the peptide after stabilizing the medium every day for 5 days.
- a nucleofector Amaxa, US / Nucleofector, Electroporation Gene Transfer, Lonza
- the present invention can be used as a cell therapy by preparing a differentiated induced pluripotent stem cells under xenopathogen free or feeder cell free conditions.
- a Vitronectin Recombinant Human protein VTN-N
- VTN-N Vitronectin Recombinant Human protein
- Treatment of the functional peptides while culturing somatic cells into which the retrodifferentiation factor is introduced is carried out in a culture plate coated with Vitronectin protein, may also promote the efficiency of dedifferentiation into induced pluripotent stem cells.
- the peptide may be treated directly in a cell culture medium or mixed with a biomaterial for culture, and the biomaterial refers to a synthetic polymer or a natural polymer.
- the synthetic polymer is preferably poloxamer, polyethylene glycol and polypropylene glycol
- the natural polymer is preferably vitronectin, collagen, gelatin, alginic acid, chondroitin sulfate, fibronectin, and extracellular matrix protein. It is not limited.
- a natural polymer, vitronectin was used.
- Such a biomaterial is preferably used by coating a culture vessel, but is not limited thereto.
- the present invention provides a culture biomaterial that can replace the mouse embryonic fibroblasts (MEF), which is an animal-derived feeder cells used in the conventional iPSC culture (iPSC) By using it, it is possible to exclude xenopathogens generated by supplying various factors from MEF supporting cells to induced pluripotent stem cells. Therefore, the production of iPSCs using peptides of the present invention will improve both xenopathogen and efficiency of inducing differentiation to be considered when using undifferentiated pluripotent stem cells as cell therapy. Can be.
- MEF mouse embryonic fibroblasts
- iPSC animal-derived feeder cells used in the conventional iPSC culture
- the functional peptide may be characterized by inhibiting NF- ⁇ B activity by inhibiting nuclear transfer of NF- ⁇ B protein in somatic cells into which the differentiation factor is introduced and inhibiting NF- ⁇ B signaling mechanism.
- E MT epithelial-mesenchymal transition
- telomere reverse transcriptase hERT
- SSEA-4 human telomerase reverse transcriptase
- MET mesenchymal-epithelial transition
- the present invention can improve the efficiency of induction of dedifferentiation, and induced pluripotent stem cells prepared by this method are pluripotent stem cells in an undifferentiated state that normally display their characteristics as pluripotent stem cells. Therefore, it is very useful to effectively manufacture a pluripotent stem cell that can be clinically applied ultimately, and to develop a mass culture system that can secure pluripotent stem cell resources that can be clinically applied.
- P1 peptide (GKCSTRGRKCCRRKK: SEQ ID NO: 1) was synthesized from the C terminus by F-moc solid phase chemical synthesis using a synthesis apparatus. In other words, it was synthesized using Rink resin (0.075 mmol / g, 100-200 mesh, 1% DVB crosslinking) combined with Fmoc- (9-Fluorenylmethoxycarbonyl) as a blocking group, and 50 mg of Rink Amide in the synthesizer. After swelling the resin with DMF after adding MBHA resin, 20% piperidine / DMF solution was used to remove the Fmoc-group.
- 0.5M amino acid solution (solvent: DMF), 1.0M DIPEA (solvent: DMF & NMP), and 0.5M HBTU (solvent: DMF) were added in sequence from the terminal C to 1, 2 hours under nitrogen stream. Reacted for a while. Each time the deprotection and coupling steps were completed, the process was washed twice with DMF and methanol. After coupling the last amino acid (coupling) deprotection (deprotection) to remove the Fmoc-group.
- P1 peptide GKCSTRGRKCCRRKK (SEQ ID NO: 1)
- P2 peptide in which the C-terminal fifth cysteine of the P1 peptide (SEQ ID NO: 1) was substituted with methionine
- P3 peptide in which the C-terminal six cysteine of the P1 peptide (SEQ ID NO: 1) was substituted by the methionine No. 3 was synthesized by F-moc solid-phase chemical synthesis using a synthesizer.
- P2 Cectide GKCSTRGRKC M RRKK (SEQ ID NO: 2)
- P3 Cectide GKCSTRGRK M CRRKK (SEQ ID NO: 3)
- the dedifferentiation factors are prepared by dividing into three different episomal vectors according to a known method and expressing p53 shRNA together (pCXLE-hOCT3 / 4-shp53, pCXLE-hSK, pCXLE-hUL) to establish dedifferentiated stem cells. Increased efficiency (Okita K et al., Nat Methods. 8 (5): 409-12, 2011). Cells into which the differentiation factor was introduced were incubated in EMEM (Eagle's minimal essential medium, ATCC) medium, and the medium was replaced at 24 hour intervals until the cell confluency became 50%.
- EMEM Eagle's minimal essential medium, ATCC
- Naphthol / Fast Red Violet dye was added to the wells and stored at room temperature in a dark space for at least 20 minutes, followed by DPBS (Dulbecco's phosphate). The dye was washed with buffered saline) and the number of colonies stained under a microscope was checked.
- dedifferentiation factor was introduced into the cells by electroporataion into human dermal fibroblasts and dispensed into 6-well coated with Vitronectin. It was. Then, the expression level of Oct4, an embryonic stem cell marker, was compared by using immunofluorescence (IF) technique.
- IF immunofluorescence
- nucleus dye (Hoechst 33342, blue) was treated at room temperature for 10 minutes and washed with buffer solution (PBS) for confocal scanning microscope (IX 70, Olympus Co, Tokyo, Japan). ), The colonies with the clearest fluorescence in the wells were photographed.
- dedifferentiation factor was introduced into the cells by electroporataion into human dermal fibroblasts and dispensed into 6-well coated with Vitronectin. It was. After 10 days of peptide stabilized cells, cells were collected in a 15 ml Falcon tube using a scraper and centrifuged at 1500 rpm for 3 minutes. After washing with buffer (PBS), 1.5 ⁇ 10 5 cells are transferred to each tube using cold buffer (cold PBS).
- THY1 10 ⁇ g / ml concentration of THY1 (cat: sc-59398) and EPCAM (cat: ab20160) antibodies were diluted in 3% BSA / PBS solution and reacted at 4 ° C for 30 minutes, followed by 3 minutes at 1500 rpm for 3 minutes. Centrifuged twice. After washing with cold buffer (cold PBS), it was transferred to a round tube and analyzed by flow cytometry (FACS). Prior to measuring samples, THY1 used human dermal fibroblasts and EPCAM used human mammary epithelial cells (HMEC) to fix the control baseline.
- HMEC human mammary epithelial cells
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Biomedical Technology (AREA)
- Zoology (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Gastroenterology & Hepatology (AREA)
- Cell Biology (AREA)
- Developmental Biology & Embryology (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Toxicology (AREA)
- Transplantation (AREA)
- Rheumatology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Peptides Or Proteins (AREA)
Abstract
Description
Claims (17)
- 하기 일반식 I의 아미노산 서열로 표시되는 펩타이드를 유효성분으로 함유하는 분화된 세포에서 유도만능줄기세포로의 역분화 촉진용 조성물:일반식 IGKCSTRGRKX1X2RRKK상기 X1 및 X2는 시스테인(C: cysteine) 또는 메티오닌(M: methionine)이다.
- 제1항에 있어서, 상기 펩타이드는 서열번호 1 내지 3으로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는 유도만능줄기세포로의 역분화 촉진용 조성물.
- 제1항에 있어서, 상기 분화된 세포는 체세포 또는 전구세포인 것을 특징으로 하는 유도만능줄기세포로의 역분화 촉진용 조성물.
- 제1항에 있어서, 상기 분화된 세포는 인간 유래인 것을 특징으로 하는 유도만능줄기세포로의 역분화 촉진용 조성물.
- 다음 단계를 포함하는 분화된 세포로부터 역분화 유도만능줄기세포의 제조방법:(a) 분화된 세포에 역분화 유도인자를 도입하는 단계; 및(b) 역분화 유도인자가 도입된 세포에 하기 일반식 I의 아미노산 서열로 표시되는 펩타이드를 처리하여 배양하는 단계;일반식 IGKCSTRGRKX1X2RRKK상기 X1 및 X2는 시스테인(C: cysteine) 또는 메티오닌(M: methionine)이다.
- 제5항에 있어서, 상기 펩타이드는 서열번호 1 내지 3으로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 펩타이드는 0.01~100μM 농도인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 분화된 세포는 체세포 또는 전구세포인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 분화된 세포는 인간 유래인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 무-이종감염(xenopathogen free) 또는 무-지지세포(feeder cell free) 조건인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 역분화 유도인자는 Oct3/4, Sox2, c-Myc, Klf4 및 Lin28로 이루어진 군에서 선택된 어느 하나 이상인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 펩타이드는 NF-κB 활성을 억제하는 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 펩타이드는 중간엽세포-상피세포 전이(Mesenchymal-Epithelial Transition, MET)를 촉진하는 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제5항에 있어서, 상기 (b) 단계는 펩타이드를 배양 배지에 직접 처리하거나, 배양용 생체재료와 혼합하여 처리하는 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제14항에 있어서, 상기 생체재료는 합성고분자 또는 천연고분자인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제15항에 있어서, 상기 합성고분자는 폴록사머, 폴리에틸렌 글리콘 및 폴르프로필렌 글리콜로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
- 제15항에 있어서, 상기 천연고분자는 비트로넥틴, 콜라겐, 젤라틴, 알긴산, 황산 콘드로이틴, 피브로넥틴 및 세포외기질 단백질로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는 역분화 유도만능줄기세포의 제조방법.
Priority Applications (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US15/514,337 US10066212B2 (en) | 2015-08-06 | 2016-07-19 | Method for preparing induced pluripotent stem cells using synthetic peptide |
JP2017543696A JP6446566B2 (ja) | 2015-08-06 | 2016-07-19 | 合成ペプチドを利用した人工多能性幹細胞の製造方法 |
EP16833217.9A EP3345999B1 (en) | 2015-08-06 | 2016-07-19 | Method for producing induced pluripotent stem cells by using synthetic peptide |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR1020150111223A KR101906557B1 (ko) | 2015-08-06 | 2015-08-06 | 합성 펩타이드를 이용한 유도만능줄기세포의 제조방법 |
KR10-2015-0111223 | 2015-08-06 |
Publications (2)
Publication Number | Publication Date |
---|---|
WO2017022981A1 WO2017022981A1 (ko) | 2017-02-09 |
WO2017022981A9 true WO2017022981A9 (ko) | 2018-02-22 |
Family
ID=57943220
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/KR2016/007857 WO2017022981A1 (ko) | 2015-08-06 | 2016-07-19 | 합성 펩타이드를 이용한 유도만능줄기세포의 제조방법 |
Country Status (5)
Country | Link |
---|---|
US (1) | US10066212B2 (ko) |
EP (1) | EP3345999B1 (ko) |
JP (1) | JP6446566B2 (ko) |
KR (1) | KR101906557B1 (ko) |
WO (1) | WO2017022981A1 (ko) |
Families Citing this family (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR102631543B1 (ko) * | 2018-03-14 | 2024-02-01 | 서울대학교산학협력단 | 세포 투과능 및 근육 재생능을 가지는 이중 기능성 펩타이드 및 이의 용도 |
KR102631542B1 (ko) * | 2018-03-14 | 2024-02-01 | 서울대학교산학협력단 | 염증 감소능 및 줄기세포로부터 연골세포로의 분화 촉진능을 가지는 이중 기능성 펩타이드 및 이의 용도 |
WO2020046002A1 (ko) * | 2018-08-31 | 2020-03-05 | 주식회사 나이벡 | 다중의 질환 바이오마커의 기능 및 발현을 억제하는 펩타이드의 신규한 용도 |
KR102244161B1 (ko) * | 2018-08-31 | 2021-04-26 | 주식회사 나이벡 | 다중의 질환 바이오마커의 기능 및 발현을 억제하는 펩타이드의 신규한 용도 |
Family Cites Families (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2011046922A1 (en) * | 2009-10-13 | 2011-04-21 | Temple University Of The Commonwealth System Of Higher Education Office Of Technology Transfer | Use of nibp polypeptides |
WO2011105869A2 (ko) * | 2010-02-26 | 2011-09-01 | 주식회사 나이벡 | 치아의 접합능을 향상시키기 위한 치과용 클린저 조성물 |
KR101400668B1 (ko) * | 2011-04-22 | 2014-05-30 | 주식회사 나이벡 | 펩타이드에 의한 표면활성형 콜라겐 차폐막 |
GB2493540A (en) * | 2011-08-10 | 2013-02-13 | Follicum Ab | Agents for stimulating hair growth in mammals |
KR101456026B1 (ko) * | 2012-09-18 | 2014-11-04 | 서울대학교산학협력단 | 종양선택적 투과기능성을 가지는 펩타이드 및 그 용도 |
KR101529634B1 (ko) * | 2013-08-28 | 2015-06-30 | 서울대학교산학협력단 | 역분화 유도를 위한 세포투과성 융합 단백질 및 그 용도 |
-
2015
- 2015-08-06 KR KR1020150111223A patent/KR101906557B1/ko active IP Right Grant
-
2016
- 2016-07-19 EP EP16833217.9A patent/EP3345999B1/en active Active
- 2016-07-19 US US15/514,337 patent/US10066212B2/en active Active
- 2016-07-19 JP JP2017543696A patent/JP6446566B2/ja active Active
- 2016-07-19 WO PCT/KR2016/007857 patent/WO2017022981A1/ko active Application Filing
Also Published As
Publication number | Publication date |
---|---|
KR101906557B1 (ko) | 2018-10-11 |
JP2017533731A (ja) | 2017-11-16 |
US20170275594A1 (en) | 2017-09-28 |
US10066212B2 (en) | 2018-09-04 |
JP6446566B2 (ja) | 2018-12-26 |
WO2017022981A1 (ko) | 2017-02-09 |
EP3345999B1 (en) | 2023-10-18 |
KR20170017385A (ko) | 2017-02-15 |
EP3345999A1 (en) | 2018-07-11 |
EP3345999A4 (en) | 2019-01-16 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Demestre et al. | Formation and characterisation of neuromuscular junctions between hiPSC derived motoneurons and myotubes | |
WO2017022981A9 (ko) | 합성 펩타이드를 이용한 유도만능줄기세포의 제조방법 | |
KR20060002033A (ko) | 계통이 예정된 신경세포 전구체 | |
GB2414995A (en) | Neural stem cells | |
US20240191205A1 (en) | Efficient and non-genetically modified ipsc-induced, industrialized single clone selection platform, and use | |
US20190010455A1 (en) | Isolation And Use Of Pluripotent Stem Cell Population From Adult Neural Crest-Derived Tissues | |
WO2021086063A1 (ko) | 3차원 뇌 오가노이드를 이용한 뇌신경계질환 치료용 줄기세포 스크리닝 방법 | |
KR20010021499A (ko) | 계통이 예정된 신경세포 전구체 | |
MX2012002117A (es) | Eficiencia mejorada para la generacion de celulas madre pluripotentes inducidas a partir de las celulas somaticas humanas. | |
WO2012008733A2 (ko) | 1기 태반조직 유래 줄기세포 및 이를 함유하는 세포치료제 | |
WO2019198962A1 (ko) | 소변유래줄기세포로부터 제작된 유도만능줄기세포의 증식 및 분화 촉진 방법 | |
WO2016208879A1 (ko) | 인 비트로 섬유증 모델, 그의 제조방법 및 용도 | |
US20180371423A1 (en) | Non-viral ipscs inducing method, compositions, kits and ipscs | |
WO2013109763A2 (en) | Activation of innate immunity for nuclear reprogramming of somatic cells | |
WO2020045990A1 (ko) | 합성 메신저 rna를 이용하여 소변세포를 신경줄기세포로 직접 역분화하는 방법 | |
WO2011126177A1 (ko) | 인간 줄기세포의 활성을 증가시키는 방법 | |
WO2019231111A1 (ko) | 성체 줄기세포 배양액 및 그 제조방법 | |
Shutova et al. | Generation of iPS cells from human umbilical vein endothelial cells by lentiviral transduction and their differentiation to neuronal lineage | |
WO2013124309A1 (en) | Direct reprogramming of somatic cells into neural stem cells | |
Kaini et al. | Recombinant xeno-free vitronectin supports self-renewal and pluripotency in protein-induced pluripotent stem cells | |
CN107201341B (zh) | 一种毛囊干细胞的制备方法 | |
WO2022031095A1 (ko) | 췌장암 오가노이드의 제조 방법 | |
WO2022055239A1 (ko) | 기능성 신경교세포를 단기간에 효율적으로 분화시키는 방법 | |
WO2021241782A1 (ko) | 세포외기질 모티프를 포함하는 세포배양용 융합 단백질 및 이의 용도 | |
WO2019117454A1 (ko) | 세포 소기관 스트레스 조절 인자를 이용하는 고효율 세포전환용 배지 첨가제 |
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: 16833217 Country of ref document: EP Kind code of ref document: A1 |
|
WWE | Wipo information: entry into national phase |
Ref document number: 15514337 Country of ref document: US |
|
ENP | Entry into the national phase |
Ref document number: 2017543696 Country of ref document: JP Kind code of ref document: A |
|
REEP | Request for entry into the european phase |
Ref document number: 2016833217 Country of ref document: EP |
|
NENP | Non-entry into the national phase |
Ref country code: DE |